SYSTEMS AND METHODS TO DETECT INFECTIOUS DISEASES

专利摘要:
SYSTEMS AND METHODS FOR DETECTING INFECTIOUS DISEASES.Systems, methods, and biases for detecting the presence of markers indicative of one or more of a plurality of infectious agents without a single clinical sample, or in a plurality of aliquots from a single clinical sample, are provided. The systems, methods and devices described herein may be point-of-service systems, the methods and devices, configured for use at a point-of-service location, where a point-of-service location may be a location in the which a sample is obtained from a subject.
公开号:BR112016004994A2
申请号:R112016004994-2
申请日:2014-09-05
公开日:2021-05-11
发明作者:Pranav Patel；Elizabeth Holmes；Scott Tabakman；Kamila Belhocine；Aaron Richardson；Josephine Lee；Sharada Sivaraman；Sheena Menzes；Surekha Gangakhedkar；Clarissa Lui
申请人:Theranos Ip Company, Llc；
IPC主号:

专利说明:

[0001] [0001] Infectious diseases, or bacterial, viral, or otherwise, present acute and chronic challenges to health. Many common infections affect the respiratory tract. Respiratory tract diseases, especially infectious respiratory diseases of viral and bacterial origin, are prevalent in patients of all ages, although they are often more severe in the very young and the very old. Viruses include DNA viruses and RNA viruses. Bacteria include Gram negative and Gram positive bacteria, and may include mycoplasma (bacteria without cell walls). In addition to disease-causing bacteria, some diseases, such as, for example, respiratory diseases, can be caused by other microorganisms, such as yeast, fungi, and others, from small disease-causing organisms.
[0002] [0002] An example of a common viral origin of respiratory (and other) disorders in patients is the flu ("flu") virus. Influenza ("flu") refers to illness caused by one of several related RNA viruses in the family Orthomyxoviridae, typified by fever, headache, fatigue, and other symptoms. There are different types of flu; influenza A and influenza B are both almost equally prevalent in humans. Identifying the flu strain in a sample can help suggest treatments, can help suggest preventative measures to be taken, and can help control these infections in a population.
[0003] [0003] Examples of common bacterial causes of respiratory (and other) disorders in patients include whooping cough, pneumonia and tuberculosis. Pertussis is caused by Bordetella pertussis and is characterized by bouts of violent coughing, which can persist for weeks. Pneumonia is the name given to respiratory illnesses characterized by fluid in the lungs, coughing, fever, vomiting, fatigue, and other symptoms. Pneumonia can be caused by a bacterial or viral infection; Determining the cause of a specific case is critical in determining the patient's course of treatment. Pneumonia causes include Streptococcus pneumonia. Staphylococcus aureus, adenovirus, influenza virus, respiratory syncytial virus, Pneumocystis carinii (a fungus), and other agents. Tuberculosis is caused by the Mycobacterium, tuberculosis, is characterized by coughing including spitting blood, chest pain, chills, fever, night sweats and other symptoms, and can be fatal.
[0004] [0004] The agents that cause infectious respiratory diseases typically differ between upper respiratory tract diseases and lower respiratory tract disorders; Thus, the variety or range of viral or bacterial agents found in patients suffering from upper respiratory disorders may be different than those bacterial or viral agents found in patients suffering from lower respiratory tract disorders. However, the diagnosis and treatment of successful respiratory diseases often requires the identification of disease-causing organisms present in a clinical sample obtained from a subject suffering from, or suspected of suffering from, an infectious respiratory disorder. Differentiating between typical upper respiratory tract organisms and those typical of lower respiratory diseases can also be critical in the diagnosis and treatment of successful respiratory diseases, in addition, the identification of other symptoms and sequelae of respiratory disorders can aid the diagnosis successful and in the treatment of respiratory diseases.
[0005] [0005] Sexually transmitted diseases, whether they present specific viral or bacterial public health problems, or not, as some patients are reluctant to recognize the risks of, or possible exposure to such diseases, and may be reluctant to be tested for these diseases. However, the lack of testing and the consequent lack of information about the disease state can lead to an increased spread of such diseases, and treatment delays for those affected.
[0006] [0006] Some diseases can be detected by blood tests (eg, dengue virus, Epstein-Barr virus, trypanosome diseases, Plasmodium diseases, and others). Some diseases can be detected by analyzing swabs of, or fluid obtained from, swabs, such as throat swabs, nasal swabs, cheek swabs, or other swabs. Diseases can also be detected by analyzing urine and other clinical specimens.
[0007] [0007] In order to be effective in treating such infectious disorders, testing must be timely. However, current testing methods and systems are often time-consuming, inconvenient for patients, may require sampling methods or quantities that are painful or uncomfortable for the patient, and may be expensive. Methods that require large amounts of sample, or that require incubation of a sample for a day or days, are often ineffective in timely detection or identification of the cause of a respiratory disorder, and therefore may not be useful in diagnosis or treatment. of infectious diseases respiratory disorders.
[0008] [0008] In addition, many infectious respiratory diseases present many of the same or similar symptoms, so useful and effective testing requires testing for the presence of multiple agents, and multiple types of agents (eg, viral, bacterial and fungal). However, present methods are often limited to trials of a single agent or agent type, or only a small number of possible TS Agen, limiting the usefulness of the results and increasing the likelihood that the causal agent cannot yet be identified. .
[0009] [0009] Thus, improved methods, systems, and assays for detecting and identifying agents that cause disease, such as influenza, respiratory diseases, sexually transmitted diseases, blood diseases, viral diseases, bacterial diseases, and other diseases, are desired. INCORPORATION BY REFERENCE
[0010] [0010] All publications, patents, and patent applications mentioned in this specification are hereby incorporated by reference to the same extent as if each particular publication, patent, or patent application were specifically and individually indicated to be incorporated by reference, ABSTRACT
[0011] [0011] Systems, methods, and biases for detecting the presence of markers indicative of one or more of a plurality of infectious agents in a single clinical sample, or in a plurality of aliquots from a single clinical sample, are provided. The systems, methods and devices described herein may be point-of-service systems, the methods and devices, configured for use at a point-of-service location, where a point-of-service location may be a location in the which a sample is obtained from a subject.
[0012] [0012] In embodiments, applicants describe systems, methods and devices for assaying for the presence of one or more of a plurality of markers indicative of an infectious disease in a single clinical sample of small volume, or aliquots thereof. In embodiments, the system, method or device is a point of service (POS) system, method or device. In embodiments, the sample is collected at the POS site, and is analyzed in a device at the POS site. In embodiments, analysis of the small volume clinical sample is completed in a short period of time. In embodiments, the infectious disease comprises a respiratory disease. In embodiments, the infectious disease comprises a respiratory disease selected from an upper respiratory disease and a lower respiratory disease. In embodiments, the infectious disease comprises a sexually transmitted disease.
[0013] [0013] In embodiments, applicants describe systems, methods and devices for detecting the presence of one or more of a plurality of markers indicative of an infectious disease in a single clinical sample of small volume, or aliquots thereof. In embodiments, the system, method or device is a POS system, method or device. In embodiments, the sample is collected at the POS site, and is analyzed in a device at the POS site. Within embodiments, analysis of the small volume clinical sample is completed in a short period of time.
[0014] In embodiments, the infectious disease is a bacterial disease, or a viral disease, or other type of disease, and analysis of the small volume of the clinical sample determines whether the infectious disease is a bacterial disease, a viral disease, or another type of illness. Determining the type of infectious disease aids in determining the type of treatment to provide the subject, for example, where the determination indicates that the infectious disease is a fungal disease, the subject should be treated with antifungal drugs; where the determination indicates the infectious disease is a fungal infection, the matter should be treated with antifungal drugs; and so on.
[0015] [0015] In embodiments, the infectious disease is a bacterial disease. In embodiments, analysis of the small volume clinical sample determines whether the infectious disease is a bacterial disease. In embodiments where analysis of the small volume clinical sample determines that the infectious disease is a bacterial disease, said determination indicates the use of antibiotics in the treatment of that disease. In embodiments, the infectious disease is a viral disease. In embodiments, analysis of the small volume clinical sample determines whether the infectious disease is a viral disease. In embodiments where analysis of the small volume clinical sample determines that the infectious disease is a viral disease, said determination indicates the use of antiviral drag in the treatment of that disease. In embodiments where analysis of the small volume clinical sample determines that the infectious disease is a viral disease, said determination indicates that antibiotics should not be used in the treatment of that disease. In embodiments, the infectious disease is a disease bacterial, or a viral disease. In embodiments, analysis of the small volume clinical sample determines whether the infectious disease is a bacterial disease or a viral disease. Likewise, where small volume clinical sample analysis determines the infectious disease is a fungal disease, the individual should be treated with antifungal drags; where the determination indicates the infectious disease is a fungal infection, the matter should be treated with antifungal drugs; and so on.
[0016] [0016] In embodiments, the infectious disease comprises a respiratory disease. In embodiments, the infectious disease comprises a respiratory disease selected from an upper respiratory disease and a lower respiratory disease. In embodiments, analysis of the small volume clinical sample determines whether the infectious disease is an upper respiratory disease or a lower respiratory disease. In embodiments, analysis of the small volume clinical sample determines the type of upper respiratory mask'-disease or a lower respiratory disease present in the small volume clinical sample. For example, in embodiments, the upper or lower respiratory disease is a bacterial disease, or a viral disease, or other type of disease, and analysis of the small volume clinical sample determines whether the upper or lower respiratory disease is a disease. bacterial, a viral disease, or another type of disease. In embodiments where analysis of the small volume clinical sample determines that the upper or lower respiratory disease is a bacterial disease, said determination indicates the use of antibiotics in the treatment of said disease. clinical sample in small volume determines that the upper or lower respiratory disease is a viral disease, this determination indicates the use of antiviral drags in the treatment of this disease. In embodiments where analysis of the small volume clinical sample determines that the upper or lower respiratory disease is a viral disease, said determination indicates that antibiotics should not be used in the treatment of that disease. Likewise, where small volume clinical sample analysis determines the upper or lower respiratory disease is a fungal disease, the subject should be treated with antifungal drugs; where the determination indicates the infectious disease is a fungal infection, the matter should be treated with antifungal drugs; and so on.
[0017] [0017] In embodiments, the infectious disease comprises a sexually transmitted disease. In embodiments, analysis of the small-scale clinical sample determines the type of STD present in the small-volume clinical sample. For example, in embodiments, the STD is a bacterial disease, or a viral disease, or other type of disease, and small clinical sample volume analysis determines whether the STD is a bacterial disease, a disease viral, or other type of disease. In embodiments where small scale clinical sample analysis which determines the sexually transmitted disease is a bacterial disease, said determination indicates the use of antibiotics in the treatment of said disease. In embodiments where small-scale clinical sample analysis, which determines the sexually transmitted disease is a viral disease, said determination indicates the use of antiviral drugs in the treatment of that disease. In embodiments where small-scale clinical sample analysis, which determines the sexually transmitted disease is a viral disease, said determination indicates that antibiotics should not be used in the treatment of that disease. Likewise, where small volume clinical sample analysis determines the sexually transmitted disease is a fungal disease, the subject should be treated with antifungal drugs; where the determination indicates the infectious disease is a fungal infection, the matter should be treated with anti-yeast drags; and so on.
[0018] [0018] In embodiments of systems, methods and devices configured for testing a plurality of markers, and in systems, methods and devices for detecting a configured plurality of markers, the markers may be indicative of respiratory diseases; In embodiments, markers may be indicative of upper airway disease; In embodiments, markers may be indicative of lower respiratory tract diseases. In embodiments, applicants describe systems, methods and devices configured to test for a plurality of markers, wherein the respiratory disease markers are indicative of two or more of the group of respiratory disease markers consisting of adenovirus B, adenovirus C, E adenovirus, Bordetella pertussis, Mycobacterium tuberculosis (MTB), Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), group A streptococci and group B streptococcus. In embodiments, applicants describe systems, methods and devices configured to test for a plurality of markers, wherein the respiratory disease markers are indicative of two or more of the group of respiratory disease markers consisting of adenovirus B, adenovirus C , E adenovirus, Bordetella pertussis , Bordetella parapertussis, mycobacterium tuberculosis (MTB), Staphylococcus aureus, methicillin esistant Staphylococcus aureus (MRSA), group A streptococci, Group B streptococci, catarrhals Moraxella, Enterobacter aerogenes Streptococcus aureus, Haemfluus pneumoniae , Parainfluenza Virus 1, Parainfluenza virus 2, Parainfluenza virus 3, Coronavirus OC43, Coronavirus L63, Coronavirus MERS, Coronavirus HKUl, Coronavirus 229E, Klibsseila pneumonia PHOE, Klebsiella pneumonia KPC, Bocavirus type 2.4 and Bocavirus type 1.3. In embodiments, respiratory disease markers are indicative of three or more, or four or more, or five or more, or six or more, or SE Ven or more, or eight of that group of disease markers. respiratory.
[0019] [0019] In embodiments of systems, methods and devices configured to test for a plurality of markers, and in systems, methods and devices configured to detect a plurality of markers, the markers may be indicative of sexually transmitted diseases. In embodiments, applicants describe systems, methods and devices configured to test for a plurality of markers, wherein sexually transmitted disease markers of diseases are indicative of two or more of the group of markers consisting of herpes simplex virus (HSV) , human immunodeficiency virus (HIV), streptococcus B, and treponema pallidum. In embodiments, applicants describe systems, methods and devices configured to test a plurality of markers, wherein the sexually transmitted disease markers are indicative of two or more of the group consisting of HIV-2 markers. Group A, HIV-2, Group B, HIV-1 group M, hepatitis B, hepatitis Delta, herpes vims simplex (HSV), Streptococcus B, and Treponema pallidum. In embodiments, sexually transmitted disease markers are indicative of three or more, four or that group of sexually transmitted disease markers.
[0020] [0020] In embodiments of systems, methods and devices configured to test for a plurality of markers, and in systems, methods and devices configured to detect a plurality of markers, the markers may be indicative of influenza. In embodiments. Applicants describe systems, methods and devices configured for testing for a plurality of markers, wherein influenza markers are indicative of influenza A and influenza B. In embodiments, applicants describe systems, methods and devices configured for test for a plurality of markers, w here influenza markers are indicative of two or more of the group of markers consisting of the following forms of influenza: H1N1 (seasonal), H1N1 (new), H3N2, H7N9 (hemagglutinin marker gene ( HA) and the neuraminidase gene (nA)), and H5N 1. In embodiments, influenza markers are indicative of three or more, or four or more, or five of said group of influenza markers. In embodiments, the markers may be influenza matrix protein influenza markers, or may be influenza protein neuraminidase markers, or may be influenza hemagglutinin markers, or other influenza markers. In embodiments, analysis of the small volume clinical sample determines whether the infectious disease is an influenza virus. In embodiments, analysis of the small volume clinical sample determines the type of influenza present in the small volume clinical sample. In embodiments where analysis of the small volume clinical sample determines that the infectious disease is an influenza virus (which is a viral disease), said determination indicates that antibiotics should not be used in the treatment of that disease. In embodiments where analysis of the small volume clinical sample determines that the infectious disease is an influenza virus, said determination indicates that antiviral drugs should be used in the treatment of said disease.
[0021] [0021] In embodiments of systems, methods and devices configured to test a plurality of markers, and in systems, methods and devices configured to detect a plurality of markers, the markers may be indicative of diseases and markers of susceptible diseases. detected by analysis of a blood sample. In embodiments, such disease and disease markers that can be detected by analysis of a blood sample include West Nile virus, Epstein-Barr virus, Plasmodium, Trypanosoma cruzi, and Dengue virus (including types 1, 2, 3, and 4).
[0022] [0022] Samples from the throat of an object can be obtained, for example, by a throat rod; Samples obtained from the nose of an object can be obtained, for example, by a nasal swab. In embodiments, samples obtained from a subject's throat and nose can be tested together. In embodiments, testing samples obtained from the throat or nose, or from both the nose and throat, can be tested by nucleic acid analysis; or by amino acid analysis (for example, ELISA or other antibody-based or protein-based binding analysis); or by general chemical analysis; or by cytometric analysis; or by combinations thereof. For example, samples can be tested by nucleic acid analysis and by amino acid analysis. Such assays can be used to determine how long a subject has had an infection, for example, by looking at the delay in raising antibody levels indicative of a specific disease in the sample; or following the increase in antibody levels indicative of a specific disease in the specimen over time (eg, by repeated testing over time). Likewise, such assays can be used to detect, or to determine the effect of treatment by observing the delay in increasing levels of antibodies indicative of a specific disease in the sample; or following the increase in antibody levels indicative of a specific disease in the closest specimen (eg, by repeated testing over time). In embodiments, throat and nose samples can be included in a single solution, and tested together. In embodiments, the throat and nose samples may be in separate containers (e.g., sample containers), but both included in a single cartridge, and the separate vessels tested at the same time. This type of testing at the same time may comprise testing the pots separately, or may include mixing the contents of the containers and testing the mixture.
[0023] [0023] In embodiments, applicants describe systems, methods and devices configured to identify, or calculate, or otherwise determine the stage of an infection in an individual by detecting, or determining amounts of, or both , both nucleic acid markers indicative of a participating infection and antibody markers indicative of the same infection. Such systems, methods and devices can be used to detect, measure and track such markers over time, effective in providing an estimate or determination of how recently an infection has occurred. Such systems, methods and devices can be used to detect, measure and control such markers over time, effective to assist in assessing the current situation of an individual suffering from an infection. Such systems, methods and devices can be used to detect, measure and monitor such markers over time, effective to aid in determining the prognosis of the probability of an individual suffering from an infection. For example, where nucleic acid markers indicative of a particular infection are relatively numerous, while antibody or other protein markers indicative of the particular infection are relatively scarce, then one can estimate or determine that the infection is a recent infection; However, where nucleic acid markers indicative of a particular infection are relatively numerous, and antibody or other protein markers indicative of the particular infection are also relatively numerous, then one can estimate or determine that the infection is not. it is a recent infection, as the subject has had time to produce antibodies specific to the infection. Whenever nucleic acid markers indicative of a particular infection are relatively sparse, and antibody or other protein markers indicative of that particular infection are also relatively numerous, then one can estimate or determine that infection at a late stage , and indicates that the infection is decreasing, since such observations indicate that the matter is to overcome the infection.
[0024] [0024] In embodiments, applicants describe systems, methods and devices configured to test a plurality of markers, and disclose systems configured to detect a plurality of markers, where the markers are indicative of a plurality of infectious diseases in a single small volume clinical sample, or aliquots thereof. In embodiments, systems, methods, and devices can be configured to test for, or detect, markers indicative of more than about 8 different diseases, or more than about 8 different diseases, or more than about of 12 different diseases, or more than about 16 different diseases, or more than about 20 different diseases, or more than about 25 different diseases, or more than about 35 different diseases, or more than about 45 different diseases different, or more than about 60 different diseases. In embodiments, systems, methods and devices can be configured to test for, or to detect, a plurality of nucleic acid markers and protein markers, each marker being indicative of at least one of a plurality of diseases or conditions. In embodiments, the systems, methods, and devices can be configured to test for, or detect a plurality of nucleic acid markers, protein markers, and cytometry markers, each marker being indicative of at least one of a plurality of diseases or conditions. In embodiments, systems, methods, and devices can be configured to test for, or to detect, a plurality of nucleic acid markers, protein markers, cytometry markers, cytokines, and inflammation markers, each marker or cytokine being indicative of at least one of a plurality of diseases or conditions. Within embodiments, the systems, methods and devices comprise a system service point. In embodiments, the sample can be collected at the service site, and can be analyzed in a device at the POS site. In embodiments, small scale clinical sample analysis can be completed in a short period of time.
[0025] [0025] In embodiments, the infectious disease comprises a respiratory disease. In embodiments, the infectious disease comprises a respiratory disease selected from an upper respiratory disease and a lower respiratory disease. In embodiments, analysis of the small volume clinical sample determines whether the infectious disease is an upper airway disease, or a lower respiratory disease. In embodiments, analysis of the small volume clinical sample determines the type of upper respiratory disease or a lower respiratory disease present in the small volume clinical sample, in embodiments, the respiratory disease comprises a respiratory disease caused by a causative agent of disease selected from a virus, a bacterium, a yeast, a fungus, a mycoplasma, and other micro-organisms. In embodiments, small-scale clinical sample analysis determines the type of upper respiratory disease or a lower respiratory disease present in the small volume clinical sample. In embodiments, analysis of the small volume clinical sample determines whether the upper or lower respiratory disease of a respiratory disease is a viral disease, or a bacterial disease, or some other type of disease. In embodiments where analysis of the small volume clinical sample determines that the disease is a viral disease, such determination indicates that antibiotics should not be used in the treatment of that disease. In embodiments where analysis of the small volume clinical sample determines that the disease is a viral disease, said determination indicates that antiviral drugs should be used in the treatment of said disease. In embodiments where analysis of the small clinical sample volume and determines that the disease is a bacterial disease, said determination indicates that antibiotics should be used in the treatment of said disease.
[0026] [0026] In embodiments, the infectious disease comprises a sexually transmitted disease. In embodiments, the sexually transmitted disease comprises a sexually transmitted disease caused by a disease-causing agent selected from a virus, a bacterium, a yeast, a fungus, a mycoplasma, and other microorganisms. In embodiments, analysis of the small-scale clinical sample determines the type of STD present in the small-volume clinical sample. In embodiments, analysis of the small volume clinical sample determines whether the sexually transmitted disease is a viral disease, or a bacterial disease, or some other type of disease. In embodiments where analysis of the small volume clinical sample determines that the disease is a viral disease, such determination indicates that antibiotics should not be used in the treatment of that disease. In embodiments where analysis of the small volume clinical sample determines that the disease is a viral disease, said determination indicates that antiviral trails should be used in treating said disease. In embodiments where analysis of the small volume clinical sample determines that the disease is a bacterial disease, said determination indicates that antibiotics should be used in the treatment of that disease.
[0027] [0027] Applicant further describes a method of determining the response status to a disease in a subject, the method comprising: a) introducing a clinical sample to a sample processing device, said sample having been obtained from an individual suspected of suffering from a disease caused by a disease-causing organism, said clinical sample having a volume not greater than 500 microliters, wherein the device comprises: i) a sample handling system; lo) a detection station; and iii) a test station comprising at least a first and a second independently detachable test unit; b) with the help of the sample handling system, transfer a portion of the clinical sample to each of the first and second test units, in which an assay for the detection of a nucleic acid indicative of the disease-causing organism is performed in said first test unit, and an assay for the detection of antibodies to the disease-causing organism is performed in the second test unit; c) transferring the first and second test units to the detection station with the help of the sample handling system; d) obtaining data measurements with the aid of the detection station, data measurements comprising determining the level of nucleic acid indicative of a disease-causing organism in the sample, and determining the level of antibodies directed at that organism of disease in the referred sample; ee) ai) to determine that the infection is a recent infection, and at an early stage of the disease, where the level of nucleic acid indicative of a disease-causing organism is high and the level of antibodies directed to that disease-causing organism is it low or normal; ii) the determination that the infection is a non-recent rather than an early stage of the disease, where the level of nucleic acid indicative of a disease-causing organism is high and the level of antibodies directed to that disease-causing organism is tall; and iii) the determination that the infection is a decreasing infection, and at a late stage of the disease, where the level of nucleic acid indicative of a disease-causing organism is low or normal and the level of antibodies directed to that disease-causing organism. disease is high, where a normal level of a marker is the level of that marker determined from a healthy population of normal individuals, where a high level is one that significantly exceeds a normal level, as determined in a healthy population, and a level low is one that is at or below the normal level, as determined in a healthy population.
[0028] In embodiments, the method of determining the response status to a disease in a subject further comprises detecting the level of inflammatory cytokines. In embodiments of the method of determining the state of response to a disease in a subject, the device further comprises a cytometry station comprising an imaging device and a stage for receiving a microscope cuvette, and the method further comprises imaging a white blood cell in a blood sample obtained from the subject. In embodiments, imaging white blood cells in a blood sample obtained from the subject comprises detecting the level of a type of white blood cell in a blood sample obtained from the subject and determining whether said detected level of said type of blood. white blood cell is above, in, or below a normal level for that type of white blood cell, wherein the normal level for that type of white blood cell is determined by the level of that type of white blood cell, in blood samples from a healthy population.
[0029] [0029] 'T ESTs can be for the detection of markers indicative of any infectious disease. For example, illnesses that can be tested for include respiratory illnesses, and include respiratory illnesses and lower respiratory illnesses. Markers can include nucleic acid markers, protein markers, polysaccharide markers, cellular markers (including cells and cell organelles or fragments), and other markers. Markers can include markers for viral infections, bacterial infections, fungal infections, fungal infections, mycoplasma infections and for other infections. Samples can be tested for markers indicative of inflammation. Samples can be tested for cytokines. Samples can be tested for inflammatory cytokines. Samples can be tested for anti-inflammatory tokines cy. The amount of dilution of a sample, or the detection level of a marker, can be determined by a subject's past condition or history.
[0030] [0030] Test results can be obtained within three hours, or two hours, or an hour or half an hour, or Jess from the time a sample is placed in a testing device for analysis. A sample can be placed in a testing device for analysis within five hours, or four hours, or three hours, or two hours, or an hour or half an hour, or less than the time a sample was obtained from a subject. Test results can be obtained within eight hours, seven hours, or six hours or five hours, or three hours, or two hours, or an hour or half an hour, or less than the time a sample was obtained from a subject. .
[0031] [0031] In embodiments, the applicant describes a method of Ting detecting a disease marker, comprising: a) introducing a cartridge comprising one or more samples to an automatic sample processing device, said cartridge being configured to containing at least one sample and being configured to perform a swab, wherein said automatic sample processing device comprises: i) a sample handling SY rod configured to transport at least a portion of a sample and being configured to transport a mobile unit trial independently; and ii) an optical detector; b) contacting a sample, or a portion thereof, with a mobile test unit, or a reagent, or both, for carrying out an assay for the detection of a disease marker, said contact comprising transport, with aiding said sample handling system, at least a portion of said sample, or a mobile testing unit, or a reagent, or combinations thereof; c) positioning said sample, or part thereof, in a suitable location for detecting an optical signal from the sample, or part thereof by said optical detector; and d) detect the presence of a disease marker. In embodiments, such a method may comprise performing two or more assays for detecting disease markers, and detecting two or more disease markers in said one or more samples, or in one or more portions. of the same. In embodiments, the sample has a volume of less than about 500 microliters (JJL). In embodiments, the one or more samples comprises a blood sample; or comprises a sample obtained using a swab; or comprises both a blood sample and a sample obtained using a cotton swab. In embodiments, a sample obtained using said swab can be obtained by scraping a mouth, throat, nasal passage, vaginal area, or other body cavity of a subject. In embodiments, the method comprises detecting the presence of a nucleic acid disease marker and a disease marker protein.
[0032] The methods described herein include performing two or more assays for detecting disease markers, and detecting two or more disease markers in said samples, or in one or more portions thereof. In embodiments, the methods comprise detecting the presence of a nucleic acid disease marker and a disease marker protein. In embodiments, the disease marker is selected from a disease nucleic acid marker, a protein disease marker, a saccharide, a prostaglandin, a cytokine, histamine, a steroid, and an inflammation marker. In embodiments, two or more disease markers are detected, wherein the disease markers are selected from a disease nucleic acid marker, a disease marker protein, a saccharide, a prostaglandin, a cytokine, histamine, a steroid, and a marker of inflammation. In embodiments, a marker is a marker for inflammation selected from prostaglandins, tumor necrosis factor alpha (TNF-α), inierleukin-1 (IL-1), interleukin-8 (IL-8), interleukin- 12 (IL-12), interferon gamma (IF-γ), bradykinin, complement system molecules, blood clotting factors, C-reactive protein, erythrocyte sedimentation rate (ESR), white blood cell count , and morphological changes in blood and other cells.
[0033] [0033] In embodiments, a marker is a marker for a disease-causing agent, wherein said disease-causing agent is selected from the group of disease-causing organisms consisting of a virus, a bacterium, a mycoplasma , a fungus, a yeast and other microorganisms. In embodiments, a disease marker for a disease-causing agent is selected from the group consisting of matrix protein Influenza A, Influenza H3N2, Influenza ΗΊΝ 1 seasonal, Influenza ΗΤΝΊ novel, Influenza B, Streptococcus pyogenes (A), Mycobacterium Tuberculosis, Staphylococcus aureus (MR), Staphylococcus aureus (RS), Bordetella pertussis (whooping cough), Streptococcus agalactiae (B), Influenza H5M 1, Influenza virus H7N9, Adenovirus B, adenovirus C, adenovirus E, hepatitis b. Hepatitis C, Hepatitis Delta, Treponema pallidum, HSV-1, HSV-2, HIV-1, HIV-2, Dengue I, Dengue 2, Dengue 3, Dengue 4, Malaria, West Nile Virus, Trypanosoma cruzi (Chagas), Klebsiella pneumoniae (Enterobacteriaceae spp), Klebsiella pneumoniae carbapenemase (CPK), Epstein Ban Vims virus (mono), rhinovirus, Parainfluenza virus (1), Parainfluenza virus (2), parainfluenza vims (3), Parainfluenza virus (4a), vims parainfluenza (4b), respiratory syncytial virus (RSV) A, respiratory syncytial VIMS (RSV) B, Coronavirus 229E, Coronavirus HKUl, Coronavirus OC43, Coronavirus NL63, new coronavirus, bocavirus, human metapneumovirus (MPVH), Streptococcus pneumoniae (Penic R) , Streptococcus pneumoniae (S), Mycoplasma pneumoniae, Chlamydia pneumoniae, Bordetella parpertussis, Haemophilus influenzae (Broad R), Haemophilus influenzae (broad S), Moraxella catarrhalis, Pseudomonas spp
[0034] In embodiments, the methods comprise detecting a disease marker in a blood sample and detecting a disease marker in a sample obtained from a swab, which one of said disease markers is a marker for inflammation, and one of said disease markers of a marker of a disease-causing agent. In embodiments, such a marker for inflammatory disease is selected from prostaglandins, tumor necrosis factor alpha (TNF-a), inte feukin~l (IL-1), interleukin-8 (11.-S;. Interleukin-12 (IL-l2), interferon gamma (IF-y), bradykinin, complement system molecules, blood clotting factors, C-reactive protein, erythrocyte sedimentation rate (ESR), white cell count of the blood, and morphological changes in blood and other cells and such a marker for disease a disease-causing agent is selected from the group of disease-causing organisms consisting of a virus, a bacterium, a mycoplasni, a fungus, a yeast, and other microorganisms.
[0035] [0035] In embodiments, a marker is a marker of a disease selected from influenza, a respiratory disease, a sexually transmitted disease, and another infectious disease. In embodiments where the disease is influenza, the disease marker is selected from ΗΓΝ1 (seasonal), ΗΓΝ1 (new), H3N2, H7M9, and H5N1. In embodiments, the disease is respiratory disease selected from an upper respiratory disease and a lower respiratory disease. In embodiments where the disease is a respiratory disease, the marker may be a marker of a disease-causing organism selected from the group consisting of adenovirus B, adenovirus C, adenovirus E, Bordetella pertussis, Mycobacterium tuberculosis ( MTB), Staphylococcus aureus, Methicillm-Resistant Staphylococcus aureus (MRSA), group A streptococci, Group B streptococci, Moraxella catarrhalis, Enterobacter aerogenes, Haemophilus parainfluenza, Metapneumo Virus. Streptococcus pneumonia, Parainfluenza virus 1, Parainfluenza virus 2, Parainfluenza virus 3, coronavirus OC43, coronavirus NL63, Coronavirus MERS, Coronavirus HKU1, Coronavirus 229E, Klibsieila pneumonia PHOE, Klebsiella pneumonia KPC, Bocavirus type 2.4 and Bocavirus Type 1,3 .
[0036] [0036] In embodiments where the disease is a sexually transmitted disease, the marker may comprise a marker for a disease-causing organism indicative of a sexually transmitted disease selected from herpes simplex virus (HSV), human immunodeficiency virus (HIV) , HIV-2, Group A, HIV-2 Group B, HIV-1 group M, hepatitis B, hepatitis Delta, herpes simplex virus (HSV), Streptococcus B, and Treponema pallidum.
[0037] In embodiments, the method comprises detecting a disease marker in a blood sample and detecting a disease marker in a sample obtained from a swab, wherein at least one of said disease markers is a marker for a disease-causing organism indicative of a respiratory disease selected from the group consisting of adenovirus B, adenovirus C, adenovirus E, Bordetelia pertussis, Mycobacterium tuberculosis (MTB), methicillin-resistant Staphylococcus aureus, Staphylococcus aureus (MRSA) , group A streptococci, Group B streptococci, Moraxella caiarrhaiis, aerogenes Enierobacter, Haemophilus parainfluenza, Metapneumo Virus, Streptococcus pneumonia, Parainfluenza Virus 1, Parainfluenza virus 2, Parainfluenza Virus 3, Coronavirus HKUERS, 63, Coronavirus coronavirus NL43, Coronavirus Coronavirus NL43 229E, Klibsieila pneumonia PHOE, Klebsiella pneumonia KPC, Bocavirus type 2.4 and Boeavirus type 1.3.
[0038] [0038] In embodiments where the disease is an infectious disease, the disease marker may comprise a marker for an infectious disease causing agent selected from the group consisting of West Nile virus, Epstein-Barr virus , Plasmodium, Trypanosoma cruzi, and a Dengue Virus.
[0039] In embodiments comprising detecting a disease marker in a blood sample and detecting a disease marker in a sample obtained from a swab, wherein at least one of said disease markers is a marker for a disease-causing organism indicative of a sexually transmitted disease selected from herpes simplex virus (HSV), human immunodeficiency virus (HIV), Group o HIV-2 A, HIV-2 Group B, HIV-1 group M, hepatitis B, hepatitis Delta, herpes simplex virus (HSV), Streptococcus B, and treponema pallidum.
[0040] [0040] In embodiments comprising detecting a disease marker in a blood sample and detecting a disease marker in a sample obtained from a swab, wherein at least one of said disease markers is a marker for a disease-causing agent selected from the group consisting of West Nile Vims, Epstein-Barr virus, Plasmodium, Trypanosoma cruzi, and a Vims Dengue.
[0041] [0041] In TH and method embodiments of detecting a disease marker, the method is a point-of-service method performed at a point-of-service location. In embodiments of methods of detecting a disease marker, the method can be performed in less than about 40 minutes. In embodiments comprising detecting a disease marker in a blood sample and detecting a disease marker in a sample obtained from a stem, the method is a point-of-service method performed at a point-of-service location. service. In embodiments comprising detecting a disease marker in a blood sample and detecting a disease marker in a sample obtained from a stem, the method can be performed in less than about 40 minutes,
[0042] [0042] For example. Applicant here describes a method for determining the stage of an infection in an individual suffering from an infection, comprising: Testing at least one sample, or an aliquot or aliquots thereof, obtained from said subject 1) for the presence of a nucleic acids indicative of the infection, and 2) for the presence of an antibody indicative of infection, and determining whether the relative amounts of a) nucleic acids indicative of infection and b) antibodies indicative of infection indicate that the intection is a recent, characterized infection. i) higher relative amounts of nucleic acids indicative of infection compared to the relative amount of antibodies indicative of infection indicate that the infection is a recent infection, and ii) a significant amount of antibodies against the infectious agent indicate the infection is not a recent infection. In embodiments of such a method, the sample from at least one comprises a blood sample. In embodiments of such a method, the sample of at least one compound is a sample selected from a pharyngeal sample, a buccal swab sample, a nasal swab sample, a saliva sample, and a blood sample. In embodiments of such a process, where a significant amount of antibody against the infectious agent is detected, and where nucleic acid markers indicative of the infectious agent are relatively sparse, then the method indicates that the infection is a stage late and that the infection is decreasing.
[0043] [0043] In embodiments, such a method may further include testing a sample or samples for an inflammation marker. In embodiments, the inflammation marker may be selected from a prostaglandin, tumor necrosis factor alpha (TNF-a), interleukm-1 (IL-1), interleukin-8 (IL-8), interleukin-12 (IL-12), interferon gamma (TF-γ), bradykinin, a complement system molecule, a blood clotting factor, and a morphological change in blood or other cells. In embodiments, the inflammation marker is a cytokine selected from a lympbokine,
[0044] [0044] In embodiments, a process described herein comprises testing to determine whether the individual is suffering from a bacterial infection, a viral infection, a fungal infection, a mycoplasma infection, a fungal infection, infection by another, or the combination of them. In embodiments, such assays comprise determining whether markers indicative of viral infection or markers indicative of bacterial infection are detected, effective to determine whether the individual is suffering from a bacterial infection or a viral infection.
[0045] [0045] In embodiments of the methods described herein, the methods further comprise the prescription of a suitable prescription for the treatment of the infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises prescribing an antibiotic when the assay determines that the subject is suffering from a bacterial infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises prescribing an anti-mycoplasmic drug when the assay determines that the subject is suffering from a mycoplasma infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises prescribing an anti-viral scavenger when the assay determines that the subject is suffering from a viral infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises avoiding prescribing an antibiotic, wherein the test determines that the subject is suffering from a viral infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises avoiding the prescription of an antibiotic, and providing the prescription of an antiviral drug, when the assay determines that the subject is suffering from a viral infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises prescribing an antifungal drug when the test determines that the subject is suffering from a fungal infection. In embodiments, the methods disclosed herein comprise providing a suitable prescription for the treatment of said infection comprises prescribing an anti-yeast drug when the test determines that the subject is suffering from a yeast infection.
[0046] [0046] In embodiments, the methods comprise detecting a disease, wherein the detected disease is caused by a disease-causing agent selected from the group of disease-causing organisms consisting of a virus, a bacterium, a mycoplasma, a fungus, a yeast, and other microorganisms, and further comprising providing a prescription for the proper treatment of said virus, bacteria, mycoplasma, fungi, yeast, or other microorganism.
[0047] [0047] In embodiments, the methods are point-of-service methods performed at a point-of-service location. In embodiments, the methods comprise performing a plurality of assays on a single, small volume clinical sample, or aliquots thereof, and can be performed in less than about 40 minutes. In embodiments, the infection is caused by a disease selected from influenza, a respiratory disease, a sexually transmitted disease, and another infectious disease. In embodiments where the infection comprises influenza, influenza may be selected from H1N1 (seasonal), H1N1 (new), H3N2, H7N9 and H5N 1. In embodiments where the infection comprises a respiratory disease, the infection can be selected from an upper respiratory disease and lower respiratory disease. In embodiments, the respiratory disease is selected from the group consisting of adenovirus B, adenovirus C, adenovirus E, Bordetella pertussis, Mycobacterium tuberculosis (MTB), Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), Streptococcus from the group A, group B streptococci, Moraxella catarrhal] s, Enterobacter aerogenes, Haemophilus parainfluenza, Metapneumo Vims, Streptococcus pneumonia, Parainfluenza Virus 1, Parainfluenza Vims 2, Parainfluenza Vims 3, Coronavirus OC43, Coronaviruses 2NL63 , Klibsieila pneumonia PHOE, Klebsiella pneumonia KPC, Bocavirus type 2.4 and Bocavirus type 1, 3.
[0048] In embodiments, the infection comprises a sexually transmitted disease selected from a disease caused by herpes vims simplex (HSV), human immunodeficiency virus (HIV), Group o HIV-2a, HIV-2, of the group B, HIV-1 group M, hepatitis B, hepatitis Delta, heipes vims simplex (HSV), Streptococcus B, and Treponema pallidum. In embodiments, the infection comprises a disease caused by a disease-causing infectious agent selected from the group consisting of matrix protein Influenza A, influenza H3N2, seasonal influenza HlNl, novel influenza H LNL, influenza B, Streptococcus pyogenes (A ), Mycobacterium Tuberculosis, Staphylococcus aureus (MR), Staphylococcus aureus (RS), Bordetella pertussis (whooping cough), Streptococcus agalactiae (B), influenza H5N1, H7N9 influenza, Adenovirus B, adenovirus C, adenovirus E, hepatitis b. Hepatitis C, Hepatitis Delta, Treponema pallidum, HSV 1, HSV-2, HIV-1, HIV-2, Dengue 1, Dengue 2, Dengue 3, Dengue 4, Malaria, West Nile Virus, Trypanosoma cruzi (Chagas), Klebsiella pneumoniae (Enterobacteriaeeae spp), Klebsiella pneumoniae carbapenemase (CPK), Epstein Barr virus (mono), rhinovirus, vims parainfluenza (1), parainfluenza virus (2), parainfluenza vims (3), vims parainfluenza (4a), parainfluenza virus ( 4b ), Respiratory VIMS syncytial (RSV) A VIMS syncytial respirator (RSV) B, Coronavirus 229E, Coronavirus HKU1, Coronavirus OC43, Coronavirus NL63, Novel Coronaviras, Bocavirus, human metapneurnovirus (MPVH), Streptococcus pneumoniae (Penic R), Streptococcus pneumoniae (S), Mycoplasma pneumoniae. Chlamydia pneumoniae, Bordetella parpertussis, Haemophilus influenzae (ampic R), Haemophilus influenzae (ampic S), Moraxella catarrhalis, Pseudomonas spp (aeruginosa), Haemophilus parainfluenza, Enterobacter cloacae (Enterobacteriaeeae spp), Enterobacteraebacteriae aerogenes (Enterobactereeae sppae) spp),
[0049] [0049] In embodiments, the infection comprises a bacterial infection caused by bacteria selected from the group consisting of Bordetella pertussis, Mycobacterium tuberculosis (MTB), Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), group A streptococci , Group B streptococci, Moraxelia catarrhalis, Enterobacter aerogenes, Haemophilus parainfluenza, Streptococcus pneumonia, Klibsiella PHOE pneumonia, Klebsiella KPC pneumonia and treponema pallidum.
[0050] In embodiments, the infection comprises a viral infection caused by a virus selected from the group consisting of an influenza virus, herpes simplex virus (HSV), human immunodeficiency virus (HIV), Group HIV -2 A, HIV-2 Group B, HIV-1 Group H, hepatitis B, hepatitis Delta, herpes simplex virus (HSV), West Nile Vims, Epstein-Barr virus, dengue virus, adenovirus B, adenovirus C , adenovirus E virus, Metapneumo virus, Parainfluenza virus 1, Parainfluenza virus 2, Parainfluenza Virus 3, Coronavirus OC 3, coronavirus NL63, Coronavirus MERS, Coronavirus HKU1, Coronavirus 229E, Bocavirus type 2.4 and Bocavirus type 1.3. In embodiments of influenza infections, influenza can be selected from HIN I (seasonal), H1N1 (new), H3N2, H7N9, and HSNL influenza viruses. In embodiments, a viral infection comprises infection by a Dengue virus, wherein said dengue virus is selected from dengue virus type 1, Dengue virus type 2, Dengue virus type 3, and Dengue VIMS type 4.
[0051] [0051] A bill for testing can be automatically generated in the POS. The invoice amount can be calculated by tests performed, or in accordance with the test results. A bill for the test may be automatically sent to the subject's insurance provider. Payment for the test may be obtained automatically from the subject, or from the subject's insurance carrier, or fro another source.
[0052] [0052] A prescription for the treatment of a detected disorder can be provided at the POS site. A prescription for the treatment of a detected disorder can be filled at the POS site. A bill for the filled prescription can be generated automatically. A bill for prescription can be automatically sent to the subject's insurance provider. Payment for the completed prescription may be obtained automatically from the subject, or from the subject's insurance provider, or from another source.
[0053] [0053] Therefore, Applicants provide systems, methods and devices for the rapid analysis of a small volume clinical sample in a short period of time. Such rapid analysis includes testing for the presence of markers indicative of a plurality of disease-causing agents in a short period of time. In embodiments, such disease-causing agents include agents that cause upper respiratory disease, and include agents that cause minor respiratory-disorders. In embodiments, such systems, methods and devices are configured to detect one or more indicators of ignition. In embodiments, such systems, methods and devices are configured to detect one or more cytokines. In embodiments, such systems, methods and devices are configured to detect one or more inflammatory cytokines. In embodiments, such systems, methods and devices are configured to detect one or more anti-inflammatory cytokines.
[0054] In embodiments, Applicants provide systems, methods and devices for detecting a plurality of pathogens in a single clinical sample, or in a plurality of aliquots from a single clinical sample. In embodiments, a single clinical sample may be a small clinical sample volume of blood, saliva, tears, nasal swabs, throat swabs, mouth swabs (e.g., cheek swabs), vaginal discharge, or other bodily fluid, the tissue, the secretion or excretion made from a subject. In embodiments, a single clinical sample has a volume of less than about 500 µl, or less than about 250 µl., or less than 150 µl, or less than about 100 or less than about 50 µl, or less than about 25 µl or less than about 10 µl, or less than about 5 µl. ,, or less than about 1 μ3_ ,, or less.
[0055] [0055] In embodiments, clinical samples may be obtained at a point-of-service location (POS). The POS location can be, for example, a pharmacy, supermarket, hospital, clinic, doctor's office, or other location. Clinical samples can be tested at the POS site for various markers indicative of agents that can cause one or more of a plurality of diseases (e.g., at least eight, or at least 10, or at least 12, or at least 20, or at least 30, or at least 40, or at least 50, or at least 60, or more markers, indicative of the same or similar numbers of different diseases). The test can be completed in a short period of time. In embodiments, the short period of time can be measured from when the sample is inserted into a device or system to perform an analysis. In embodiments, the short period of time can be measured from when the sample is obtained from the subject.
[0056] In embodiments, clinical samples can be analyzed at a POS location. In embodiments, clinical samples obtained at a POS location can be analyzed at the same POS location. In embodiments, clinical samples may be obtained at a point-of-service (POS) location and may be analyzed at a different location. In embodiments, clinical samples perhaps analyzed in a short period of time, for example, in a period of time that is less than about 5 hours, or less than about 4 hours, or less than about 3 hours, or less of about 2 hours, or less than about 1 hour, or less than about half an hour.
[0057] [0057] In embodiments, Applicants provide devices (e.g., cartridges) for use in performing assays for detecting a plurality of pathogens in a single clinical sample, or in a plurality of aliquots from a single clinical sample.
[0058] [0058] In embodiments, devices for use in performing assays for detecting a plurality of disease-causing agents as described herein may further include a space, or vessel, for holding a rod; or it may further include a space, or a vessel, for holding two pads, or a plurality of rods. In embodiments, such devices may further include two spaces, or two vessels, for holding two pads; or it can include a plurality of spaces, or ships, for realizing a plurality of rods. In embodiments, a single roving can be placed in a single space, or container; in embodiments, two pads can be piaced into a single space, or vessel; and in embodiments, a plurality of swab samples may be placed in a single space, or vessel. Thus, in embodiments, a swab can be placed in a container, and, in embodiments, a swab, or a plurality of rods, can be placed in a single vessel. In embodiments, a plurality of smear samples may be piaced in a plurality of vessels. A vessel such for holding a swab or containers for holding swabs may contain a reagent, or a diluent, or other solution for use with a sample or swabs. In embodiments, a container for holding a swab can be used to provide a clean swab for use in obtaining a sample. In embodiments, a container for holding a swab can be used to i) provide a clean swab for use in obtaining a sample and also to ii) receive the swab after its use in obtaining a sample from an individual. . In embodiments, a container for holding a plurality of swab samples can be used to provide a plurality of cleaning rods for use in obtaining a sample. In embodiments, a container for holding a plurality of swab samples can be used to i) provide a plurality of cleaning rods for use in obtaining a sample and also to ii) receive one or more of the plurality of swabs after the its use in obtaining a sample from a subject.
[0059] [0059] For example, a throat rod and a nasal swab can be obtained from a subject. A nasal swab can be useful for testing for upper respiratory ailments, and a throat rod can be useful for testing for lower respiratory ailments. In embodiments, the IHE throat swab can be placed in a container on one device (e.g., a cartridge), and the nasal swab can be placed on a different container on the device. These containers may contain a reagent, or a diluent, or other solution for use with the swabs; such reagents may be different for throat swabs and nasal cotton swabs. In embodiments, the throat and nasal stem can be placed in the same vessel in one device. The container may contain a reagent, or a diluent, or other solution for use with these smears. The device can be placed in an analysis device, or within an analysis system for analysis. Such analysis devices and analysis systems can be placed in the same place as the one where the sample was obtained; or such analysis devices and analysis systems may be in a different location or locations than the location where the sample was taken.
[0060] [0060] In embodiments, a device may be or comprise a cartridge configured to contain a reaction vessel or a plurality of reagent vessels. In embodiments, a device can be or comprise a cartridge configured to contain a reaction vessel or a plurality of reaction vessels. In embodiments, a device can be a cartridge configured to contain a cytometry cuvette, or a plurality of cytometry cuvettes. In embodiments, a device can be or comprise a cartridge configured to contain a waste container, or a plurality of garbage containers. In embodiments, a device can be or comprise a cartridge configured to contain a sample; In embodiments, a sample can be contained in a sampling device. In embodiments, a device can be or comprise a cartridge configured to contain a sample collection container.
[0062] [0062] Accordingly, applicants describe systems for detecting the presence of one or more of a plurality of markers indicative of an infectious disease, from a small volume clinical sample, comprising: a) a sample handling system;
[0063] [0063] Accordingly, applicants describe systems for detecting the presence of one or more of a plurality of markers indicative of an infectious disease of a small volume clinical sample, comprising: a) a sample handling system; b) a detection station comprises an optical sensor; c) a fluidly isolated sample collection unit configured to hold a clinical sample; d) a test station comprising at least a first, second, and third fluidly isolated test unit, wherein the first unit comprises a first reagent, the second unit comprises a second reagent, and the third unit comprises a third reagent ; and e) a controller, wherein the controller comprises a local memory and is operative}' coupled to the sample handling system and the detection station; where the system is configured to run the tests with any one or more of the first, second and third test units: where the controller's local memory comprises a protocol comprising instructions for: i) the direction of the sample handling system to transfer a portion of the clinical sample to the first test unit, the second test unit and the third test unit; and ii) direct the sample handling system to transfer the first test unit, the second test unit, and the third test unit to the detection station. In embodiments, rod SY can include only two test units; or it can include four test units; or it can include more than four test units.
[0064] [0064] In embodiments, the system is a point-of-service system. In embodiments, the system is contained within a housing. In embodiments, the system is located at a point-of-service location, and is configured to be used in analyzing a sample at said point-of-service location. In embodiments, the system is a point-of-service system configured to perform a plurality of assays on a single, small volume sample or aliquots thereof.
[0065] [0065] Applicants further describe systems for detecting the presence of one or more of a plurality of markers indicative of an infectious disease, from a clinical sample of small volume, comprising: a) a sample handling system; b) a detection station comprises an optical sensor; c) a fluid handling system configured to transport liquids between components of said system, wherein said fluid transport comprises transporting isolated aliquots of fluid; d) a fluidly isolated sample collection unit configured to hold a clinical sample; e) a test station comprising at least a first, second, and third fluidly isolated test unit, wherein the first unit comprises a first reagent, the second unit comprises a second reagent, and the third unit comprises a third reagent ; and f) a controller, wherein the controller comprises a local memory and is operatively coupled to the sample handling system and the detection station; wherein the system is configured to run tests with any one or more of the first, second, and third test units; wherein the controller buffer comprises a protocol comprising instructions for: i) directing the sample handling system to transfer a portion of the clinical sample to the first test unit, the second test unit and the third test unit; and ii) direct the sample handling system to transfer the first test unit, the second test unit, and the third test unit to the detection station.
[0066] [0066] In embodiments, the system is a point-of-service system. In embodiments, the system is contained within a housing. In embodiments, the fluid handling system is configured to transport fluid within said housing. In embodiments, the system is located at a point-of-service location, and is configured to be used in analyzing a sample at said point-of-service location. In embodiments, the system is a point-of-service system configured to run a plurality of assays on a single small volume sample or on aliquots thereof.
[0067] [0067] In embodiments, applicants disclose a clinical sample processing system, comprising: a) a sample handling system; b) a detection station comprises an optical sensor; c) a fluidly isolated sample collection unit configured to hold a clinical sample; d) an assay station comprising at least a fluidly isolated first, second, and third assay unit, wherein the first unit comprises an antibody, the second unit comprises an oligonucleotide, and the third unit comprises a chromogen or a dye or another marker; and e) a controller, wherein the controller is operatively coupled to the sample handling system, wherein the sample handling system is configured to transfer a portion of the clinical sample from the sample collection unit to each of the first units of assay, the second assay unit, and the third assay unit, and the device is configured to perform an immunoassay, a nucleic acid assay, and a general chemistry assay comprising a chromogen. In embodiments, the system is a point-of-service system. In embodiments, the system is contained within a housing. In embodiments, the system is located at a point-of-service location, and is configured to be used in analyzing a sample at said point-of-service location. In embodiments, the system is a point-of-service system configured to perform a plurality of assays on a single, small volume sample or aliquots thereof.
[0068] [0068] In embodiments, applicants disclose methods of performing at least 4 different assays selected from immunoassays, nucleic acid assays, cytometric assays of, and general chemistry assays on a small volume clinical sample, which comprises: a) introducing a clinical sample having a volume that is not greater than 500 microliters into a sample processing device, the device comprising: i) a sample handling system; ii) a detection station; iii) a cytometry station comprising an imaging device and a stage for receiving a microscopy cuvette; and iv) a test station comprising at least one independently mobile first, second, third, and fourth test unit; b) with the aid of the Han d Ling system sample, the transfer of a portion of the clinical sample to each of the first, and the second, third, fourth test units, in which a different test is performed in each of the first, second, third and fourth test units; C) with the help of the sample handling system, the transfer of the first, second, third, and fourth assay units to the detection station or cytometry station, in which the assay units comprising immunoassays or general chemistry assays are transferred to the detection station and the assay units comprising cytometric assays are transferred to the cytometry station; d) with the aid of the detection station or cytometry station, obtaining measurements of data from the assay performed in each of the first, second, third, and fourth assay units.
[0069] [0069] In embodiments, the methods are point-of-service methods. In embodiments, the system used in carrying out the methods is contained within a housing. In embodiments, the methods are performed at a point-of-service location, and may be used in analyzing a sample at said point-of-service location. In embodiments, the methods comprise point of service methods for performing a plurality of assays on a single small volume sample or on aliquots thereof.
[0070] [0070] In embodiments, the methods include methods of determining the type of infection suffered by a subject. Methods of determining the type of infection as disclosed herein include, without limitation, methods as described herein which comprise determining whether an individual suffers from a bacterial, viral, yeast, fungal, and other infection. For example, methods of determining the type of infection as disclosed in the present invention include methods of determining whether a subject is suffering from a bacterial infection or a viral infection. In embodiments, the methods include methods for detecting, identifying, quantifying, and combinations thereof, markers in a sample indicative of the type of infection suffered by a subject. Methods of detecting, identifying, quantifying, and combinations thereof, markers in a sample indicative of the type of infection, as disclosed herein include, without limitation, methods as described herein comprising detecting, identifying, quantifying, and combinations thereof, markers in a sample indicative of a bacterial, viral, yeast, fungal, and other infection. For example, methods as described in the present invention include methods for detecting, identifying, quantifying, and combinations thereof, markers in a sample indicative of a bacterial infection or a viral infection.
[0071] The methods disclosed herein can be used to determine whether a subject is suffering from, for example, a bacterial, viral, yeast, fungal, and other infection. Determining the type of infection as disclosed herein can be used to guide therapy of the subject suffering from infection. Determining the type of infection as disclosed herein can be used to guide the selection of pharmaceuticals for the treatment of the subject who suffers from infection. Determining the type of infection as disclosed herein can be used to guide the selection of the dosage, or dosage regimen, of drugs used to treat the subject suffering from the infection. For example, the methods disclosed herein can be used to determine whether an individual is suffering from a bacterial infection or a viral infection. The determination of whether a subject suffers from a bacterial infection or a viral infection as disclosed herein can be used to guide therapy of the subject suffering from the infection. The determination of whether a subject suffers from a bacterial infection or a viral infection as disclosed herein can be used to guide the selection of pharmaceuticals for treating the subject suffering from the infection. The determination of whether a subject suffers from a bacterial infection or a viral infection as disclosed herein can be used to guide the selection of a pharmaceutical, dosage selection, dosage regimen, or a combination thereof, used in the treating the subject suffering from the infection. For example, where the infection is determined to be a bacterial infection, antibiotics may be prescribed; however, where the infection is determined to be a viral infection, antibiotics are not indicated, and, in embodiments, will not be fixed. Determining that a subject is suffering from a viral infection may allow the subject to avoid unnecessary treatment and expense (eg where antibiotic therapy is avoided when the infection is identified as being a viral infection). Determining that a subject is suffering from a viral infection may allow the subject to obtain more appropriate therapy directed at viral infections, as opposed to antibiotic therapy which is directed at bacterial infections.
[0072] [0072] Likewise, the determination of whether a subject suffers from a yeast, fungus, and other infection, as opposed to a bacterial or viral infection, may guide or determine the therapy provided to an individual suffering from an infectious disease, including guidance or determining the selection of a pharmaceutical, dosage selection, dosage regimen, or a combination thereof, used in the treatment of the subject suffering from infection. Determining the type of infection may allow the object to obtain a More appropriate therapy targeted at the specific type of infection suffered by the subject, as opposed to inappropriate, or less specific, therapy that may not be as effective for this type of infection.
[0073] [0073] Accordingly, applicants disclose methods of the present invention for providing a prescription for the treatment of an infectious disease in an individual, comprising: providing a clinical sample obtained from a subject; analyzing said clinical sample, wherein the analysis includes testing for, or detecting the presence of, a plurality of disease markers in the clinical sample; determining an appropriate treatment for a disease indicated by the presence of a marker detected by said analysis; and provide a prescription for said appropriate treatment. Applicants further describe methods of the present invention for providing a prescription for the treatment of an infectious disease in a subject, comprising: providing a clinical sample obtained from a subject; analyzing said clinical sample at a point of service (POS) location, the analysis including testing for, or detecting the presence of, a plurality of disease markers in the clinical sample; determining an appropriate treatment for a disease indicated by the presence of a marker detected by said analysis; and provide a prescription for said appropriate treatment.
[0074] [0074] In embodiments of methods for Ding providing a prescription for the treatment of an infectious disease in an individual, analyzing the sample comprises analyzing to determine whether the individual suffers from a bacterial infection, a viral infection, a fungal infection , a yeast infection, other infection, or combination thereof In embodiments of methods for providing a prescription for the treatment of an infectious disease in an individual, analyzing the sample comprises analyzing to determine whether the individual is suffering from an infection. bacterial or a viral infection. In embodiments of methods for providing a prescription for the treatment of an infectious disease in an individual, where sample analysis determines whether the subject is suffering from a bacterial infection, providing a prescription for said treatment comprises the proper prescription of an antibiotic. In embodiments of methods for providing a prescription for the treatment of an infectious disease in an individual, where analysis of the sample determines that the subject is suffering from a viral infection, providing a prescription for said treatment appropriately comprises avoiding the prescription of an antibiotic , and may include prescribing an anti-viral drug. In embodiments of methods for providing a prescription for the treatment of an infectious disease in an individual, where analysis of the sample determines that the subject is suffering from a fungal infection, providing a prescription for said treatment suitable comprises prescribing an antifungal damn it. In embodiments of methods for providing a prescription for the treatment of an infectious disease in an individual, where analysis of the sample determines that the subject is suffering from a bacterial infection which is a mycoplasma infection, providing a prescription for said suitable treatment comprises the prescription of an anti-mycoplasmal drug. In embodiments of methods for providing a prescription for the treatment of an infectious disease in an individual, where analysis of the sample determines that the subject is suffering from a yeast infection, providing a prescription for said treatment suitable comprises the prescription of a drug anti-yeast.
[0075] [0075] Therefore, the systems, devices and methods disclosed herein are point of service methods. In embodiments, the systems described herein, including the systems used in carrying out the methods described herein, can be contained and the methods carried out within a housing. In embodiments, the devices described herein can be placed or used within an enclosure, for example, an enclosure containing a system disclosed herein. In embodiments, the systems described herein may be located at a POS location, and the methods may be performed at a point-of-service location. The systems and methods disclosed herein can be used in analyzing a sample at said point-of-service location. In embodiments, the systems and methods comprise point of service methods for performing a plurality of assays on a single small volume sample or on aliquots thereof.
[0076] [0076] In embodiments, applicants describe systems, methods and devices for detecting one or more of a plurality of markers indicative of a disease in a clinical sample obtained at A (POS) point-of-service location. In embodiments, such a clinical sample is a small volume clinical sample. In embodiments, the one or more markers are detected in a short period of time. In embodiments, the sample is obtained at a POS location. In embodiments, systems and devices may be located at a POS location. In embodiments, detection of one or more markers is performed at a POS location. In embodiments, the diseases are infectious diseases. In embodiments, the diseases are caused by a disease-causing agent selected from the group of disease-causing organisms consisting of a virus, a bacterium (including a mycoplasma), a fungus, a yeast, and other microorganisms. . In embodiments, the diseases are infectious respiratory diseases, and can be upper respiratory diseases, and can be lower respiratory diseases.
[0077] [0077] Therefore, in embodiments, applicants describe POS systems, methods and devices. In embodiments, such POS systems, methods and devices comprise automated POS systems, methods and devices. In embodiments, for example, applicants describe an automated POS system, automated methods, and devices thereof, for detecting one or more of a plurality of markers indicative of a disease in a clinical sample obtained at a capture site; such a disorder may be a respiratory disorder, and may be a disorder caused by a disease-causing agent selected from among (it group of disease-causing organisms consisting of a virus, a bacterium (including a mycoplasma), a fungus, a yeast, and other microorganisms. In embodiments, such automated POS systems, automated methods, and devices thereof, are configured for use at POS sites, and for use with samples obtained at POS locations. such automated POS systems, automated methods, and devices thereof, are configured for use in a single, small-volume clinical sample. In embodiments, such automated POS systems, automated methods, and devices thereof, are configured to detect, if present , one or more of a plurality of markers indicative of a disorder in a clinical sample within a short period of time.
[0078] [0078] In embodiments, such automated POS systems, methods, and devices are located in a POS location selected from a pharmacy, a supermarket, a clinic, a hospital, and a physician's office. In embodiments, a prescription for a treatment is issued at said POS location. In embodiments, a prescription for a treatment is issued at said location POS in accordance with the results of such testing performed by an automated system of POS systems, methods and devices located at the POS location. In embodiments, a prescription for a treatment is filled in at said POS location, where the prescription was issued for a treatment according to respiratory disease, GN performed by an automated system of POS systems, methods and devices located in the POS site. In embodiments, a bill for testing is issued at the POS site; a law like that can be issued automatically. In embodiments, a. bill for a. prescription is issued at the POS site, where the prescription was issued for a treatment is in accordance with the results of such tests performed by an automated system of POS systems, methods and devices located at the capture site; a law like that can be issued automatically. In embodiments, a bill for a test or prescription may be issued from a subject's insurance carrier POS location; a law like that can be issued automatically. In embodiments, an automatic payment may be made for a test performed or a prescription filled to from a subject's insurance carrier POS location, pursuant to an invoice automatically issued from the POS location.
[0079] [0079] Accordingly, applicants disclose devices configured to measure or detect a disease-causing agent or marker indicative of a disease-causing agent in a sample in accordance with a method described herein. Such a sample may be a small volume clinical sample. Such devices can be configured to measure or detect a particular disease-causing agent or marker indicative of a particular disease-causing agent in a sample in less than about three hours, or less than about two hours, or in less than about of one hour, or, in embodiments, in less than about 40 minutes or in less than about 30 minutes.
[0080] [0080] The devices described herein can be configured to perform an assay for the detection or measurement of a plurality of disease-causing agents or markers indicative of the same. The devices described herein can be configured to perform an assay for detecting or measuring a particular disease-causing agent or indicative marker thereof and also to perform an assay which comprises measuring a morphological characteristic of a cell in the sample. The devices described herein can be configured to perform an assay for measuring a disease-causing agent or indicative marker thereof and also to perform an assay comprising the measurement of another substance, e.g., a cytokine, a prostaglandin, histamine , a steroid (e.g., a glucocorticoid, or other steroid), a vitamin, a hormone, a drug or metabolite of a drug, or other analyte. Such devices can be configured in which the tests, or the order of execution of tests, which are performed by said device can be changed by communication with another device.
[0081] [0081] The methods and compositions described herein provide rapid assays that require only small amounts of sample, such as only small amounts of saliva, urine, blood, or fluid, into which a throat swab, buccal swab, or nasal secretion has been immersed ΐη embodiments, a plurality of samples, including a plurality of small samples, may comprise a plurality of sample types, such as saliva, urine, blood, or fluid samples, may be supplied to, and analyzed by, the systems. , devices and methods disclosed here at. The methods, devices, and systems described herein are configured to perform such rapid assays that require only small amounts of sample. Methods, apparatus and systems disclosed herein are configured to perform such rapid assays on a plurality of sample types, and may require only small amounts of each sample type. The methods, devices and systems described herein are configured to perform multiple assays on a sample, or on a plurality of sample types, and can be used to screen for one or more of a plurality of diseases. Methods, apparatus and systems described herein are configured to perform multiple assays on a sample, or on a plurality of sample types, and can be used to screen for diseases caused by one or more than twenty bacteria, yeast, fungi, mycoplasma, Archea, fungi, yeasts, parasites and other microorganisms. Hence the methods. Devices and systems described herein provide rapid testing, requiring only small clinical samples, and thus provide advantages over other methods, devices and systems. BRIEF DESCRIPTION OF THE DRAWINGS
[0082] [0082] Fig. 1 A provides a graphical summary of the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample by a variety of markers and for two different concentration ranges of the markers (10 C μl and 100 C / ID, where "C / μΓ means copies per microliter (μl, )). The times are labeled "LOD" ("delay length"). The vertical axis is shown in fluorescence units relative (relative fluorescence units, RFU) in thousands.
[0083] [0083] Fig. IB provides a bar graph showing the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample for the indicated markers for various diseases (at 00 C / U!).
[0084] [0084] Fig. IC provides a bar graph showing the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample for the markers indicated by various strains of influenza and to identify targets (at 100 °C / μl).
[0085] [0085] Fig. ID provides a bar graph showing the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample for the indicated markers of various respiratory diseases (at 100 °C / µl).
[0086] [0086] Fig. IE provides a bar graph showing the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample for the indicated markers for various sexually transmitted diseases (a 100 °C / μl).
[0087] [0087] Fig. SE provides a bar graph showing the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample for the indicated markers for various diseases that can be detected in the blood (at 100 °C / μl) .
[0088] [0088] Fig. 2A shows amplification over time, showing detection of influenza A (seasonal H11 strain) marker sometimes well before any significant amounts of amplification of the target non-nucleic acid t message has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[0089] [0089] Fig. 2B shows the detection limit of influenza A virus (seasonal strain HlNl) in a sample. The height of the bars indicates the time until the copy number shows an inflection (rises significantly above the background level), with the horizontal axis indicating the initial copy numbers of the influenza A virus message (seasonal strain HlNl); "NTC" indicates "no template controls" (no additional copies of the target marker). FIG. 2B shows detection of influenza A virus (seasonal H11 strain) marker sometimes well before any significant amounts of non-target message nucleic acid amplification has occurred.
[0090] [0090] Fig. 3A shows amplification over time, which shows detection of influenza A (new Hl l strain) marker sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[0091] Fig. 3B shows the detection limit of influenza A virus (new HI strain) in a sample. The height of the bars indicates the time until the copy number shows an inflection (rises significantly above the background level), with the horizontal axis indicating the initial copy numbers of the influenza A virus message (new HLN strain G); "NTC" indicates "no template controls" (no additional copies of the target marker). FIG. 3B shows detection of the influenza A (new HlNl strain) marker sometimes well before any significant amounts of amplification of the non-target nucleic acid message has occurred,
[0092] [0092] Fig. 4A shows amplification over time, showing detection of influenza A (H3N2 s train) marker sometimes well before any significant amounts of amplification of the non-target nucleic acid message have occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands,
[0093] [0093] Fig. 4B shows the detection limit of influenza A virus (strain H3N2) in a sample. The height of the bars indicates the time until the copy number shows an inflection (rises significantly above the background level), with the horizontal axis indicating the initial copy numbers of influenza A virus (strain H3N2) of the message; "NTC" indicates "no template control" (no appended copies of target marker). Fig. 4B shows detection of the influenza A (strain H3N2) tag sometimes well before any significant amounts of amplification of the non-target nucleic acid message has occurred.
[0094] [0094] Fig. 5A shows amplification over time, which shows the detection of influenza virus A (H7N9 sirain) marker in the iimes well before any significant amounts of non-target message nucleic acid amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands,
[0095] [0095] Fig. 5B shows the detection limit of influenza A virus (strain H7N9) in a sample. The height of the bars indicates the time until the copy number shows an inflection (rises significantly above the background level), with the horizontal axis indicates the initial copy numbers of the influenza A message (strain H7N9); "NTC" indicates "no template controls" (no additional copies of the target marker). FIG. 5B shows detection of influenza A virus (H7N9 strain) marker sometimes well before any significant amounts of non-target message nucleic acid amplification has occurred.
[0096] [0096] Fig. 6A shows amplification over time, which shows detection of influenza A virus (strain H5N1) marker sometimes well before any significant amounts of non-target message nucleic acid amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[0097] [0097] Fig. 6B shows the detection limit of influenza A virus (H5N 1 strain) in a sample The height of the bars indicates the time until the copy number shows an inflection (raises significantly above the background level), with the horizontal axis indicates the initial copy numbers of the influenza A message (strain Ή5Ν1); "NTC" indicates "no template controls" (no additional copies of the target marker). FIG. 6B shows detection of the influenza A (H5N1 sirain) marker sometimes well before any significant amounts of amplification of the non-target nucleic acid message has occurred.
[0098] [0098] Fig. 7 A shows amplification over time, which shows the detection of influenza B marker sometimes well before any significant amounts of non-target message nucleic acid amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[0099] [0099] Fig. 7B shows the detection limit of influenza B in a sample. The height of the bars indicates the time until the copy number shows an inflection (rises significantly above the background level), with the horizontal axis indicating the initial influenza B message copy numbers; "NTC" indicates "no template controls" (no additional copies of the target marker). FIG. 7B shows detection of the influenza B marker sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred.
[00100] [00100] Fig. 8 A shows amplification over time, showing detection of the Influenza Matrix protein marker sometimes well before any significant amounts of amplification of the non-target nucleic acid message has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands,
[00101] [00101] Fig. 8B shows the detection limit of the influenza marker protein matrix in a sample. The height of the bars indicates the time until the copy number shows an inflection (rises significantly higher than the background level), with the horizontal axis indicating the initial copy numbers of influenza matrix protein message; "NTC" indicates "no template controls" (no additional copies of the target marker). FIG. SB shows detection of the influenza marker matrix protein sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred.
[00102] [00102] Fig. 9A shows amplification over time, which shows detection of a tuberculosis marker (Myobacterium tuberculosis) sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00103] [00103] Fig. 9B shows the detection limit of tuberculosis in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly above the background level), with the numbers "TB 1000" indicating 1000 copies of the TB marker message, "TB 100" indicating 100 copies of TB marker message and "TB 10" indicating 10 copies of TB marker message; "NTC" indicates "no template controls" (no additional copies of the target marker).
[00104] [00104] Fig. 10A shows amplification over time, which shows detection of a staphylocccus marker (Staphylococcus aureus) sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00105] [00105] Fig. lob shows the detection limit of Staphylococcus aureus in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly above the background level), with the horizontal axis indicating the number of copies of Staphylococcus aureus message: "NTC" indicates "no model controls " (not added copies of target marker).
[00106] [00106] Fig. 1 1A shows amplification over time, which shows detection of a staphylocccus marker (Methiciliin-resistant Staphylococcus aureus - MRSA) sometimes well before any significant amounts of nucleic acid message amplification non-target occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The Vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00107] [00107] Fig. 1 IB shows the detection limit of MRSA Staphylococcus aureus in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly above the background level), with the horizontal axis indicating the number of copies of the Staphylococcus aureus MRSA message; "NTC" indicates "no template controls" (no additional copies of the target marker).
[00108] [00108] Fig. 12A shows amplification over time, which shows detection of a streptococcal marker (Group A streptococci) at times well before any significant amounts of amplification of the non-target nucleic acid message has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00109] [00109] Fig. 12B shows the detection limit of Group A Streptococcus in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly higher than the background level), with the horizontal axis indicating the number of copies of the Streptococcus A message group; "NTC" indicates "no template controls" (no additional copies of the target marker).
[00110] [00110] Fig. 13A shows amplification over time, which shows detection of a Bordetella pertussis marker sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00111] [00111] Fig. 13B shows the detection of Bordetella pertussis in a sample. The height of the bars indicates the time until the copy number shows an inflection (raises significantly above the background level), with the horizontal axis indicating the number of copies of the Bordetella pertussis message (as final DNA copy per μl, ); "NTC" indicates "no template controls" (no additional copies of the target marker).
[00112] [00112] Fig. 14A shows amplification over time, which shows detection of an adenovirus B marker sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00113] [00113] Fig. 14B shows the detection limit of adenovirus B in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly higher than the background level), with the horizontal axis indicating the number of copies of the adenovirus B message; "NTC" indicates "no template controls" (no additional copies of the target marker).
[00114] [00114] Fig. 15A shows amplification over time, which shows detection of an adenovirus C marker sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[001153] [001153] fig. 15B shows the limit of detection of adenovirus C in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly higher than the background level), with the horizontal axis indicating the number of copies of the Adenovirus C message; "NTC" indicates "no template controls" (no additional copies of the target marker).
[00120] [00120] Fig. 18A shows amplification over time, which shows detection of a Treponema pallidum marker sometimes well before any significant amounts of non-target nucleic acid message amplification has occurred. The horizontal axis is termed "cycles", although temperature cycling was not used; each unit of "cycles" is about a minute, so the horizontal axis numbers can be read in terms of minutes. The vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00121] [00121] Fig. 18B shows the detection limit of the presence of Treponema pallidum in a sample. The height of the bars indicates the time until the number of copies shows an inflection (rises significantly above the background level), with the horizontal axis indicating the number of copies of the Treponema pallidum message; "NTC" indicates "no template controls" (no additional copies of the target marker).
[00122] [00122] Fig. 19A shows the amplification over time, the increase in fluorescence relative to about 15 to 20 minutes indicating the presence of a seasonal marker HlNl influenza virus. The horizontal axis is in "minutes, the vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU).
[00124] [00124] Fig. 2.0A shows an exemplary vessel for holding a rod (a swab vessel) and an exemplary cartridge (which includes wells and wells of reagents and vessels, and is configured to hold the reagent vessels, reaction, and other containers and utensils). The arrows moving away from the swab vessel indicate how the wick container can be placed inside a receptacle on the cartridge.
[00125] [00125] Fig. 20B shows an exemplary swab container (configured to hold a rod) and an exemplary cartridge (which includes reagent wells and wells and vessels, and is configured to hold reagent vessels, reaction vessels, and other containers and utensils), in addition to the wells and wells configured to contain reagent containers, reaction vessels, and other containers and utensils, as shown in the embodiment of fig. 20A, the exemplary cartridge shown in Fig. 20B includes wells and wells suitable for holding other sample containers, e.g., blood or urine vessels, in addition to smear vessels. The arrows moving away from the swab vessel indicate how the wick container can be placed inside a receptacle on the cartridge.
[00126] [00126] Fig. 20C shows an exemplary swab container, and an exemplary cartridge that includes wells and wells for carrying a swab rod and vessel, as well as wells and wells configured to hold the reagent vessels, reaction vessels, and other containers and instruments (which may optionally include other sample containers, eg blood vessels or urine samples). The arrows moving away from the rod indicate how the swab can be placed inside a swab receptacle on the cartridge. The arrows moving away from the swab vessel indicate how the wick container can be placed inside a swab container receptacle on the cartridge.
[00127] [00127] Fig. 21 shows examples of cotton swabs that can be used to obtain samples from the throat, nasal passages, cheeks, or other body locations of individuals.
[00129] [00129] Fig. 23A shows several nomenclature panels of influenza types of influenza that may be identified by the methods and devices discussed here.
[00130] [00130] Fig. 2.3B shows the inflection times for various types of influenza that can be identified by the methods and devices discussed here.
[00131] [00131] Fig. 24A shows several respiratory disease panels naming types of respiratory diseases, which can be identified by the methods and devices discussed here.
[00132] [00132] Fig. 24B shows the inflection times for upper and lower respiratory tract disease types, which can be identified by the methods and devices discussed here.
[00133] [00133] Fig. 25A shows several panels of hospital-acquired infectious diseases nomenclature types of respiratory diseases, which can be identified by the methods and devices discussed here,
[00134] [00134] Fig. 25B shows the inflection times for various hospital-acquired infectious disease panels of nomenclature types of respiratory diseases, which can be identified by the methods and devices discussed here.
[00135] [00135] Fig. 26 shows results of an influenza assay
[00136] Fig. 27 shows the specificity of target nucleic acid assays for influenza type H2N2.
[00137] [00137] Fig. 28 shows the specificity of nucleic acid assays for the seasonal type influenza H1N1 target.
[00138] [00138] Fig. 29 lists potential interfering substances for the sexually transmitted disease (STD) panel, which have been found not to interfere with nucleic acid assays.
[00139] [00139] Fig. 30 lists potentially interfering substances for sexually transmitted disease (STD) from the urine panel that have been found not to interfere with nucleic acid assays,
[00140] [00140] Fig. 31 lists potential interfering substances for the blood panel that have been found not to interfere with nucleic acid assays. DETAILED DESCRIPTION
[00141] [00141] Description and disclosure of examples of reagents, assays, methods, kits, devices and systems that can be used with the methods, assays and reagents, devices and systems described herein can be found, for example, in US Patent 8,088. 593; US patent
[00142] [00142] Disclosures of nucleic acid target detection methods include, for example, methods, assays and reagents, and devices, as disclosed in US Application Serial No. 61 / 800,606, filed March 15, 2013; US Serial Application No. 61 / 908,027, filed November 22, 2013. US Serial Application No. 62 / 001,050, filed May 20, 2014; US Serial Application No. 14 / 214,850, filed March 15, 2014; PCT / US2014 / 030034, filed on March 15, 2014; US Serial Application No. 61 / 800,241, filed March 15, 2013; and US Application Serial No. 61 / 800,340, dated March 15, 2013; the disclosures of which patent applications are hereby incorporated by reference in their entirety. Other methods for detecting nucleic acid targets include, for example, Polymerase Chain Reaction (PCR), methods described, for example, in U.S. Pat. No, 4683, 195; and, in general, Muliis et al, Cold Spring Harbor Symp. Quantity Biol. 51: 263 (1987); Erlieh, Ed., PGR Technology (Stockton Press, New York, 1989).
[00143] Disclosure of methods for deleting protein targets, including antibody methods, can be found, for example, in U.S. Pat. No. 4,376, from 1 to 10; Pat. No. 4.81 6.567; Pat. No. 7429652; The Patent
[00144] [00144] Disclosure on systems, devices and methods for analyzing clinical samples, including clinical samples, such as small volume clinical samples, and including systems, devices and methods for analyzing small volume clinical samples in short periods of time , can be found, for example, in US Patent 8,380,541; US patent
[00145] [00145] Before presenting the new target binding molecules, compositions, assays, methods and kits are disclosed and described, it should be understood that the terminology used herein is for the purpose of describing only particular embodiments and is not intended to be limiting. It is also to be understood that the present disclosure provides explanatory and exemplary descriptions and examples, so that, unless otherwise indicated, the molecules, compositions, methods, assays and kits described herein are not limited to the specific embodiments herein. described.
[00146] [00146] It should be noted that, as used in the specification and the appended claims, the singular forms "a", "an" and "o" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a salt thereof" refers to a single salt or mixtures of different salts, and the like.
[00147] In this specification and in the claims which follow, reference will be made to a number of terms which will be defined as having the following meanings:
[00148] [00148] Acronyms and abbreviations, such as "rpm" (revolutions per minute), "min" (minutes), "sec" (second), and so on, have their usual meanings.
[00149] [00149] As used herein, the terms "normal" and "normal level" refer to the levels of a marker as found in a healthy population of normal individuals. For example, the normal level for a particular type of white blood cell found is the level of that type of white blood cell found in blood samples from a healthy population of normal individuals.
[00150] [00150] As used herein, the terms "high" and "high level" and the like refer to levels that significantly exceed normal levels, i.e., a high level of a marker is one that significantly exceeds the levels of which marker that is found in a healthy population of normal individuals.
[00151] [00151] As used herein, the terms "low" and "low level" and the like refer to levels that are below normal levels, i.e., a low level of a marker is one that is below the levels of that marker. which is found in a healthy population of normal individuals.
[00152] [00152] It should be understood that where a marker is typically absent, or sparse, in normal subjects, a normal level of a marker may be very low in absolute numbers (eg, as measured by the number of markers per unit volume, or marker weight per volume unit), and remains the normal level for that marker. So, for example, where the marker is an antibody to a particular infectious disease, and healthy, normal individuals have not recently been exposed to that particular disease, normal levels of antibodies to that disease may be low in absolute terms, and levels of a subject that exceed the normal level could indicate that the subject has recently been exposed to, or suffers from, an infection or disease.
[00153] The term "isolated", as used herein, when used to describe the various nucleic acids and proteins disclosed herein, means that the nucleic acid or protein (or other molecule) has been separated and/or recovered from, by the less, a contaminant with which it is normally associated. Ordinarily, however, isolated nucleic acids and proteins will be prepared by at least one purification step.
[00154] [00154] The "portion" Tenn as used herein refers to any particular composition of matter, for example, a molecular fragment, an intact molecule, or a mixture of materials.
[00155] [00155] As used herein, the terms "disease agent - causing", "disease causing organism" and plurals and their grammatical equivalents are used interchangeably to refer to viruses, bacteria, yeasts and other microorganisms that can cause disease in a subject. Thus, when referring to diseases and their causes, the terms "agent", "organism" and plurals and grammatical equivalents are used interchangeably herein.
[00158] As used herein, the term "bacteria" refers to small unicellular organisms, prokaryotes that can infect the cells, organs, tissues or surfaces of plants or animals, including humans. The term bacteria includes Gram negative and Gram positive bacteria. Bacteria can cause illness. Mycoplasma are a form of bacteria that lack cell wailing,
[00159] As used herein, the term "drug" is used broadly to refer to any agent that can be administered to a subject for the purpose of treating a disease or condition suffered by the subject; such treatment may include prevention, amelioration of symptoms, speeding recovery, strengthening the patient in the face of a disease or condition, as well as directly combating the disease or condition. Where the disease or condition results from an infection, for example, is due to an infectious disease, the drug may be, without limitation, an antibiotic, an antiviral scavenger, an antifungal scavenger, an anti-mycoplasma drug, an anti-yeast drug, or combinations thereof.
[00160] [00160] As used herein, the term "antibiotic" is used widely to refer to drags which act to reduce or eliminate bacterial infections. Antibiotics include, without limitation, penicillin, ampiciflin, amoxicillin, tetracycline, oxytetracycliiie, doxycycline, minocycline, a sulfa-drug sulfonamide (such as, for example, sulfanilamide, sulfamethoxazole, sulfadiazine), erythromycin, ciprofloxacin, gentamicin, omitomycin , clarithromycin, a cephalosporin, for example, cefaclor, cefprozil, cefuroxime axetil, loracarbef, cefdinir, cefixime, eefpodoxime proxetil, ceftibutene, or ceftriaxone, gramicidin, valinomycin, nonactin, aiameihicin, and other antibiotics.
[00161] As used herein, the term "anti-mycoplasma" is used in a broad sense to refer to drugs that act to reduce or eliminate bacterial infections, where the bacteria are mycoplasma. Antibiotics that target the cell wall are usually ineffective against mycoplasma, which lack wailing cells. Antibiotics, such as, for example, piasmoein, doxocycline, minocycline, gramicidin, valinomyein, nonactin, a] ametbicin, macrolide antibiotics, and others can be used to treat mycoplasma infections.
[00162] As used herein, the term "antiviral" is used in a broad sense to refer to drugs that act to reduce or eliminate viral infections. Antiviral drugs include, for example, zanamivir, oseltamivir, acyclovir, adefovir, darunivir, famciclovir, ganciclovir, nexavir, rifampicin, pieconaril, amantadine, rimantadine, and others.
[00163] As used herein, the term "antifungal" is used in a broad sense to refer to drugs that act to reduce or eliminate fungal infections. ant-fungal drugs include, for example, amphotericin, nystatin, candicin, filipin, hamycin, netanicin, rimocydin, bifonazole, clotrimazole, other imidazoles, triazoles, and thiazoles, and others. Some drags that can be used to treat yeast infections may also be suitable for treating yeast infections.
[00164] As used herein, the term "anti-yeast" is used in a broad sense to refer to drugs that act to reduce or eliminate fungal infections. Anti-yeast medications include, for example, antimycotics, such as, for example, clotrimazole, nystatin, fluconazole, ketoconazole, amphotericin, gentian violet, and other dr. Some dr gs that can be used to treat yeast infections may also be suitable for treating yeast infections.
[00165] As used herein, the phrase "nucleic acid markers indicative of" a particular infection, refers to nucleic acid molecules (including single-stranded and double-stranded DA and RNA molecules) and fragments thereof, which are derived from from disease-causing organisms, or are copies of, or are substantially similar to, or are complementary to nucleic acid molecules from the organism causing that particular infectious disease. Detection of nucleic acid markers indicative of a particular infection in a sample indicates that the disease-causing organism is, or was, present in the sample and, therefore, that the subject has been exposed to the disease-causing organism, and is likely to suffer or have suffered from a specific infection caused by that same disease-causing organism.
[00166] [00166] As used herein, the phrase "antibody markers indicative of a particular infection, refers to antibodies (or parts thereof or fragments thereof) that are directed to an antigen or antigens found on the organisms that cause that infectious disease in particular. Detection of antibody markers indicative of a particular infection in a sample indicates that the disease-causing organism is, or was, present in the sample and, therefore, that the subject has been exposed to the disease-causing organism, and is likely to suffer or have suffered. specific infection caused by that particular disease-causing organism.
[00167] As used herein, a nucleic acid comprising a nucleotide compound molecule, and refers to deoxyribonucleic acid (DNA) and ribonucleic acid for RNA molecules ().
[00170] As used herein, a "target" nucleic acid or molecule refers to a nucleic acid of interest. A nucleic acid/molecule target can be of any type, including single-stranded or double-stranded DNA or RNA (eg mRNA).
[00171] As used herein, a nucleic acid molecule that is described as containing the "sequence" of a template or other nucleic acid may also be considered to contain the template or other nucleic acid itself (e.g., a molecule that is described as containing the sequence of a template can also be described as containing the template), unless the context clearly dictates otherwise.
[00172] As used herein, "complementary" sequences refer to two nucleotide sequences which, when aligned antiparallel to each other, contain several individual nucleotide bases which are paired with each other. It is not necessary for all nucleotide bases in two sequences to pair with each other for sequences to be considered "complementary". Sequences can be considered as complementary, for example, if at least 30%, 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95 %, 98%, 99%, or 100% of the nucleotide bases in two sequences pair with each other. Furthermore, sequences can still be considered "complementary" when the overall lengths of the two sequences are significantly different from each other. For example, a 15 nucleotide primer can be considered "complementary" to a larger polynucleotide containing hundreds of Nucl eotides if several individual nucleotide bases of the nucleotide base pair long polynucleotide when the primer is antiparallel aligned to a particular region of the polynucleotide. larger polynucleotide.
[00173] As used herein, a "concatemer" refers to a nucleic acid molecule which contains within it two or more copies of a particular nucleic acid, wherein the copies are linked in series. Within the concatemer, the copies of the particular nucleic acid can be linked directly to one another, or they can be indirectly linked (for example, there can be nucleotides between the copies of the particular nucleic acid). In one example, the particular nucleic acid may be that of a double-stranded nucleic acid template, such that a concatemer may contain two or more copies of the double-stranded nucleic acid template. In another example, the particular nucleic acid can be that of a template polynucleotide, such that a concatemer can contain two or more copies of the template polynucleotide.
[00174] [00174] As used herein, a "saccharide" is a molecule comprising one, some, or several sugar moieties, and includes monosaccharides, oligosaccharides, and polysaccharides.
[00175] As used herein, a protein comprises a molecule composed of amino acids, the amino acids covalently linked by amide bonds. The terms "peptide", "polypeptide" and "protein" can be used interchangeably to refer to molecules composed of amino acids linked by peptide bonds. Individual amino acids can be referred to as "residues" of a polypeptide or protein. The amino acid sequences of the polypeptides disclosed herein can be identified by SEQ ID NO: presented as a sequence of letters, where the letters have the following meanings: TABLE IB
[00176] As used herein, a "cytokine" is a naturally occurring protein molecule in mammals frequently released in response to injury, infection, inflammation, or other stressor. Cytokines include iymphokines, interleukins, chemokines, interferons, and other cytokines. Cytokines can be inflammatory cytokines (which tend to cause inflammation, also called pro-inflammatory cytokines), or they can be anti-inflammatory cytokines (which tend to suppress inflammation). Inflammatory cytokines include, for example, interleukin-1 (IL-1), interleukin-6 (IL-6), interleukin-12 (IL-12), interleukin 18 (TL-18), interferon gamma (IF - γ ) , and tumor necrosis factor alpha (TNF-a). anti-inflammatory cytokines include, for example, interleukin-10 (EL-10). Some cytokines can have various actions or effects (for example, interleukin-6 (IL-6) has inflammatory and anti-inflammatory effects), [001773 As used herein, the terms "inflammation marker", "inflammatory marker", and plurals and their grammatical equivalents refer thereto to markers that can be detected in a sample, and that can be identified in a sample, that indicate the presence or level of inflammation in the subject from which the sample was obtained. . Markers of inflammation include both peptide and non-peptide markers; for example, inflammation markers include, without limitation, prostaglandins, tumor necrosis factor alpha (TNF-a), interleukin-1 (IL-1), interleukin-8 (IL-8), interleukin-12 (IL-12) , interferon gamma (lf-γ), bradykinin, complement system molecules, blood clotting factors, C-reactive protein, erythrocyte sedimentation rate (ESR), white blood cell count, and moiphological changes in blood and others cells.
[00178] The term "antibody" is used in the broadest sense and specifically covers individual monoclonal antibodies (including agonist and antagonist antibodies), antibody compositions with polyepitopic specificity, and antibody fragments, and includes human humanized antibodies. Monoclonal antibodies are obtained from a population of substantially homogeneous antibodies, that is, the individual antibodies that make up the population are identical except for possible naturally occurring mutations that may be present in smaller amounts. The most abundant class of antibodies is the IgG class, characterized by having molecular weights of about 150 kD.
[00179] The term "monoclonal antibody" (mAb) as used herein refers to an antibody obtained from a population of substantially homogeneous antibodies, ie, the individual antibodies making up the population are identical except for possible naturally occurring mutations which may be present in smaller amounts. For example, monoclonal antibodies can be produced by the meihod hybridoma first described by Kohler et al., Nature, 256:. 495 (1975), or can be produced by recombinant DNA methods (see, for example, US Patent No. 4,816,567 to Cabilly. et al).
[00180] The term "intact antibody" refers to the complete antibody, or the amino acid sequence of the complete antibody, of which an antibody fragment is a part. It should be understood that an antibody fragment can be produced by partial digestion (e.g., by papain or pepsin) of an intact antibody, or can be produced by recombinant or other means.
[00182] An antibody fragment is, or comprises, a polypeptide having a primary structure consisting of an uninterrupted sequence of contiguous amino acid residues that have the amino acid sequence of an intact antibody. Examples of antibody fragments include Fab, Fab', Fab'-SH, F fragments (ab', Fd, Fc, Fv fragments, diabodies, and any other "non-single chain antigen binding unit" as described, for example , in Pat. No. 7,429,652.
[00183] The term "diabodies" refers to small antibody fragments with two antigen-binding sites, which fragments comprise a heavy chain variable domain linked to a light chain variable domain on the same polypeptide chain. Using a linker that is too short to allow pairing between the two domains on the same chain, the domains are forced to pair with the complementary domains of another chain and create two antigen-binding sites. Diabodies are more fully described in, for example, EP 404,097; WO 93/1161; and Hollinger et al, Proc. Natl. Academic Sci. LISA, 90:6444-6448 (1993).
[00184] As used herein, an "antigen-binding antibody fragment" is any antibody fragment which retains the ability to bind the specific target to which the antibody specifically binds intact. An antigen-binding antibody fragment may have (eg, lower) different binding affinity for the target antigen than the intact antibody. As used herein, unless otherwise indicated, an antibody fragment is an antigen-binding antibody fragment.
[00185] An antibody that "specifically binds to" or is "specific to" a particular polypeptide or, an epitope on a particular polypeptide is one that binds to that particular polypeptide or epitope on a particular polypeptide, without substantially binding to any other polypeptide or polypeptide epitope.
[00186] The terms "antigen", "target molecule", "target polypeptide", "target epitope", and the like are used herein to denote the molecule specifically bound by an antibody or antibody fragment.
[00187] As used herein, a "label", a "label", a "portion marker" and a "label portion" refer to a detectable compound or composition that is directly or indirectly conjugated to the antibody in a manner to generate a " labeled "antibody. A label or marker provides a detectable signal for at least the period of time during which a signal is to be observed. The label or label may be detectable on its own (for example radioisotope labels or fluorescent labels) or, in the facility of an enzyme label, it may catalyze a chemical change to a compound or substrate composition that is detectable.
[00188] [00188] A label or marker moiety can be, for example, a dye, an epitope tag, a fluorescent moiety, a luminescent moiety, a chemiluminescent moiety, an enzyme label, a magnetic label, a paramagnetic label, an agent contrast, a nanoparticfe, a radioisotope, biotin, streptavidin, and a fire extinguisher. A nanoparticle can be a particle of an element, such as a gold or alloy or composite nanoparticle, such as a quantum dot (a particle of a semiconductor material), or another particle with a size typically in the range of about 1 race for about 100 run.
[00189] [00189] A label or marker portion may provide a signal by reflecting, or modulating, the energy that impinges on the label portion or a marker, a label or marker portion may provide a signal by emitting, or increasing, a detectable signal. Likewise, a label or marker portion may provide a signal by decreasing, or extinguishing, a signal (eg, extinction of a signal). It should be understood that a label or marker portion may be directly detectable (e.g. it may provide a detectable signal without further measurements or energy input), or may use or require energy, a substrate, a binding partner, or any other action in order to provide a detectable signal. An enzyme label may be suitable for use with a binding partner or substrate; for example, a peroxidase such as horseradish peroxidase can serve as a marker, as they can be used to detect the presence of a target, or to measure the amount of target, when used with, for example, diaminobenzidine or other suitable molecule for use with a peroxidase; On the other hand, a non-limiting example, luciferase can serve as a marker, as it can be used to detect the presence of a target, or to measure the amount of target, when used with luciferin.
[00190] [00190] As used herein, the term "chromogen" refers to a compound that can be readily converted to a dye or other colored compound.
[00191] As used herein, "BSA" means bovine serum albumin; "PEG" means polyethylene glycol: "ELISA" means enzyme-linked immunosorbent assay; and other terms, abbreviations and acronyms have the standard meanings understood in the biological and chemical arts IHE e.
[00192] [00192] A composition may include a buffer. Buffers include, without limitation, phosphate, citrate, ammonium, acetate, carbonate, tris(hydroxymethyl) aminomethane (TR1S), 3-(non-morphoi)propanesulfonic acid (MOPS), 3-morpholmo-2-hydroxypropanesulfonic acid (MOPSO). ), 2-(N-morpholino)ethanesulfonic acid (MES), N-(2-acetamido)-iminodiacetic acid (ADA), piperazine-N, N'-bis (2-ethanesulfonic acid) (PIPES), Ν- (2 .-acetamido) -2-aminoethanesulfonic (ACES), cholamine chloride, N,N-bis(2-hydroxyethyI) -2-aminoethanesulfonic (BBS), 2-[[1,3-d hydroxy-2-(hydroxymethyl) propan-2-yl]amino]ethanesulfonic (TES), 4-i2-hydroxyethyI) - 1 ethanesulfonic piperazine (HEPES), acetamidogycine, tricine (N-(2-hydroxy-1,1-bis(hydroxymethyl)ethyl)glycine ), glycinamide, and bicin (2-(bis(2-hydroxyethyl)amino)acetic) buffers. Buffers include other organic acid buffers in addition to the phosphate, citrate, ammonium acetate, carbonate, and buffers explicitly mentioned here,
[00193] [00193] Composition A may include a physiologically acceptable winner. For example, a physiologically acceptable vehicle can be an aqueous pH buffered solution. Examples of physiologically acceptable carriers include buffers such as phosphate, citrate, and other organic acids as discussed above; antioxidants including ascorbic acid; low molecular weight (less than about 10 residues); protein polypeptides, such as, for example, serum albumin, gelatin, cytochromes, and immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, arginine or lysine; monosaccharides, disaccharides, polysaccharides, and other carbohydrates including glucose, mannose, and dextrins; chelating agents such as ethylene diamine tetraacetic acid (EDTA); sugar alcohols such as mannitol or sorbitol; salt-forming eunterions such as sodium, potassium, calcium, magnesium and others; nonionic surfactants such as TWEEN™, polyethylene glycol (PEG), and Pluronics™; and/or other compounds known in the art. For example, a composition may include albumin, gelatin, cytochrome C, an immunoglobulin, an amino acid, agar, glycerol, ethylene glycol, a protease inhibitor, an antimicrobial agent, a chelating agent of metal, a monosaccharide, a disaccharide, a polysaccharide, a reducing agent, a chelating agent, or combinations thereof.
[00196] [00196] Biological and clinical samples may include nasopharyngeal lavage, or other fluid obtained by washing a body cavity or surface of an object, or by washing a cotton swab after applying the swab to a body cavity or surface of an object . N asal swabs, mouth swabs (including cheek swabs), swabs, vaginal discharge, stool samples, hair, Page 5: fingernail, ear wax, breath, and other solids, semi-solids, or gaseous can be transformed into an extraction buffer, for example, for a fixed or variable amount of time, prior to analysis. The extraction buffer or an aliquot thereof can then be processed similarly to other fluid samples if desired. Examples of tissue samples from the subject may include, but are not limited to, connective tissue, muscle tissue, nervous tissue, epithelial tissue, cartilage, cancerous tissue, or bone. The sample can be obtained from a human or animal. The sample can be obtained from a vertebrate, for example, a bird, fish or mammal, such as a rat, a rat, a pig, a monkey, another primate (including humans), a farm animal, a sport animal, or a pet. The sample can be obtained from an object being or dead. The sample may be obtained fresh from a subject or may have undergone some form of pre-processing, storage or transport,
[00197] [00197] As used herein, a "small volume" refers to a volume of less than about 1 mL, or less than about 500 µ, Ε, or less than about 250 µl, or less than 150 Al, or less than about 100 μl ,, or less than about 50 μί ^ , or less than about 2.5 μϋ,
[00198] [00198] A sample can be divided into two or more portions. As used herein, when referring to a sample or samples, the terms "portion" and "aliquot" and their plurals and their grammatical equivalents are used interchangeably to refer to a fractional amount of sample taken from a complete original sample. . a Such fraction may be any fraction or amount, such that a portion or aliquot may comprise most of the original sample, a large fraction of the original sample, a small fraction of the original sample, or a relatively small fraction of the original sample. The phrases "at least a. Portion", "at least an aliquot", and the like, can refer both to a fractionated portion of an original sample and to the entire original sample.
[00199] [00199] The detection of markers, and detection of disease-causing organisms (or others) may include detection of nucleic acid markers; detection of protein (peptide) markers, including detection of antibodies; detection of inflammation markers (including both peptide and non-peptide markers); and detection of other markers. Identification of markers, and disease-causing organisms (or others) may include identification of nucleic acid markers; the identification of protein (peptide) markers, including the identification of antibodies; the identification of inflammation markers (including both peptide and non-peptide markers); and identification of other markers. Detection and identification of markers and organisms may include the quantitative detection and identification of markers and such organisms.
[00200] [00200] A method can be performed in a short period of time. A device may be able to perform all steps of a process in a short period of time. A device can be able to perform all steps of a method in a single sample in a short amount of time. A device may be able to perform all steps of a process on two samples, such as a blood sample and a sample obtained from a swab, in a short amount of time. A device may be capable of performing all steps of a process on more than two samples in a short period of time. For example, from taking a sample from an individual to detecting one disease marker, or to detecting multiple disease markers, it can take about 3 hours or less, 2 hours or less, 1 hour or less, 50 minutes or less, 45 minutes or less, 40 minutes or less, 30 minutes or less, 20 minutes or less, 15 minutes or less, 10 minutes or less, 5 minutes or less, 4 minutes or less, 3 minutes or less, 2 mmutes or less, or 1 minute or less. For example, from an individual's sample collection to the transmission of data regarding, and/or the analysis of a sample or samples may take about 3 hours or less, 2 hours or less, 1 hour or less, 50 minutes or less, 45 minutes or less, 40 minutes or less, 30 minutes or less, 20 minutes or less, 15 minutes or less, 10 minutes or less, 5 minutes or less, 4 minutes or less, 3 minutes or less , 2 mmutes or less, or 1 minute or less.
[00202] [00202] The time period of accepting an e sampl within the device for detecting a disease marker, or for detecting a plurality of disease markers, or for transmitting data about, and/or analyzing a sample or samples may depend on the type or number of steps, tests or assays performed on the sample or samples. The amount of time to accept a sample, or samples, into the device for detection of a disease marker or markers, or for transmission of data and/or for analysis from the device in relation to such a sample or samples may be approximately 3 hours or less, two hours or less, 1 hour or less, 50 minutes or less, 45 minutes or less, 40 minutes or less, 30 minutes or less, 20 minutes or less, 15 minutes or less, 10 minutes or less , 5 minutes or less, 4 mmutes or less, 3 minutes or less, 2 minutes or less, or 1 minute or less.
[00203] Thus, as used herein, a "short period of time" refers to a period of time of about 5 hours or less, or about 4 hours or less, or about 3 hours or less, or about 2 hours or less, or about 1 hour or less, or about 50 minutes or less, or about 40 minutes or less, or about 30 minutes or less, or about 20 minutes or less, or about of 10 mmutes or less, or about 5 minutes or less.. a short period of time can be determined relative to an initial time; the start time can be the time when analysis of a sample started; the start time can be the time when a sample is inserted into a device for sample analysis; the start time can be the time when a sample was taken from a subject.
[00204] [00204] The terms "Point of Service" (abbreviated POS) and "point of service system", as used herein, refer to a location, and a system in that location, which is capable of providing a sendee (eg, testing, monitoring, treatment, diagnosis, guidance, sampling, identity verification (ID verification), and other services) at or near the location or location of the object. A service can be a medical service, and it can be a non-medical service. In some situations, a POS system provides a service in a predetermined location, such as a subject's home, school or work or in a supermarket, a pharmacy, a community center, a clinic, a doctor's office, a hospital, etc. . a POS system can include one or more service points for addictions. In some embodiments, a POS system is a point of care system,
[00205] [00205] A. "point of care" (abbreviated POC) is a place where medical-related care (eg, treatment, testing, monitoring, diagnosis, counseling, etc.) is provided. The POC can be, for example, that of a subject at home, at work or at school, or in a supermarket, a community center, a pharmacy, a doctor's office, a clinic, a hospital, etc. A POC system is a system that can assist in, or can be used in, providing such related medical care, and can be located at or near the location or location of the subject or subject's healthcare provider (for example, at the subject's home, work or school, or to a grocery store, a community center, a pharmacy, a doctor's office, a clinic, a hospital, etc.).
[00206] As used herein, the term "immunoassay" refers to any assay that detects, identifies, characterizes, quantifies, or otherwise measures an amino acid target of a sample (wherein an amino acid target may be a small peptide, a polypeptide, a protein, or a protein macromolecule). Immunoassays include, for example, direct binding or competitive assays using techniques such as western blots, radioimmunoassays, ELISA (enzyme-linked immunosorbent assay),
[00207] As used herein, the term "nucleic acid assay" is used to refer to any assay that detects, identifies, characterizes, quantifies, or otherwise measures a target nucleic acid in a sample (wherein an acid target Nucleic acid can be a single stranded, double stranded, or other nucleic acid molecule of any size, nucleic acid assays include polymerase chain reaction (PGR) assays (see, for example, US Patent No. 4,683,202 ), isothermal cycle mediated amplification ("LAMP") (see, for example, US Patent No. 6,410,278), and other methods, including the methods discussed below for detecting nucleic acid targets in a sample. can be detected by any suitable means, including means including nucleic acid amplification (e.g., thermal cycling amplification methods including PGR, and other nucleic acid amplification methods, ist amplification methods. thermal, including the lamp, etc) and any other method that can be used to detect the presence of nucleic acid markers indicative of a disease-causing organism in a sample.
[00208] As used herein, the term "general chemistry assay" refers to any assay that detects, identifies, characterizes, quantifies, or otherwise measures a target in a sample, other than a target, which is an acid nucleic or other form that does not utilize an antibody or other protein that specifically binds. General chemistry tests include, for example, for electrolyte tests; for vitamin levels; for levels of blood components: of trace metals; for lipids; and other targets). General chemistry tests can include, for example, tests of a basic metabolic panel [glucose, calcium, sodium (Na), potassium (K), chloride (Cl), CO ,
[00209] As used herein the term "cytometry assay" refers to any that detects, identifies, characterizes, quantifies, or otherwise measures a cell or large particle (eg, a crystal) in a sample. Imaging cytometry assays typically use other light-based techniques to detect, measure, characterize and quantify cells and particles in a sample, Systems, devices and METHODS
[00210] In embodiments, applicants describe systems, including such systems discussed above, for detecting one or more of a plurality of disease-causing agents in a clinical sample. In embodiments, applicants describe systems, including such systems discussed above, for detecting one or more of a plurality of disease-causing agents in a clinical sample, wherein said diseases include respirator diseases. In embodiments, applicants describe systems, including such systems discussed above, for detecting one or more of a plurality of disease-causing agents in a clinical sample, wherein said respiratory disease-causing pathogens selected from viral diseases, bacterial diseases, fungal diseases, Mycoplasma diseases, and other diseases.
[00211] [00211] In embodiments, the plurality of disease-causing agents causes a number of diseases, wherein said number of diseases comprises 8 or more diseases, or 10 or more diseases, or 12. or more diseases, or 14 or more illnesses, or 16 or more illnesses, or 18 or more illnesses, or 20 or more illnesses, or 30 or more illnesses, or 40 or more illnesses, or 50 or more illnesses, or 60 or more illnesses, or more . In embodiments, the plurality of disease-causing agents causes a number of diseases, wherein said diseases are selected from viral diseases, bacterial diseases, fungal diseases, Mycoplasma diseases, and other diseases.
[00212] [00212] In embodiments, the systems described herein, as well as the methods described herein, can be used to perform ail of a plurality of assays with a single small volume sample, or an aliquot or aliquots thereof. In embodiments, a small volume of sample has a volume selected from no more than about 1 ml, or no more than about 500 µl, or no more than about 250 µl, or not more than about 150 iil, or no more than about 100 p, L, or no more than about 75 iil, or no more than about 50 µl., or no more than about 25 µl . ,, or not more than about 15 μΙ_, or not more than about 10 ,, or μΙ_ not more than about 5 μl ,, or not more than about 4 μl ,, or not more than about 5 μl ,, or not more than about 5 μl about 3 μl-, or no more than about 2 μl^,
[00214] [00214] In embodiments, the plurality of disease-causing agents can cause a number of diseases, wherein said disease-causing agents are selected from myeobacterium tuberculosis, Staphylococcus aureus (including methicillin-resistant Staphylococcus aureus), streptococci (including Group A Streptococcus and Group B Streptococcus), Bordetella pertussis, adenovirus (including adenovirus B, adenovirus C, and adenovirus e), influenza, parainfluenza, respiratory syncytial virus (SV), adenovirus, corona virus, bocavirus, Haemophilus parainfluenzae, human papilloma virus (HPV), hepatitis, human vims immunodeficiency (HIV), herpes simplex virus (HSV), West Nile virus, Epstein Barr virus, rhinovirus, and other viruses. In embodiments, the plurality of disease causing agents causes a number of diseases, wherein said disease causing agents are selected from Streptococcus, Staphylococcus, Bordetella pertussis, tuberculosis, enterobacteria, Pseudomonas, dengue, malaria , tiypanosome cruzi, Treponema pallidum, mycoplasma, chlamydia, Moraxella catarrhalis, Acinetobacter, legoneilla, cols Escherichia, Candida, chlamydia. Neisseria, Trichomonas, and other disease-causing agents, micro-organisms, das, including but not limited to disease-causing agents referred to elsewhere herein.
[00215] In embodiments, the plurality of disease causing agents can cause a number of respiratory diseases, wherein said number of respiratory diseases has 8 or more respiratory diseases, or 10 or more respiratory diseases, or 12 or more diseases or 14 or more respiratory diseases, or 16 or more respiratory diseases, or 18 or more respiratory diseases, or 2.0 or more respiratory diseases, or 30 or more respiratory diseases, or 40 or more respiratory diseases, or 50 or more respiratory diseases, or 60 or more respiratory diseases, or more.
[00216] [00216] In embodiments, the plurality of disease-causing agents can cause a number of respiratory diseases, wherein said respiratory diseases are selected from viral diseases, bacterial diseases, fungal diseases, Mycoplasma diseases, and other diseases .
[00218] [00218] In embodiments, the plurality of disease-causing agents can cause influenza. In embodiments, influenza may be selected from influenza A, influenza B, influenza II in I (including seasonal forms of influenza and novel H1 1), H3N2 influenza, H7N9 influenza, H5N1 influenza and others. flu.
[00219] [00219] In embodiments, the plurality of disease-causing agents can cause a sexually transmitted disease. In embodiments, the sexually transmitted disease is selected from herpes simplex virus (HSV), human immunodeficiency virus (HIV, including HlV-1, HIV-2, including HIV-2 Group A), gonorrhea, syphilis, virus of human papilloma (PPV), Streptococcus (including Streptococcus B), Treponema pallidum, and other sexually transmitted diseases.
[00220] [00220] Other targets include drug-resistant microorganisms, including those that exhibit multi-drag resistance. Drug resistance (also called antibiotic resistance) is found where a population (or subpopulation) of a microorganism, such as a bacterium, acquires resistance or exposures to one or more drugs (eg, one or more antibiotics). Micro-organisms that are resistant to multi-drug treatment are termed as "'multi-drug resistant" and micro-organisms are termed to have or exhibit "multi-drug resistance"; either term may be abbreviated as "MDR". Resistance to one or more drugs is observed, or exhibited, when a population of microorganisms survives (and typically continues to grow W and multiply in number) despite the presence of an obstacle, or (in the case of MDR) despite the presence of multiple drugs. drug resistant organisms of particular interest include, but are not limited to, methicillin resistant Staphylococcus aureus (MRSA), vancomycin intermediate S. aureus (VISA), vancomycin resistant S. aureus (VRSA), bacteria (eg, enterobacteria ) Having extended spectrum beta-lactamase (BLEE), resistant to vancomycin Enierococcus (VRE), and resistant to multiple drugs A. Bau iannii (MRAB). drug-resistant target organisms, including MDR target organisms, can be identified using nucleic acid markers,
[00221] [00221] Many antibiotic compounds include a β- lactam ring (a ring of four carbon atoms); for example, penicillin is an antibiotic with a β-lactam ring. Many bacteria have β-lactamase enzymes that can cleave a β-lactam ring, and thus protect bacteria against such antibiotics. Enzymes that can cleave a β- lactam ring, which are often found in Gram-negative bacteria, include the ETM and Rob β-lactamase enzymes. -1, although blaTEM-2 and a blaROB- and others are also found); other drug resistance markers available in disease-causing organisms include the KPC resistance gene (found in Klebsiella pneumonia (KPC) carhapenemas); mecA and MECC resistance genes (responsible for resistance to β-lactam containing antibiotics such as methicillin); vancomycin A and B resistance genes (vanA vanB e) can be found in disease-causing bacteria such as, for example, vancomycin-resistant enterococci; and other drag-resistance markers.
[00222] Disease-causing organisms of interest include viruses of the filoviridae family of viruses (phyllus virus), which include Ebola virus, Marburg virus, and Cueva virus. Ebola virus cause Ebola virus disease (also known as Ebola hemorrhagic fever); Marburg virus also cause hemorrhagic fever, Marburg hemorrhagic fever; and Cueva virus such as lloviu VIMS (LLOV), may be endemic in France, Spain or Portugal. Such viruses can be detected and can be identified in a sample from nucleic acid markers specific for these viruses, from protein (peptide) markers specific for these viruses, and from other markers specific for those viruses.
[00223] [00223] Phyllum viruses commonly cause hemorrhagic fevers and related illnesses. These and other diseases can be detected, and can be identified, by identifying nucleic acid markers, peptide markers, and other markers, alone or in combination, as disclosed herein. Phyllo viral diseases, other hemorrhagic fevers and other diseases , including many tropical diseases, for example Dengue 1, Dengue 2, Dengue 3, Dengue 4, malaria, typhoid and other diseases, have many harmful effects, including bleeding and internal bleeding, and can cause electrolyte disturbances, can cause anemia , and may cause other symptoms and effects. Integrated electrolyte assays (eg, for sodium, potassium, and other electrolytes, including sodium and potassium together, and including sodium and potassium together with other electrolytes) can identify individuals who suffer from imbalances. electrolytes, and can thus identify individuals suffering from hemorrhagic fever, anemia, or both. Tests for hemoglobin, iron and other tests can identify individuals suffering from anemia, from hemorrhagic fever, or both. Anemia can be due to bleeding, parasitic infection (eg, hookworm), both bleeding and parasitic infection, and other causes. Thus, in addition to testing for nucleic acid, peptide, and other markers, these hemorrhagic and other diseases can be detected, and can be identified, with integrated electrolyte measurements; with hemoglobin measurements; with iron measurements; and with other general chemistry tests, alone or in combination. In embodiments, measurements of integrated electrolytes, hemoglobin, iron, or other general chemistry assays can predict indications that a subject is suffering from anemia, hemorrhagic fever such as Ebola, Marburg, or another disease such as malaria or typhoid fever ; such indication can be used to suggest further tests for markers of such diseases. Thus, in embodiments, electrolyte testing including sodium and potassium, or other general chemistry tests, can be performed concurrently when testing for Ebola, Marburg, or other bleeding disorder (e.g., when testing nucleic acid, peptide, or other markers for such diseases).
[00224] [00224] In embodiments, such indications derived from general chemistry test results (for example, from the results of integrated measurements electrolytes, hemoglobin, iron, or other general chemistry tests) may automatically trigger reflex tests for one or more disease markers hemorrhagic fever, or malaria, typhoid, or other disease marker, and can automatically trigger reflex tests for combinations or for all of these markers. In embodiments, a subject may be tested for other diseases and not (initially) for Ebola, Marburg, or other bleeding disorder (e.g., a patient who is weak, or febrile, cannot initially be tested for the Ebola, Marburg, or other hemorrhagic disease); based on the results of integrated measurements of electrolytes, hemoglobin, iron, or other general chemistry tests, a reflex test for Ebola, Marburg, or other hemorrhagic disease can be performed automatically, or can be ordered by a healthcare professional. In embodiments, a patient or individual may not wish to be tested for Ebola, Marburg, or other bleeding disorder, or other disease of interest (e.g., a patient may fear quarantine, or may fear that the cost of the assay may be prohibitive ) and therefore cannot initially be tested for Ebola, Marburg, or other bleeding disorders; based on the results of integrated measurements of electrolytes, hemoglobin, iron, or other general chemistry tests, a reflex test for Ebola, Marburg, or other hemorrhagic disease can be performed automatically, or can be ordered by a healthcare professional. In embodiments, such a reflex test for Ebola, Marburg, or other bleeding disease, or other diseases in question may be performed if the patient is first tested for one or more infectious diseases other than Ebola, Marburg, or other bleeding disease , and the initial test panel does not show the infection of any known disease (and therefore further testing would be needed in order to identify other possible sources of the individual's disease or condition).
[00225] The methods, devices and systems described herein can be used to detect, and can be used to identify, disease-causing organisms in normal or healthy individuals and populations.
[00226] [00226] These disease-causing organisms and such benign organisms can be detectable, and can be identified, in samples obtained from several individuals, for example, once or on a continuous basis, in order to determine a baseline or normal level found in a normal (healthy) population. Such detection of causes of benign diseases and organisms may include detection of nucleic acid markers; detection of protein (peptide) markers, including detection of antibodies; detection of inflammation markers (including both peptide and non-peptide markers); and detection of other markers. This identification of benign disease causers and organisms can include the identification of nucleic acid markers; the identification of protein (peptide) markers, including the identification of antibodies; the identification of inflammation markers (including both peptide and non-peptide markers); and identification of other markers. Detection and identification of markers may include quantitative detection and identification of such a marker. Differences between results obtained from an individual subject sample and normal baseline results or obtained from a population of comparable healthy individuals can be used to improve the probability of detecting whether or not an individual subject suffers. of a disease or condition. For example, the determination of an initial or a normal level for an individual subject aids in detecting, identifying and diagnosing disease conditions or progression to a disease or detrimental condition by comparing results for the individual subject to baseline levels. or normal found on a. population of comparable healthy individuals. Such comparisons between an individual subject's results and baseline or normal levels found in a healthy population can be used to determine whether the individual INDI subject is likely to suffer from an infection. Such comparisons might include analysis of symptoms, if any, of the individual subject compared to the symptoms (or lack thereof) found in a healthy population.
[00227] [00227] In embodiments, such a system is a point-of-service system (POS system), wherein a POS system is located at a service location point. In embodiments, a POS system is located at a service location point and is configured to accept a clinical sample obtained from an individual at the POS location. In embodiments, a POS system is located at a service location point and is configured to accept a clinical sample obtained from an individual at the POS location, and is further configured to analyze the clinical sample at the POS location. In embodiments, the sample is a small volume clinical clinical sample. In embodiments, the clinical sample is analyzed in a short period of time. In embodiments, the short period of time is determined in relation to the time at which the analytical sample started. In embodiments, the short period of time is determined with respect to when the sample was inserted into a device for analyzing the sample. In embodiments, the short period of time is determined in relation to when the sample was obtained from the subject.
[00229] [00229] Applicants disclose methods of the present invention for detecting the presence of a target influenza VIMs molecule in a sample are described herein, wherein the presence of a plurality of possible target influenza viruses are tested from a single sample within a short period of time. In embodiments, the plurality of target possible influenza viruses comprise at least 5 possible target influenza viruses, or at least 10 possible target influenza viruses, or at least 15 possible target influenza viruses, or at least 20 possible target flu viruses, or at least 25 possible target influenza viruses, or at least 30 possible target influenza viruses, or at least 35 possible target influenza viruses, or at least 40 possible target influenza viruses, or at least 45 possible target influenza viruses, or at least 50 possible target influenza viruses, or at least 55 possible target influenza viruses, or at least 60 possible target influenza viruses, or at least 64 possible target influenza viruses, or at least 65 possible target flu viruses, or more.
[00230] Applicants herein further describe methods for detecting the presence of a plurality of target molecules in a single sample within a short period of time, wherein the plurality of target molecules comprises nucleic acid molecules, or protein molecules, or saccharides, or cytokines, or steroids, or histamine, or other molecules. Applicants further describe methods of the present invention for detecting the presence of a plurality of target molecules in a single sample within a short period of time, wherein the plurality of target molecules comprises nucleic acid molecules and the protein molecules. Appiicanis further discloses herein methods for detecting the presence of a plurality of target molecules in a single sample within a short period of time, wherein the plurality of target nucleic acid molecules comprises molecules, protein molecules, inflammation markers and cytokines. Applicants further describe methods of the present invention for detecting the presence of a plurality of target molecules in a single sample within a short period of time, wherein the plurality of target molecules comprises nucleic acid molecules, protein molecules and saccharides.
[00231] [00231] Applicants disclose devices described herein for use in systems and methods as disclosed herein. Such devices include, for example, devices that comprise a holder configured to accept and retain a clinical sample (e.g., a clinical sample contained within a a sampling device); a reagent container or a plurality of reagent vessels; and a reaction vessel, or a plurality of reaction vessels. In embodiments, such devices can be further configured to accept and retain one or more of a cytometry cuvette or cuvettes; a waste container or other containers; a tip or tips, set to aspirate or release fluid; and other tools.
[00232] Applicants further describe herein assays for detecting one or more of a plurality of target molecules in a single sample within a short period of time, wherein the plurality of target molecules comprises one or more of the target molecules. nucleic acid, protein molecules, saccharides, inflammation markers and cytokines. In embodiments, such assays can be configured for use with systems and devices as disclosed herein.
[00233] Accordingly, applicants disclose systems, devices and methods herein, including the following examples of integrated systems.
[00234] [00234] 1) An integrated system for providing tests and diagnosis of an individual suspected of suffering from a disease, said system comprising a means for obtaining a sample (which may include, for example, a sample collection device comprising a lancet, a syringe, a needle and tube, or other blood collection device, or a nasal swab, a mouth swab (eg, a cotton swab), a throat swab, a vaginal swab, or other swab , and fluid in which to immerse the swab following contacting the swab with an object): a cartridge comprising reagents for assays for disease; a device for carrying out a plurality of assays for detecting a plurality of diseases; means a device for displaying / communicating the detection of one or more of said diseases. Such integrated systems can be configured for uses where the sample is a small volume sample; for uses where detection is performed in a short period of time; or for use both where the sample is a small volume sample and where detection is performed in a short period of time.
[00235] [00235] 2) An integrated system for providing testing and diagnosis of an individual suspected of suffering from a respiratory disease, said system comprising a means for obtaining a sample (which may include, for example, a nasal swab, a swab a throat, mouth swab (eg, a face swab), a vaginal swab, or other swab, and in which the fluid to immerse the next swab contacts the swab with a subject); a cartridge comprising the reagents for testing for respiratory disorders; a device for carrying out a plurality of tests for detecting a plurality of respiratory diseases; a device/means for the display to communicate the detection of one or more of said respiratory disorders. Such integrated systems can be configured for uses where the sample is a small volume sample; for uses where detection is performed in a short period of time; or for use both where the sample is a small volume sample and where detection is performed in a short period of time.
[00236] [00236] 3) An integrated system for providing testing, diagnosis and prescription of an individual suspected of suffering from a respiratory disease, said system comprising a means for obtaining a sample (which may include, for example, a nasal swab, a throat swab, a mouth swab (eg, a face swab), a vaginal swab, or other swab, and wherein the fluid for dipping the next swab contacts the swab with a subject); a cartridge comprising the reagents for testing for respiratory disorders; a device for carrying out a plurality of tests for detecting a plurality of respiratory diseases; a device/means for displaying/communicating the detection of one or more of said respiratory disorders; and means for providing a prescription for the treatment of a respiratory disorder detected in said sample. Such integrated systems can be configured for uses where the sample is a small volume sample; for uses where detection is performed in a short period of time; or for use both where the sample is a small volume sample and where detection is performed in a short period of time.
[00237] [00237] 4) An integrated system for providing testing, diagnosis, prescription and treatment of an individual suspected of suffering from a respiratory disease, said system comprising a means for obtaining a sample (which may include, for example, a cotton swab nasal, throat swab, a mouth swab (eg, a face swab), a vaginal swab, or other swab, and in which the fluid to immerse the next swab contacts the swab with a subject); a cartridge comprising the reagents for testing for respiratory disorders; a device for carrying out a plurality of tests for detecting a plurality of respiratory diseases; a device/means for displaying/communicating the detection of one or more of said respiratory disorders; means for providing a prescription for the treatment of a respiratory disorder detected in said sample; and means for providing/selling/delivering a treatment (drug/pill/shot) to said subject, pursuant to said prescription. Such integrated systems can be configured for uses where the sample is a small volume sample; for uses where detection is performed in a short period of time; or for use both where the sample is a small volume sample and where detection is performed in a short period of time.
[00238] [00238] Methods for detecting the presence of a target influenza vims molecule in a sample are described herein, wherein the presence of a plurality of possible target influenza viruses are tested from a single sample within a short period of time. . In embodiments, the plurality of possible target flit viruses comprise at least 5 possible target influenza viruses, or at least 10 possible target influenza viruses, or at least 15 possible target influenza viruses, or at least 20 influenza viruses. possible target influenza, or at least 25 possible target influenza viruses, or at least 30 possible target influenza viruses, or at least 35 possible target influenza viruses, or at least 40 possible target influenza viruses, or at least 45 influenza viruses possible target influenza, or at least 50 possible target influenza viruses, or at least 55 possible target influenza viruses, or at least 60 possible target influenza viruses, or by the feast 64 possible target influenza viruses, or at least 65 influenza viruses possible target flu, or more. In embodiments, a short period of time is a period of time that is IVF and hours or less, or is four hours or less, or is three hours or less, or is two hours or less, or is an hour or less, or it's 50 minutes or less, or it's 40 minutes or less, or it's 30 minutes or less, or it's 20 minutes or less, or it's 10 minutes or less, or it's 5 minutes or less.
[00239] [00239] Methods for detecting the presence of a respiratory disease causing agent in an individual suspected of having a respiratory disorder, wherein the presence of a plurality of possible respiratory disease causing agents are tested from a single sample using two nucleic acid assays and protein assays, wherein the nucleic acid assay comprises detecting the presence of target nucleic acid sequences, and wherein the protein comprises assaying for detecting the presence of target proteins that have target amino acid sequences. In embodiments, target nucleic acid sequences may comprise sequences that are at least 8 nucleotides, or at least 10 nucleotides, or at least 15 nucleotides, or at least 20 nucleotides, or at least 30 nucleotides, or at least 40 nucleotides , or at least 50 nucleotides, or more, that are identical or very similar to target nucleotide sequences. In embodiments, target amino acid sequences may comprise sequences that are at least 8 amino acids, or at least 10 amino acids, or at least 15 amino acids, or at least 20 amino acids, or at least 30 amino acids, or at least 40 acidic amino acids. , or at least 50 amino acids, or more, which are identical or very similar with the target amino acid sequences.
[00240] [00240] In embodiments, a respiratory disease causing agent is detected if more than a minimum level of such respiratory disease causing agents is detected in a small sample - VOLUME obtained from a subject, where the small volume sample is tested for the presence of a plurality of causative agents of respiratory diseases. In embodiments, the small volume sample is 150 μΙ_ or less in terms of volume, ie 75 μl , or less in terms of volume, ie 50 , ul or less in terms of volume, or is 25 iL or less in terms of volume, ie 15 } iL or less in terms of volume, or is 10 p, L or less of volume, ie 5 μΕ or less in volume.
[00241] [00241] Embodiments of the methods described herein include methods in which a disease-causing agent is detected if more than a minimal level of such disease-causing agents is detected in a small volume sample obtained from a subject. For example, such methods include methods in which a small volume sample is tested for the presence of a plurality of disease-causing agents. In embodiments of such methods, what minimum level can be set to a level that is determined by the state of the subject. The minimum level can be set to a higher level for individuals who show symptoms of active infection. The minimum level can be set at a lower level for individuals who are receiving treatment for an infection at the time of sampling. time of taking the sample, for example, who have recently been treated for an infection before the time of taking the sample.
[00242] [00242] In embodiments, the sample may be diluted before being tested for the presence of a plurality of disease-causing agents. In embodiments, such a sample dilution is greater for individuals who have a condition indicating that they may have higher levels of disease-causing agents than subjects who do not have that condition, or than individuals who have a different condition.
[00243] [00243] Embodiments of the methods described herein include methods in which a respiratory disease causing agent is detected if more than a minimal level of such respiratory disease causing agents is detected in a small volume sample obtained from a subject . For example, such methods include methods in which a small volume sample is tested for the presence of a plurality of respiratory disease causing agents. In embodiments of such methods, what minimum level can be set to a level that is determined by the state of the subject. The minimum level can be set to a higher level for individuals who show symptoms of active infection. The minimum level can be set at a lower level for individuals who are receiving treatment for an infection at the time of obtaining the sample. The minimum level can be set at a lower level for individuals who have received treatment for an infection before the time of taking the sample, for example, who have recently received treatment for an infection before the time of taking the sample.
[00244] [00244] In embodiments, the sample may be diluted before being tested for the presence of a plurality of disease-causing agents, such as respiratory disease-causing agents. In embodiments, such a sample dilution is higher in individuals who have a condition that indicates they may have higher levels of disease-causing agents, such as respiratory disease-causing agents, than subjects who do not have that condition, or than individuals who have a different condition.
[00245] For example, conditions that indicate that a subject may have higher levels of disease-causing agents, such as respiratory disease-causing agents, including individuals with an active infection; individuals who have a cough, including a persistent cough; individuals who have a fever; individuals who report chills; individuals who report fatigue; individuals who report a headache; individuals who have sweats; and individuals who have or report other symptoms indicative of an active infection.
[00246] For example, conditions that indicate that a subject may not have higher levels of disease-causing agents, such as respiratory disease-causing agents, include subjects currently receiving treatment for the infection; subjects who have recently received treatment for the infection; subjects currently receiving, or who have recently received treatment for, a cough, including a persistent cough; one; chills; fatigue; headache; sweats; or other symptoms, or an indication of an infection.
[00247] [00247] In embodiments of methods where a disease-causing agent, such as a respiratory disease-causing agent, is detected, if more than a minimum level of such disease-causing agent (e.g., a disease-causing agent respiratory disease) is detected in a small volume sample obtained from a subject, where the small volume sample is tested for the presence of a plurality of disease causing agents, such as respiratory disease causing agents, the minimum level is set at a higher level for individuals who have not been recently diagnosed with a disease, such as a respiratory disease, than the minimum level set for individuals who are newly diagnosed with a disease (such as a respiratory disease), in embodiments , the sample may be diluted prior to testing to detect the presence of said plurality of disease-causing agents, such as disease-causing agents. respiratory diseases. In embodiments, such a sample dilution is greater for individuals who have not recently been diagnosed with a disease, such as a respiratory disease, than the dilution of samples obtained from individuals who have recently been diagnosed with a disease, such as as a respiratory disease.
[00248] [00248] In embodiments, the sample can be further tested for the presence of indicators of inflammation. For example, the sample can be further tested for the presence of above-normal levels of glucocorticoids such as cortisol (e.g., in blood or saliva, tears, or other body fluid or sample obtained by a swab). For example, the sample can be further tested for the presence of above-normal levels of histamine (for example, in blood or saliva, tears, or other body fluid or swab-obtained samples). Other indicators of inflammation include, without limitation, increased levels of prostaglandins, increased levels of inflammatory cytokines (including, for example, tumor necrosis factor alpha (TNF-a), interleukin-1 (IL-1), interleukin- 8 (IL-8 ), interleukin-12 (IL-12), interleukm-18 (IL-18), and interferon gamma (IF-γ)), bradykinin, complement system molecules, blood clotting factors, C-reactive protein, erythrocyte sedimentation rate (ESR), white blood cell count, morphological changes in blood and other cells, and other molecular and cellular markers indicative of inflammation.
[00249] In embodiments, the sample can be further tested for the presence of a cytokine or a plurality of cytokines. In embodiments, the sample can be further tested for the level of a cytokine or a plurality of cytokines. In embodiments, the target cytokine is selected from a lymphokine, a chemokine, an mterleukin, and an interferon. In embodiments, target cytokines are selected from lympliokines, chemokines, inter-leukms, and interferons. In embodiments, the cytokine can be an inflammatory cytokine. Within embodiments, the cytokine can be an anti-inflammator cytokine. In embodiments, a target cytokine can be an mterleuki (IL) selected from IL-1, IL-2, IL-3, IL-4, 1L-5, 1L-6, 1L-7, ίΙ, -8, ίί ., - 9, IL-10, ii .- i 1, IL-12, IL-13, 1L-I4, IL-15, IL-1 6, IL-17, IL-1 8, 1L - 19, 11 .-
[00250] [00250] In embodiments, the same sample can be assayed for disease-causing agents and for cytokines. In embodiments, the same sample can be assayed for agents causing respiratory diseases and for cytokines.
[00251] In embodiments, the sample can be further tested for the presence of antibodies against a disease-causing agent. In embodiments, the same sample can be tested for a plurality of disease-causing agents, and for antibodies to a plurality of disease-causing agents.
[00252] In embodiments, the sample can be further tested for the presence of antibodies against a causative agent of respiratory disease. In embodiments, the same sample can be tested for a plurality of respiratory disease causing agents and for antibodies to a plurality of respiratory disease causing agents.
[00253] [00253] Methods for detecting the presence of a target influenza virus molecule in a sample are described herein, wherein the presence of a plurality of possible target influenza viruses are tested from a single sample using two acid tests nucleic acid and protein assay, wherein the test nucleic acid comprises detecting the presence of target nucleic acid sequences, and wherein the protein comprises assaying for detecting the presence of target proteins that have target amino acid sequences. In embodiments, target nucleic acid sequences may comprise sequences that are at least 8 nucleotides, or at least 10 nucleotides, or at least 15 nucleotides, or at least 20 nucleotides, or at least 30 nucleotides, or at least 40 nucleotides , or at least 50 nucleotides, or more, that are identical or very similar to target nucleotide sequences. In embodiments, target amino acid sequences may comprise sequences that are at least 8 amino acids, or at least 10 amino acids, or at least 15 amino acids, or at least 20 amino acids, or at least 30 amino acids, or at least 40 acidic amino acids. , or at least 50 amino acids, or more, which are identical or very similar with the target amino acid sequences.
[00254] In embodiments, the plurality of possible influenza viruses t Arget comprise at least 5 possible target influenza viruses, or at least 10 possible target influenza viruses, or at least 15 possible target influenza viruses, or at least 20 possible target influenza viruses, or at least 25 possible target influenza viruses, or at least 30 possible target influenza viruses, or at least 35 possible target influenza viruses, or at least 40 possible target influenza viruses, or at least 45 possible target influenza viruses, or at least 50 possible target influenza viruses, or at least 55 possible target influenza viruses, or by the feast 60 possible target influenza viruses, or at least 64 possible target influenza viruses, or at least 65 possible target influenza viruses, or more.
[00255] [00255] The assays and methods described here can be performed on a device, or by a system, for processing a sample. Such a sample may be a small volume clinical sample. The assays and methods described herein can be easily incorporated into and used in the device for processing a sample, or a system for processing a sample, which can be an automated testing device, or it can be an automated testing system. . Such a device, and such a system, can be useful for practicing the methods disclosed herein. For example, a device can be useful for receiving a sample, a device can be useful for preparing or processing a sample. device can be useful for performing a test on a sample. A device can be useful for obtaining data from a sample. A device can be useful for transmitting data obtained from a sample. A de-vice can be useful for removing a sample after processing or testing a sample.
[00256] [00256] The tests and methods described herein can be easily incorporated into and used in an automated testing device,
[00257] [00257] A device can be part of a system, a component of which can be a sample processing device. A device can be a sample processing device. A sample processing device can be configured to facilitate taking a sample, preparing a sample for testing at a clinic, or performing a chemical reaction with one or more reagents or other chemical or physical treatment, as disclosed herein. A sample processing device can be configured to obtain data from a sample. A sample processing device can be configured to transmit data obtained from a sample. A sample processing device can be configured to analyze data from a sample, a sample processing device can be configured to communicate with another device, or by a laboratory, or an individual affiliated with a laboratory, for analysis of data obtained from a sample.
[00258] [00258] A sample processing device can be configured to be placed inside or over an object. A sample processing device can be configured to accept a sample from a subject, either directly or indirectly. A sample can be, for example, a blood sample (for example, a sample obtained from a finger prick, or a venipuncture, or an arterial blood sample), a urine sample, a biopsy sample, a tissue slice, stool sample, or other clinical sample; a water sample, a soil sample, a food sample, an air sample; or other sample. A blood sample can comprise, for example, whole blood, plasma, or serum. A sample processing device can receive a sample from the subject via a housing of the device. Sample collection can take place at a sample collection site, or elsewhere. The sample can be supplied to the device at a sample collection site.
[00259] [00259] In some embodiments, a sample processing device may be configured to accept or realize a cartridge. In some embodiments, a sample processing device can comprise a cartridge. The cartridge is removable from the sample processing device. In some embodiments, a sample may be supplied to the sample processing device cartridge. Alternatively, a sample may be supplied to another portion of a sample processing device. The cartridge and/or deputy can comprise a sample collection unit that can be configured to accept a sample.
[00260] [00260] A cartridge may include a sample, and may include reagents to be used in treating or testing a sample, disposables to be used in treating or testing a sample, or other materials. Upon placement of a cartridge into, or insertion of a cartridge into, a sample processing device, one or more components of the cartridge may be brought into fluid communication with other components of the sample processing device. For example, if a sample is collected on a cartridge, the sample can be transferred to other portions of the sample processing device. Likewise, if one or more reagents are provided in a cartridge, the reagents can be transferred to other portions of the sample processing device, or other components of the sample processing device can be brought into the reagents. In some embodiments, reagents or components of a cartridge may remain on the edge of the cartridge, in some embodiments, no ffuidics are included that require tubes or that require maintenance (e.g., manual or automated maintenance).
[00261] [00261] A. sample or reagent may be transferred to a device such as a sample processing device. A sample or reagent can be transferred within a device. Such sample or reagent transfer can be accomplished without providing a continuous fluid path from the cartridge to the device. Such sample or reagent transfer can be accomplished without providing a continuous fluid path within a device. In embodiments, such sample or reagent transfer may be performed by a sample handling system (e.g., a pipette); For example, a sample, reagents, or an aliquot thereof can be aspirated with a tip opening transfer component, such as a pipette tip, which can be operably linked to a sample handling system that transfers the tip, with the sample, reagents, or aliquot of the latter contained within the tip, to a location on or within the sample processing device. The sample, reagents, or even an aliquot can be deposited at a location on or within the sample processing device. Sample and the reagent, or several reagents, can be mixed using a sample handling system in a similar manner. One or more components of the cartridge can be transferred in an automated manner to other portions of the sample processing device, and vice versa.
[00262] [00262] A device, such as a sample processing device, may have a fluid handling system. A fluid handling system can perform, or can assist in performing, transport, dilution, extraction, aliquoiting, mixing, and other actions with a fluid, such as a sample. In some embodiments, a fluid handling system may be contained within a housing of the device, a fluid handling system may allow for the collection and delivery, processing and/or transport of a fluid, dissolution of dry reagents. , mixing the liquid and/or dry reagents with a liquid, as well as the collection and delivery, processing and/or transportation of non-fluidic components, samples, or materials. The fluid can be a sample, a reagent, diluent, wash, dye, or any other fluid that can be used by the device, and can include, but are not limited to, homogeneous liquids, different liquids, emulsions, suspensions, and other fluids. . A fluid handling system, including, without limitation, a pipette, may also be used to transport containers (with or without fluid contained therein) around the device. The fluid handling system may dispense or aspirate a fluid. The sample can include one or more particles or solid matter floating within a fluid.
[00263] [00263] In embodiments, a SY fluid handling rod may comprise a pipette, pipette tip, syringe, capillary, or other component. The fluid handling system may have a portion with an inner surface and an outer surface and an open end. The fluid handling system may comprise a pipette, which may include a pipette body and a pipette nozzle, and may comprise a pipette tip. A pipette tip may or may not be removable from a pipette nozzle. In embodiments, a fluid handling system can use a pipette coupled with a pipette tip; a tip can be disposable. The tip can form a fluid tight seal when coupled with a pipette. A pipette tip can be used once, twice or more times, in embodiments, a fluid handling system can use a pipette or the like of addiction, with or without a pipette tip, to aspirate, dispense, mix , transport, or otherwise manipulate the fluid. Fluid can be dispensed from the fluid handling system when desired. Fluid can be contained within a pipette tip before being dispensed, for example, from a hole in the pipette tip. In embodiments, or during use cases, all of the liquid may be dispensed; In other embodiments, or instances during use, a portion of the fluid within a tip may be dispensed. A pipette can selectively aspirate a fluid. The pipette can aspirate a selected amount of fluid. The pipette may be capable of actuating stirring mechanisms to mix the liquid within the tip or within a vessel. The pipette can incorporate tips or vessels to create continuous flow loops for mixing, including materials or reagents that are in non-liquid form. A pipette tip can also facilitate mixing by controlled delivery of multiple fluids simultaneously or in succession, as in 2-piece substrate reactions.
[00264] [00264] The fluid handling system may include one or more independent hydraulicaliv or fluidically isolated units. For example, the fluid handling system can include one, two, or more pipette tips. Pipette tips can be configured to accept and confine a fluid. The tips can be fluidly insulated from or hydraulically independent of each other. The fluid contained within each tip may be hydraulically isolated or fluidly independent of one another in fluid tips and other fluids within the device. The fluidly isolated or hydraulically independent units can be movable relative to other portions of the vice and/or each other. The independent units are fluidly isolated or can be individually hydraulically movable. A fluid handling system can comprise one or more base or support. A base or support can support one or more pipette or pipette units. A base or support can connect one or more pipettes of the fluid handling system with one another.
[00265] [00265] A sample processing device may be configured to perform processing steps or actions on a sample obtained from a subject. Sample processing may include sample preparation, including, for example, sample dilution, aliquoting a sample, extraction, contact with a reagent, filtration, separation, centrifugation, or other preparatory action, transformation or step. A sample processing device can be configured to perform one or more sample preparation action or step on the sample. Optionally, a sample can be prepared by a chemical reaction and/or physical processing step. The sample or step preparation action may include one or more of the following procedures: centrifugation, separation, filtration, dilution, enrichment, purification, precipitation, incubation, pipetting, transport, chromatography, cell lysis, cytometry, spraying, milling , activation, uitrasonication, micro column processing, magnetic bead processing, nanoparticle processing, or other sample action preparation or steps. For example, sample preparation can include one or more steps to separate blood into serum and/or particle fractions, or to separate any other sample into multiple components. Sample preparation may include one or more steps of diluting and/or concentrating a sample, such as a blood sample, or other clinical samples. Sample preparation may include the addition of an anticoagulant or other ingredient to a sample. Sample preparation can also include purification of a sample. In embodiments, all sample processing, preparation or testing actions or steps are performed by a single device. In embodiments, all sample processing, preparation or testing actions or steps are performed within a single device housing. In embodiments, most sample processing, preparation or assay actions or steps are performed by a single device, and may be performed within a single device housing. In embodiments, many of sample processing, preparation or testing actions or steps are performed by a single device, and can be performed within a single vice housing. In embodiments, sample processing, preparation or testing actions or steps may be performed by more than one device.
[00267] [00267] A sample processing device can be configured to perform a plurality of assays on one sample. In embodiments, a sample processing device may be configured to run a plurality of assays on a single sample. In embodiments, a sample processing device can be configured to perform a plurality of assays on a single sample, where the sample is a small sample. For example, a small sample must have a sample volume that is a small volume of less than about 1000 μΕ, or less than about 500 μΕ, or less than about 250 μΕ, or less than about 150 Ah, or less than about 100 μl , or less than about 75 ΐ ^ , or less than about 50 μl , or less than about 40 μl , or less than about 20 μl. , or less than about 10 il, or other small volume. A sample processing device may be capable of performing multiplex assays on a single sample, a plurality of assays may be performed simultaneously; can be run sequentially; or some tests can be run simultaneously,
[00268] [00268] In embodiments, all of a plurality of trials can only be performed in a short period of time. In embodiments, such a short time period comprises less than about three hours, or less than about two hours, or less than about an hour, or less than about 40 minutes, or less than about 30 minutes, or less than about 25 minutes, or less than about 20 minutes, or less than about 15 minutes, or less than about 10 minutes, or less than about 5 minutes, or less than about 4 minutes, or less than about 3 minutes, or less than about 2 minutes, or less than about 1 minute, or other short period of time.
[00269] [00269] A sample processing device can be configured to detect one or more signals related to the sample. A sample processing device can be configured to identify one or more properties of the sample. For example, the sample processing device can be configured to detect the presence or concentration of an analyte or a plurality of analytes or a disease condition in the sample (e.g., in or through a bodily fluid, secretion, tissue, or otherwise sample). Alternatively, the sample processing device may be configured to detect a signal or signals which may be analyzed to detect the presence or concentration of one or more analytes (which may be indicative of a disease condition) or a disease condition. in the sample. Signals can be analyzed on board the device, or at another location. Running a clinical test may or may not include any analysis or comparison of the collected data.
[00270] [00270] A chemical reaction or other processing step can be performed, with or without the sample. Examples of steps, tests, or assays that can be prepared or administered by the device may include, but are not limited to, immunoassay, nucleic acid assay, receptor-based assay, cytometric assay, colorimetric assay, enzyme assay, electrophoretic assay , electrochemical assay, spectroscopy assay, chromatography assay, microscopic assay, topographic assay, calorimetric assay, turbidmefric assay, agglutination assay, radioisotope assay, viscosimetric assay, coagulation assay, coagulation time assay, synthesis assay protein, histological assay, culture assay, osmotic pressure assay, and/or other types of assays, centrifugation, separation, filtration, dilution, enrichment, purification, precipitation, spraying, incubation, pipetting, transport, cell lysis, or other sample preparation action or steps, or combinations thereof. Steps, tests, or assays that can be prepared or performed by the device can include imaging, including microscopy, cytometry, and other techniques that prepare or use imaging. Steps, tests or assays that can be prepared or performed by the device may further include an assessment of the histology, morphology, kinematics, dynamics and/or state of a sample, which may include such assessment for cells.
[00272] [00272] A device can be configured to prepare a sample for disposal, or to dispose of a sample, such as a clinical sample, after processing or testing a sample.
[00273] [00273] In embodiments, a sample processing device may be configured to transmit data obtained from a sample. In embodiments, a sample processing device may be configured to communicate over a network. A. Sample processing device can include a communication module that can interact with the network. A sample processing device can be connected to the network via a wired or wireless connection. The network can be a local area network (LAN) or a wide area network (WAN), such as the Internet. In some embodiments, the network can be a personal area network. The network can include the cloud. The sample processing device can be connected to the network without the need for a vice intermediary, or an intermediary device may be required to connect a sample processing device to a network. A sample processing device can communicate over a network with another device, which can be any type of network device, including but not limited to a personal computer, server compitter or laptop computer; personal digital assistants (PDAs) as a Windows CE device; phones, such as cell phones, smartphones (eg iPhone, Android, Blackberry, etc.), or location-aware, handheld phones (such as GPS); a mobile device, such as a network-connected roaming device; a wireless device, such as a wireless electronic device or any other device capable of wirelessly communicating with a computer network; or any other type of network device that can possibly communicate or view a network and handle electronic transactions. This communication may include providing data to a cloud computing infrastructure or any other type of data storage infrastructure that can be accessed by other devices.
[00274] [00274] A sample processing device can provide data about a sample from, for example, a healthcare professional, a healthcare professional location such as a laboratory, or an affiliate thereof. One or more of a laboratory, healthcare professional, or subject may have a network device capable of receiving or accessing data provided by the sample processing device. A sample processing device can be configured to provide data regarding a sample from a database. A sample processing device can be configured to provide data regarding a sample from an electronic medical record system, a laboratory information system, a laboratory automation system, or other system or software. A sample processing device can provide data in the form of a report.
[00275] [00275] A. laboratory, device, or other entity or software may perform analysis on data about a sample in real time. A software system may perform chemical sis analysis and/or pathological analysis, or these may be distributed among combinations of laboratory, clinical, and specialty personnel or experts. Analysis may include qualitative and/or quantitative assessment of a sample. Data analysis may include qualitative and/or quantitative assessment of a subsequent sample. Optionally, a report can be generated based on raw data, pre-processed data, or analyzed data. This report can be prepared so as to maintain the confidentiality of the data obtained from the sample, the identity and other information relating to the subject from which a sample was obtained, the analysis of the data, and other confidential information. The report and/or data can be transmitted to a healthcare professional. Data obtained by a sample processing device, or analysis of that data, or reports, may be provided to a database, an electronic medical records system, a laboratory information system, a laboratory automation system , or other system or software.
[00276] Markers indicative of a disease, such as a respiratory disease, include nucleic acid markers. Such nucleic acid tags include, for example, viral nucleic acids, or parts thereof; bacterial nucleic acids, or portions thereof; and nucleic acids, or parts thereof, derived from other microorganisms. Methods for detecting nucleic acid markers in a sample, including a small volume sample, include methods in which small amounts of nucleic acid can be amplified (e.g., copies made). For example, polymerase chain reaction (PCR) and related methods are common nucleic acid amplification methods. PCR is discussed, for example, in Pat. No. 4,683,195; and, in general, Mullis et al, Cold Spring Harbor Symposium on Quantitative Biology, Volume 51: 263 (1987); Eriich, ed., PCR Technology (Stockton Press, New York, 1989). PCR is one, but not the only, example of a nucleic acid polymerase reaction method for amplifying a nucleic acid test sample comprising using a known nucleic acid as a primer and a nucleic acid polymerase to amplify or generate a specific piece of nucleic acid. Further discussion of PCR and other methods can be found, for example, in the manual by Moleeuj AR Clonin A Laboratory by Green and Sambrook, Cold Spring Harbor Laboratory Press, 4th edition 2012, which is incorporated herein by reference in its entirety. PCR and many other amplification methods must be performed at several different temperatures, requiring temperature changes used during PCR. reaction ("thermal cycles"). Other amplification methods, such as, for example, isothermal cycle-mediated ("LAMP") amplification (see, for example, US Patent No. 6,410,278), and other methods, including the methods discussed below, require less or less extensive than that of PCR thermal cycling, or do not require thermal cycling.
[00277] [00277] Methods for nucleic acid amplification that do not require thermal cycling are described in United States Patent Application 61/800,606, filed March 15, 2013, incorporated herein by reference in its entirety. Such methods can be used to detect nucleic acid markers of disease, such as respiratory disease, in small volume samples in short periods of time. Such methods are discussed below, and examples of results obtained with such methods, from small samples and over short periods of time, are presented in the Figures and Examples described herein. In the following, such methods are referred to as "non-cycling amplification methods".
[00278] [00278] amplification methods without the nucleic acid amplification cycle can be applied to double stranded DNA A. However, as a target nucleic acid molecules need not be limited to double stranded DNA targets; for example, double-stranded DNA for use in non-cycling applications. The amplification methods described herein can be prepared from viral RNA, or mRNA, or other single-stranded RNA target sources, by reverse transcriptase. In another example, double-stranded DNA for use in the non-cycling amplification methods described herein can be prepared from single-stranded DNA targets by a DN polymerase. Such methods can be applied as an initial step, before applying the non-cycling amplification methods discussed below,
[00279] [00279] Amplification of a double-stranded DNA target, for example, starts with a primary double-stranded DNA to be amplified (referred to as the "primary nucleic acid" in the following). Primary nucleic acid contains a target region called a template region; the template region as a template sequence. Such a template double-stranded region contains a first strand of DNA and a complementary second strand of DNA, and includes terminal nucleotides on one strand and a 3' to 5 terminal nucleotides on the other strand that are complementary to the other.
[00280] [00280] A first primer and a second primer are provided, which each have tail regions and template binding regions; the primer array binding regions are complementary to the target template regions. The tail regions of the primers can contain three components: a) the 5' terminal nucleotide of the primer, b) an innermost nucleotide, where the innermost nucleotide is downstream of the 5' end of terminal nucleotides, and c) a mid-section between the 5' terminal nucleotide and the innermost nucleotide, which comprises one or more nucleotides. Furthermore, at least portions of both regions of the primer tail can be complementary to each other when correctly aligned.
[00282] The primary nucleic acid can be treated with a polymerase and a first copy of the first primer under conditions such that the template binding region of the first copy of the first primer hybridizes to the first strand of the nucleic acid template. Under these conditions, an extension product of the first copy of the first primer is formed. Polymerase, which may have strand displacement activity, can catalyze the formation of the first copy extension product of the first primer. The first copy of the first primer may be covalently linked to the synthesized extension product such that the first copy of the first primer (which is complementary to the first strand of the nucleic acid template) becomes par for the molecule described herein as the "product first copy first initiator extension extension ". The template binding region, but not the tail region of the first copy of the first primer hybridizes to the first strand of the nucleic acid template. Examples of conditions suitable for polymerase-based nucleic acid synthesis are known in the art and are provided, for example, in Molecular Cloni maintained: A Laboratory Manual, B. Green and J. Sambrook, Cold Spring Harbor Laboratory Press (2012) , which is incorporated herein by reference in its entirety.
[00283] The extension product of the first copy of the first primer can be treated with a polymerase (which may have strand displacement activity) and with the second primer under conditions such that the template binding region of the second primer hybridizes to the product extension of the first copy of the first initiator. In this way, an extension product of the second initiator can be formed. The polymerase can DISPL ace the first part of the eic acid template Nucl from the extension product of the first copy of the first primer during synthesis of the extension product of the second primer. The second primer can be covalently linked to the synthesized extension product , such that the second primer becomes part of the molecule described herein as the "second primer extension product." The extension product of the second primer is complementary to the extension product of the first copy of the first primer. The late Temp binding region but not the tail region of the second primer can hybridize to the extension product of the first copy of the first primer when the second primer hybridizes to the extension product of the first copy of the first primer.
[00284] The extension product of the second primer can be treated with a polymerase (which may have strand displacement activity) and a second copy of the first primer, so as to provide an extension product of the second copy of the first primer. During generation of the extension product from the second copy of the first primer, the second copy of the first primer may be covalently linked to the synthesized extension product, such that the second copy of the first primer becomes part of the molecule described herein as the "extension product of the second copy of the first initiator." The extension product of the second copy of the first primer is complementary to the extension product of the second primer.
[00285] [00285] The generation of the extension product of the second copy of the first primer may result in the generation of a molecule comprising the extension product of the second copy of the first primer and the extension product of the second primer, which may be referred to herein as the "secondary nucleic acid", A secondary nucleic acid may comprise the "terminal region of the extension product of the second primer (and its complement) and may comprise the 3'3 terminal region of the extension product of the second copy of the first primer (and its complement.) Secondary nucleic acid molecules include sequences from the template region adjacent to tail sequences. In embodiments, double-stranded nucleic acids are produced in which the complementary template and tail region sequences align. In practice, multiple copies (e.g., two or more) of the secondary nucleic acid are produced by any process by which a nucleic acid having the ger structure. aI derived nucleic acid can be generated, including by practicing the non-cycling amplification methods discussed herein, In this way, copy pairs of the derived nucleic acid can be provided. Other copy numbers can then be generated, for example, by repeating the previous steps and methods. For example, the complete process as described above for generating a nucleic acid derived from a primary nucleic acid can be repeated twice in order to generate a two-pair of secondary nucleic acid copies; more repetitions can be performed to amplify the number of additional copies, eg to exponentially amplify the number of copies (eg by powers of two).
[00287] [00287] Furthermore, since secondary nucleic acid molecules include sequences from the template region adjacent to tail sequences, partially double-stranded nucleic acids can be produced in which the tail region sequences and line hybridize. Since these tail region sequences are associated with single-stranded template regions, a cross-over structure having two nucleic acid strands together in possession of the HY bridized tail region sequences is produced. These cross-over structures can be extended by a polymerase to form extension products of both component strands. These extension products, which may be referred to as "wires". Concatemer Two concatemer strands can be annealed together, and can be collectively referred to as a concatemer; such concatemers can contain two or more copies of the nucleic acid template.
[00288] [00288] In some embodiments, even longer concatemers can be formed. For example, it can hybridize concatemers together; or two concatemer molecules can form a crossover structure similar to those formed by the shorter molecules called concatemer strands, as discussed above, followed by a larger concatemer molecule containing four copies of the nucleic acid template. In another example, a nucleic acid and a secondary concatemer can form a cross-over structure, followed by a larger molecule containing three copies of the nucleic acid template. In some embodiments, multiple concatemers of several different lengths can be generated simultaneously.
[00289] [00289] Thus, concatemers generated according to these methods can be of any length of nucleotides. In some embodiments, the concatemer molecules generated herein can be at least 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 400, 500, 600, 700, 800, 900, 1000, 1500, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000, 15000, 20000, or 25000 nucleotides in length. Concatemers generated according to these methods can contain any number of copies of a nucleic acid template. In some embodiments, the concatemer molecules generated herein may contain at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1, 12, 13, 14, 15, 16, 17 , 18, 19, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, or 100 copies of a nucleic acid template. Other examples are provided, and more detail of these and other examples is provided in US Patent Application 61/800,606, filed March 15, 2013.
[00290] [00290] The progress of a method provided herein can be monitored in several different ways. In one embodiment, the reaction can be assayed for a nucleic acid amplification product (for example, to the level of the product or its rate of generation). In another embodiment, the reaction can be assayed for the activity of a polymerase along a nucleic acid template (e.g., for movement of a polymerase along a template strand). Thus, in some embodiments, events of a method provided herein may be observed due to accumulation of product from a method (which may be during or after completion of method steps), or due to detectable events that occur during the steps of a method of ,
[00291] [00291] The presence of amplified nucleic acids can be assayed, for example, by detection of reaction products (amplified nucleic acids or reaction by-products) or by detecting probes associated with the progress of the reaction.
[00292] [00292] In some embodiments, the reaction products can be identified by staining the products with a dye. In some embodiments, a dye may have greater fluorescence when bound to a nucleic acid than it is not bound to a nucleic acid. In some embodiments, a dye can intercalate with a double-stranded nucleic acid, or can bind to an outer region of a nucleic acid. Nucleic acid dyes that can be used with the methods and compositions provided in the present invention include, for example, cyanine dyes, PicoGreen®, OiiGreen®, RiboGreen®, SYBR® dyes, SYBR® gold, SYBR Green I®, ® SYBR green II, ethidiuin bromide, dihydroethidiuni, BlueView™, TOTO® dyes, TO-PRO® dyes, POPO® dyes, YOYO® dyes, Bobo® dyes, JOJO® dyes, LOLO® dyes, SYTOX® dyes, SYTO® dyes, iodide propidium, hexidium iodide, methylene blue, DAPI, acridine orange, quinacrine, acridine dimers, 9-amino-6-chloro-2-niethoxyacridine, bisbenzimide dyes, Hoeehst dyes, 7-aminoactmomycin D, actinomyein D, hydroxystilbamidine, pyronine Y , diamond dye ™, GelRed ™, GelGreen ™ and LDS 751.
[00293] [00293] In some embodiments, reaction products can be identified through turbidity analysis of amplification reactions, for example, where the increase in turbidity is correlated with the formation of reaction products and reaction by-products (for example , magnesium pyrophosphate complexed with),
[00294] [00294] In some embodiments, reaction products can be identified by separating a reaction performed according to a method io here by gel electrophoresis, followed by staining the gel with a dye for nucleic acids. The dye can be any nucleic acid dye disclosed herein or otherwise known in the art.
[00295] In some embodiments, any method or composition known in the ail for detecting acids or processes associated with the generation of nucleic acids can be used with the methods and compositions provided herein.
[00296] In some embodiments, a nucleic acid probe that contains a nucleotide sequence complementary to a portion of a template nucleic acid strand (or the strand that has a similar or identical sequence) and that contains one or both of a fluorescent reporter (fluorophore) and a quencher are included in a reaction provided herein.
[00297] [00297] In one example, a nucleic acid probe may contain a fluorescent reporter at its 5' or 3' end, and a quencher at the other end.
[00298] [00298] In another example, a nucleic acid probe may contain a fluorescent reporter at its 5' or 3' end, and which may be hybridized to a nucleic acid primer containing a quencher. The nucleic acid primer containing an inhibitor may contain the inhibitor at a position on the primer such that when the nucleic acid probe is annealed to the primer, the fluorescent reporter is quenched.
[00299] [00299] In probes containing a fluorescent reporter and quencher pair, the fluorescent reporter and quencher can be selected so that the inhibitor can effectively kill the reporter. In some embodiments, a fluorescent reporter is paired with a quencher, where the fluorescent reporter emission maximum is similar to the inhibitor uptake maximum, Fiuorphores that can be used as the fluorescent reporter include, for example, CAL Fluor gold , CAL Fluor orange, Quasar 570, CAL Fluor Red 590, CAL Fluor Red 610, CAL Fluor Red 610, CAL Fluor Red 635, Quasar 670 (Biosearch Technologies), VIC, NED (Life Technologies), Cy3, Cy5, Cy5.5 (GE Healthcare Life Sciences), Oyster-556, Oyster 645 (Integrated DMA Technologies), LC Red 610, LC Red 610, LC Red 640, LC Red 670, LC Red 705 (Roche Aplica Science), Texas Red, FAM, TET, HEX, JOE, TMR and ROX.
[00300] [00300] In some embodiments, a method provided herein may be monitored in an apparatus that contains a fighting source and an optical sensor. In some situations, the reaction can be positioned in the path of light from the light source, and light absorbed by the sample (for example in the case of a turbid reaction), scattered through the sample (for example in the case of a turbid reaction), or emitted by the sample (for example in the case of a reaction with a fluorescent molecule) can be measured. In some embodiments, a method provided herein may be performed or monitored in a device or module therein, as disclosed in US Patent Application Serial No. 13 / 769,779, filed February 18, 2013, which is incorporated herein by reference. in your totality.
[00301] [00301] A specimen, such as a throat swab, a nasal swab, a mouth swab (eg a face swab), a vaginal swab, saliva, blood or other specimen, may be used for more than one assay . For example, a sample can be subjected to nucleic acid testing and to test for peptides indicative of an infection. In embodiments, a sample can be divided into two or more portions and each portion can be subjected to a single test, or it can be subjected to a plurality of tests. Nucleic acid testing can be used to identify nucleic acid molecules, or portions thereof, whose presence indicates the presence of disease-causing organisms (eg, viruses, bacteria, and other organisms that carry such nucleic acids). The test protein (or peptide) can be used to identify peptides or proteins, or portions thereof, whose presence indicates the presence of disease-causing organisms (for example, organisms expressing such proteins or peptides). Test protein or peptide can be used to identify disease-causing organisms (eg, viruses, bacteria and other organisms) in a sample, and can also be used to identify antibodies directed against such agents that may be present in a sample. Thus, forms of protein (or peptide) tests include testing for the presence of antibodies against targets whose presence indicates the presence of disease-causing organisms. Such test nucleic acid and protein (or peptide) can be used to identify or estimate, or otherwise determine the stage at which an infection in an individual is at the time the sample was taken, by detecting or determining the amounts of, or both, both nucleic acid markers indicative of an infection and protein (or peptide) markers indicative of the same particular infection (such protein markers of the same infection include antibodies to the microorganism causing the infection, as well as markers of protein present in or in the microorganism itself).
[00302] Nucleic acid markers of an infection include DNA and RNA molecules, and fragments thereof unique to the infectious agent (eg, viral nucleic acids, or bacterial nucleic acids, or other nucleic acids from any other infectious microorganism) . Peptide or protein of an infection markers include peptides or proteins specific for the infectious agent (e.g. bacterial peptides); cytokines and other peptides produced in response to infection; and antibodies produced in response to infection.
[00303] [00303] For example, where nucleic acid markers indicative of a particular infection are relatively numerous, while antibody markers indicative of the particular infection are relatively sparse, then the infection is a recent infection; However, where nucleic acid markers indicative of a particular infection are relatively numerous, and antibody markers indicative of that particular infection are also relatively numerous, then the infection is not a recent infection since the object has had time to produce antibodies specific to the infection. Where nucleic acid markers indicative of a particular infection are relatively sparse, and antibody markers indicative of that particular infection are relatively numerous, then the infection may be diminishing and at a late stage. Other protein markers (other than antibodies to the infectious organism) being produced by the disease-causing organism itself, such as viral coat proteins, bacterial cell wall proteins, bacterial toxins, and other non-antibody markers, typically follow a course temporal more similar to that of nucleic acid markers of a particular infection and less similar to that of antibody markers of a particular infection.
[00304] A typical response in killer individuals to infection by many infectious agents includes increased levels of inflammatory cytokines (including mterleukin 1 (IL-1 tumor necrosis factor), IL-6, IL-1 8, a ( TNF-a), interferon gamma (IFN-γ), and others). Cytokine levels can increase rapidly after infection.
[00305] [00305] The time course of producing antibodies against an infectious agent (eg, a virus, bacteria, or other infectious micro-organism) varies between individual subjects and from infection to infect onwards; such time limits for courses can be known or identified for different types of infections. Generally speaking, a few days or more are required before antibodies to an infectious agent are detectable in a subject; once detectable, the amount of antibody detected in a subject often grows very rapidly, and can plateau (or peak) over a period of weeks or months after infection, in addition to the type of infection, the factors that can affect the plateau (or peak) levels, and the timing at which those levels are reached, whether or not the infection is acute or chronic; the severity of the infection; other diseases or conditions that affect the subject; the subject's nutritional status; environmental factors; and other factors.
[00306] [00306] The time course of an acute infection can be short; for example, an acute infection may follow a course time measured in days or weeks. For example, many viral and bacterial infections in an otherwise healthy human subject usually go away within about a week. Levels of nucleic acid and protein markers indicative of viral, bacterial and other infections typically rise, and then fall, during the course of the infection. Initially, after infection, and following near the time of infection, infectious agent markers (eg, Nucleic acid markers and protein markers indicative of viral, bacterial, or other infection) will be detectable in samples obtained from a Object: Levels of such markers will rise from the time of infection, and will typically peak within a few days (for a short-lived infection) or within a few weeks (for a longer-term infection). Antibodies to the infectious agent can be detected within a week or two after infection, and can then increase further over several weeks (eg, for a month or more). If the infection resolves itself and the infectious agent is cleared from the subject, antibody levels will then slowly decrease over a period of months.
[00307] Long-term, or chronic infections may follow a longer temporal course. For example, the time course of viral markers and antibody formation in a person infected with the human immunodeficiency virus (HIV) may follow a time course over many months and years. Initially, HIV viral markers (eg , the viral p24 antigen, viral nucleic acids, and other viral markers produced by the virus itself) may be present (in samples obtained from a subject) at elevated levels during the first few months after infection, and may peak at approximately 6 months after infection. In contrast, anti-HIV antibodies (eg antibodies to the g L20 or other antigenic viral epitopes) are not detectable in samples obtained from an object for a few months after infection, but become detectable for about 3- 5 months after infection, and anti-HIV antibody levels rapidly rising over the next 6 months or so, continue to increase at a slower rate from about 1 year after infection to about 4 to 6 years after infection. During this time period (from about a year to about five years) the levels of viral markers can be very low; However, in the absence of treatment, as levels of T cells (eg, CD4 T cells) typically have not reached over the time period of about one year to about five years after HIV infection, the subject may begin to suffer from Systemic immune deficiency and greater loss of T cells (eg CD-8 T cells) about 5-6 years after HIV infection. Levels of HIV viral markers will typically increase as systemic immune deficiency becomes apparent 5 - 6 years after HIV infection.
[00308] [00308] The time of plates relevant to the detection of the marker may also depend on the type of sample tested for the presence of the marker. For example, in some infections, the causative organism, and the organism's nucleic acid and protein markers, may be found initially in blood, or saliva, or other fluid or tissue; and can later be found in urine or stool samples; and still later it can be detectable in tissue samples. Antibodies to such disease-causing organisms, which normally appear some time after the organism's appearance in samples, can be found first in blood and then following their appearance in blood, saliva, urine, or feces. .
[00309] [00309] The devices, systems and assays described herein may use techniques, devices, systems and assays disclosed, for example, in US Patent 8,088,593; US Patent 8,380,541; United States Patent Application Serial No. 13 / 769,798, fifed February 18, 2013; United States Patent Application Serial No. 13/769,779, filed February 18, 2013; US Serial Application No. 61 / 800,606, filed March 15,
[00310] [00310] For example, devices for use in performing assays for detecting a plurality of pathogens in a single clinical sample, or in a plurality of aliquots of a single clinical sample include cartridges including some or all of the containers, reaction containers of reagents, tools and instruments used in testing, and reagents used in testing. A cartridge can contain one or more of the following: ship reagents; reaction vessels; cytometry cuvettes; garbage containers; sample collection devices or sample collection vessels; and other vessels and materials. Such devices may include multiple vessels containing reagents for use in an assay for detecting a plurality of markers indicative of an infectious agent, for example, an upper airway infectious agent; a lower respiratory infectious agent; a causative agent of a sexually transmitted disease; a detectable agent from a sample obtained from a swab (eg, a throat swab, a nasal swab, a mouth swab (eg, a SW r ah cheek), a vaginal swab, or other swab) ; a detectable agent from a blood sample; or their combinations.
[00311] Thus, for example, a cartridge may contain a plurality of available reagents; a plurality of reagent vessels, containing reagents for detecting a marker indicative of a disease-causing agent. disease-causing agents can include agents that cause upper airway disorders, or lower respiratory tract disorders, or sexually transmitted diseases. A disease-causing agent can be detected in a blood sample. disease-causing agent A. can be detected in a specimen obtained with a swab, such as a throat swab, or a nasal swab, or a buccal swab (eg, a buccal swab), or a vaginal swab, or other specimen or their combinations.
[00312] [00312] In embodiments, a device comprising or comprising a cartridge configured to contain one or more reagent vessels, and one or more reaction vessels, for example, for use in nucleic acid assays; for use in immunoassays (for example, ELISA assays); for use in general chemistry assays (eg, for clinical electrolytes, vitamin levels, blood component levels, and other targets); for use in cytometric assays; and for their combinations. In embodiments, such a device can include reagents, reaction vessels, and tools, troughs, and other instruments for use in nucleic acid assays; for immunoassays (for example, ELISA assays); general chemistry assays (eg, for clinical electrolytes, vitamin levels, blood component levels, and other targets): cytometric assays; and for their combinations.
[00313] [00313] In embodiments, a cartridge to be used in performing assays for detecting a plurality of disease-causing agents as disclosed herein may include one or more spaces or vessels for holding a rod or smears. A single wick can be placed in a single space, or in a single vase; in embodiments, two pads can be placed in a single space, or a single vase. In embodiments, a plurality of smear samples may be placed in a single space, or a single vessel. Vessels for holding a rod, or swabs, may contain a reagent, or a diluent, or other solution for use with a rod or swab.
[00314] [00314] A nasal swab may be useful for testing for upper respiratory tract illnesses, and a throat rod may be helpful for testing for lower respiratory illnesses. In embodiments, a mouth swab (e.g., a buccal swab, a tongue swab, a gum wad, or other swab taken inside the mouth) may be used in addition to or instead of a nasal or swab throat.
[00315] [00315] In embodiments, a cartridge for use in performing assays for detecting a plurality of disease-causing agents as disclosed herein may include one or more spaces or vessels for holding a blood sample. In embodiments, a cartridge for use in performing assays for detecting a plurality of disease-causing agents as disclosed herein may include one or more spaces or vessels for holding a blood sample and may also include a or more spaces or vessels for holding a rod or swabs. Thus, a device such as a cartridge can take a blood sample and a throat swab; can take a blood sample and a nasal swab; and may hold a blood sample, a throat swab, and a nasal swab. In embodiments, a device, such as a cartridge, can take a blood sample and a mouth swab: it can take a blood sample, a mouth swab, and a nasal discharge; and can hold a blood sample, a mouth swab, a throat swab, and a nasal swab.
[00316] In embodiments, a cartridge for use in performing assays for detecting a plurality of disease causing agents as disclosed herein may include one or more spaces or vessels for holding a urine sample. In embodiments, a cartridge for use in performing assays for detecting a plurality of disease-causing agents as disclosed herein may include one or more spaces or vessels for holding a urine sample and may also include a or more spaces or vessels for holding a rod or swabs. Thus, a device such as a cartridge may contain a urine sample and a throat swab; can perform a urine sample and a nasal swab; can hold a urine sample, a throat swab, and a nasal swab; can hold a urine sample, a throat swab, a mouth swab, and a nasal swab; and can hold a urine sample, and one or more of a swab, a mouth swab, a vaginal and nasal swab, and a swab.
[00318] [00318] A cartridge may include a sample, and may include reagents to be used in treating or testing a sample, disposables to be used in treating or testing a sample, or other materials. Upon placement of a cartridge into, or insertion of a cartridge into, a sample processing device or system, one or more components of the cartridge may be brought into fluid communication with other components of the sample processing device. For example, the sample is collected in a cartridge, the sample can be transferred to other portions of the sample processing device. Likewise, if one or more reagents are provided in a cartridge, the reagents can be transferred to other portions of the sample processing device, or other components of the sample processing device can be brought into the reagents. In one embodiment, reagents or components of a cartridge may remain on the edge of the cartridge. In some embodiments, no fluidics are included that require tubing or that require MAINTENANCE (e.g., manual or automated maintenance).
[00320] [00320] As shown in Fig. 20A, fig. 20B, and fig. 20C, a container for holding a swab can be loaded into a cartridge, where it can be held until needed for analysis; the cartridge can be loaded into an analysis system or analysis device, thereby loading the rod of (and any other samples or sample containers in the cartridge as well). As shown in Fig. 2.0A, a container for holding a rod (a swab vessel 10) can be loaded into a cartridge of 20 by placement within a receptacle 30. The cartridge 20 also includes, as shown, cavities and cavities 40 to receive and store reagents and vessels. A wick container 10 may hold a rod in place within the swab vessel 10, or it may be loaded into a cartridge without a swab in place within the swab vessel 10.
[00321] [00321] As shown in Fig. 2.0B, a container for holding a rod (a swab vessel 10) can be loaded into a cartridge of 20 by placement within a receptacle 30. The cartridge 20 also includes, as shown, cavities and wells 40 for receiving and storing reagents and vessels. In the embodiment shown in Fig. 20B, cartridge 20 also includes a sample collection reservoir 50, which can hold, for example, blood, urine, or other sample. The arrows away from the swab vessel 10 indicate how the swab container 10 can be placed within a receptacle 30 in the cartridge 20.
[00322] [00322] As shown in Fig. 20C, a container for holding a rod (a swab vessel 10) can be loaded into a cartridge of 20 by placement within a receptacle 30. The cartridge 20 also includes, as shown, cavities and wells 40 for receiving and storing reagents and vessels. As shown in the embodiment of fig. 20C, cartridge 20 includes a swab receptacle 60 configured to hold a rod of 70. In embodiments (e.g., in the embodiment illustrated in Fig. 20C) of a cartridge 20 having a swab receptacle 60 may include, optionally, a sample collection container 50, which can hold, for example, blood, urine, or other sample. Such a swab 70 may be made into the swab receptacle 60 prior to its use in taking a sample. In embodiments, a swab 70 can be placed into a swab vessel 10 after collecting a swab sample 70. In the embodiment shown in Fig. 20C, swab vessel 10 can be loaded into a cartridge without a swab in place inside. of the swab vessel 10 prior to using the swab 70, and a swab vessel 10 may be replaced in a swab vessel 30, which holds swab 70 within the swab vessel 10 after taking a sample by swab 70.
[00323] [00323] Swabs can be any swab suitable for taking a sample. Several examples of swabs suitable for use in sampling are shown in Fig. 21. Swabs with flake tips (00 swab, and shorter swab 200, suitable for pediatric use), or those that are also suitable for use in the establishment of cultures of materials obtained by scraping a body orifice, a body cavity or surface of an object (300 swab), cotton swabs (400 swab), and other swabs can be used to collect a sample from a patient for use with the methods , systems and devices disclosed herein. For example, samples can be obtained by scraping a nasal cavity, throat, mouth, vagina, or other orifice, body cavity, or locating on or within a subject.
[00324] [00324] An analysis system, which may include an analysis device, such as a sample processing device, may have a fluid handling system (also referred to herein as a sample handling system). A fluid handling system can perform, or can assist in performing, transport, dilution, extraction, aliquotting, mixing, and other actions with a fluid, such as a sample. In some embodiments, a fluid handling system may be contained within a housing of the device. A fluid handling system may allow for the collection and delivery, processing and/or transport of a fluid, the dissolution of dry reagents, mixing of the liquid and/or dry reagents with a liquid, as well as the collection, deliver "-, the processing and/or transport of non-fluidic components, samples, or materials. The fluid may be a sample, a reagent, diluent, wash, dye, or any other fluid that may be used by the device, and may include, but not be limiting to homogeneous liquids, different liquids, emulsions, suspensions, and other fluids, a fluid handling system, including, without limitation, a pipette, may also be used for the transport of containers (with or without fluid contained therein) in The fluid handling system may dispense or aspirate a liquid The sample may include one or more particles or solid matter floating within a fluid.
[00326] [00326] A. fluid handling system may include one or more fiuidically isolated or hydraulically independent units. For example, the fluid handling system can include one, two, or more pipette tips. Pipette tips can be configured to accept and confine a fluid. The tips can be insulated from fiuidically or hydraulically independent of one another. The fluids contained within each tip can be hydraulically isolated or fiuidically independent of each other in fluid tips and other fluids within the device. The fiuidically insulated or hydraulically independent units can be mobile relative to other portions of the device and/or one another. Independent units fiuidically insulated or hydraulically, can be individually movable. A fluid handling system can comprise one or more base or support. A base or support can support one or more pipette or pipette units. A base or support can connect one or more pipettes of the fluid handling system with one another.
[00327] [00327] A sample processing system, which may include a sample processing device, may be configured to perform processing steps or actions on a sample obtained from a subject. Sample processing may include sample preparation, including, for example, sample dilution, aliquoting a sample, extracting, contacting with a reagent, filtration, separation, centrifugation, or other preparatory, transformation, or other action. stage. A sample processing device can be configured to perform one or more sample preparation action or step on the sample. Optionally, a sample can be prepared by a chemical reaction and/or physical processing step. The sample or step preparation action may include one or more of the following procedures: centrifugation, separation, filtration, dilution, enrichment, purification, precipitation, incubation, pipetting, transport, chromatography, Ceil lysis, cytometry, spraying, milling , activation, ulrrasonication, micro column processing, magnetic bead processing, nanoparticle processing, or other sample action preparation steps or steps. For example, sample preparation can include one or more steps to separate blood into serum and/or particle fractions, or to separate any other sample into multiple components. Sample preparation may include one or more steps of diluting and/or concentrating a sample, such as a clinical sample or biological sample, for example, from blood, urine, sputum, material obtained from a nasal swab, a throat swab, a buccal swab, or other specimen, or other clinical or biological specimens. Sample preparation can include adding an anticoagulant or other ingredients to a sample. Sample preparation can also include purification of a sample. In embodiments, all sample processing, preparation or testing actions or steps are performed by a single device. In embodiments, all sample processing, preparation or testing actions or steps are performed within a single device housing. In embodiments, most sample processing, preparation or assay actions or steps are performed by a single v-ice, and may be performed within a single device housing. In embodiments, many sample processing, preparation or testing actions or steps are performed by a single device, and may be performed within a single device housing. Within embodiments, sample processing, preparation or assay actions or steps may be performed by more than one device.
[00328] [00328] A sample processing system, which may include a sample processing device, may be configured to perform one or more runs of a sample, and to obtain data from the sample. An assay can include one or more chemical or physical treatments, and can include performing one or more chemical or physical reactions. A sample processing device can be configured to run one, two or more tests on a small sample of body fluid. One or more chemical reactions can take place in a sample with one volume, as described elsewhere herein. For example one or more chemical reaction may take place on a tablet having less than femtoliter volumes, in one example the sample collection unit is configured to receive a volume of body fluid equivalent sample for a single drop or less of blood or interstitial fluid. In embodiments, the volume of a sample can be a small volume, where a small volume can be a volume that is less than about 1000 µl, or less than about 500 µl, or Jess than about 250 μl ,, or less than about 150 μΕ , or less than about 100 μl , or less than about 75 .L , or less than 50 aboui L , or less than 40 uL aboui °, or less than about 20 or less than about 10 or other small volume. In embodiments, all sample test measurements or steps are performed on a single sample. In embodiments, all sample test measurements or steps are performed by a single device. In embodiments, all sample test measurements or steps are performed within a single device housing. In embodiments, most sample testing actions or steps are performed by a single device, and may be performed within a single device housing. In embodiments, many sample testing actions, or steps are performed by a single device, and may be performed within a single device housing. In embodiments, sample processing, preparation or testing actions or steps may be performed by more than one device.
[00329] [00329] A sample processing system, which may include a sample processing device, may be configured to perform a plurality of assays on one sample. For example, an addiction transformation sample can be configured to detect, or to identify, or to measure the identifying material of pathogens in a sample. In embodiments, a sample processing device can be configured to perform a plurality of assays on a single sample. In embodiments, a sample processing device can be configured to perform a plurality of assays on a single sample, where the sample is a small sample. For example, a small sample must have a sample volume that is a small volume of less than about 1000 or less than about 500 SL, or less than about 250, IIl, or less than about 150 μΙ_, or less than about 100 μl, or less than about 75^L, or less than about 50 μl, or less than about 40 μl, or less than about 20 μϊ_ ·, or less than about 10 μl., or other small volume. A sample processing device may be capable of performing multiplexed assays on a single sample. A plurality of tests can be run simultaneously; can be run sequentially; or some tests can be run simultaneously, while others are run sequentially. One or more assays and/or control calibrators (eg including a setup with a calibrator control for the assay/tests) may also be incorporated into the device; Control and assay runs on the calibrators may be performed simultaneously with assays performed on a specimen, or may be performed before or after assays performed on a specimen, or any combination thereof. In embodiments, all sample test measurements or steps are performed by a single device. In embodiments, all of a plurality of test actions or steps are performed within a single device housing. In embodiments, most sample testing actions, or steps, of a plurality of tests are performed by a single device, and may be performed within a single device housing. In embodiments, many sample testing actions, or steps, of a plurality of tests are performed by a single device, and may be performed within a single device housing. In embodiments, sample processing, preparation or testing actions or steps may be performed by more than one device.
[00330] [00330] In embodiments, all of a plurality of trials can be performed in a short period of time. In embodiments, such a short time period comprises less than about three hours, or less than about two hours, or less than about an hour, or less than about 40 minutes, or less than about 30 minutes, or less than about 25 minutes, or less than about 20 minutes, or less than about 15 minutes, or less than about 10 minutes, or less than about 5 minutes, or less than about 4 minutes, or less than about 3 minutes, or less than about 2 minutes, or less than about 1 minute, or other short period of time.
[00332] Systems for detecting the presence of one or more of a plurality of markers indicative of an infectious disease in a small volume of clinical sample include, for example, a) a sample handling system; b) a detection station comprises an optical sensor; c) a fhridically isolated sample collection unit configured to hold a clinical sample; d) an assay station comprising at least a first and a second fluidly isolated assay unit, wherein the first unit comprises a first reagent and the second unit comprises a second reagent; and e) a controller, wherein the controller comprises a focal memory and is operatively coupled to the sample handling system and the detection station. Such systems may be configured to perform tests with one or both of the first and second test units: wherein the controller's local memory comprises a protocol comprising instructions for: i) the direction of the sample handling system to transfer a portion of the sample clinical for the first test unit and for the second test unit; and ii) directing the sample handling system to transfer the first assay unit and the second assay unit unit to the detection station. In other embodiments, a test station in such systems may include at least a first, second, and third unit, fluidly isolated assay, wherein the first unit comprises a first reagent, the second unit comprises a second reagent, and the third unit comprises a third reactant. In other embodiments, a test station in such systems may include at least a first, second, third and fourth fluidically isolated test unit, wherein the first unit comprises a first reagent, (the second unit comprises a second reagent, the third unit comprises a third reagent, and the fourth unit comprises a fourth reagent It should be understood that embodiments of such systems may include more than four assay units; or other numbers of assay units.
[00333] [00333] In embodiments, tests are performed with any one or more of the first, second and third test units; or with any one or more of the first, and the fourth, second, third test units, wherein the controller's local memory comprises a protocol comprising instructions for: i) directing the sample handling system to transfer a portion of the clinical sample for the first test unit, the second test unit, and, where applicable, the third test unit and/or fourth test units; and ii) direct the sample handling system to transfer the first test unit, the second test unit, and, where applicable, the third test unit and/or the fourth test unit, to the detection station.
[00334] [00334] Furthermore systems for detecting the presence of one or more of a plurality of markers indicative of an infectious disease from a small volume clinical sample include: a) a sample handling system; b) a detection station comprises an optical sensor; c) a fluid handling system configured to transport liquids between components of said system, wherein said fluid transport comprises transporting isolated aliquots of fluid; d) a fluidly isolated sample collection unit configured to hold a clinical sample; e) a test station comprising at least a first, second, and third fluidly isolated test unit, wherein the first unit comprises a first reagent, the second unit comprises a second reagent, and the third unit comprises a third reagent ; and f) a controller, wherein the controller comprises a local memory and is operatively coupled to the sample handling system and the detection station. Such systems can be configured to perform tests with any one or more of the first, second, and third test units; wherein the controller buffer comprises a protocol comprising instructions for: i) directing the sample handling system to transfer a portion of the clinical sample to the first test unit, the second test unit and the third test unit; and ii) direct the sample handling system to transfer the first test unit, the second test unit, and the third test unit to the detection station. It should be understood that embodiments of such systems may include only two test units; or it can include four test units; or other numbers of bake units.
[00335] [00335] Clinical sample processing systems for use in performing assays as disclosed herein may include: a) a sample handling system; b) a detection station comprises an optical sensor; c) a fluidly isolated sample collection unit configured to hold a clinical sample; d) an assay station comprising at least a fluidly isolated first, second, and third assay unit, wherein the first unit comprises an antibody, the second unit comprises an oligonucleotide, and the third unit comprises a chromogen or a dye or another marker; and e) a controller, wherein the controller is operatively coupled to the sample handling system, wherein the sample handling system is configured to transfer a portion of the clinical sample from the sample collection unit to each of the first unit test unit, the second assay unit, and the third assay unit, and the device is configured to perform an immunoassay,
[00336] [00336] These systems may serve as point of service (POS) systems. These systems can be contained within a housing, a system located at a POS location can be configured for use in analyzing a sample at the POS location. These systems can be POS systems configured to run a plurality of assays on a single small volume sample or on aliquots thereof.
[00337] [00337] disease-causing agents such as viruses, bacteria, yeasts, fungi and other microorganisms identify nucleic acids and proteins, among others identifying characteristics that can serve as markers. Markers indicative of a disease or disease-causing agent, such as a respiratory disease, or a form of influenza, or a sexually transmitted disease, or other disease, nucleic acid markers.
[00338] As shown in the figures, markers for many diseases can be tested for, and can be detected using nucleic acid assays as disclosed herein. All disease-causing agents tested in the tests shown in Fig. 1A were detected within 40 minutes, with most agents detected within about 30 minutes. Copy number detection times of 100 copies per microliter (c / uL) were shorter than for lower copy numbers (10 c / u, L). Detection times for samples with 100 copies per μΕ are shown in Figure IB; most were close to or less than 20 minutes, with various detection times of about 15 minutes or less.
[00339] Fig. 1A provides a graphical summary of the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample by a variety of markers and for two different concentration ranges of the markers (10 e / μί and 100 c / μl, where "c μΓ means copies per microliter (μl,)). the 10 results c / μl are shown on the left, of the 100 c / μl results for each type of disease (influenza ( influenza), respiratory, and sexually transmitted diseases (STD)) shown in figu'e. the times are labeled "LOD" ("delay length") the vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU), in thousands.
[00340] [00340] Fig. IB provides a bar graph showing the durations of time from the start of the nucleic acid assay until the detection of the presence of a target nucleic acid in a sample for the indicated markers for various diseases (at 100° C / id).
[00341] More information relating to detection time for various diseases, grouped by general location of infection, or type of disease, or samples, by which diseases can be detected, are presented in Figs. 1C through IF (all at 100 c / u!). FIG. 1 C shows time durations from initiation of nucleic acid assay to detection of various influenza strains and identification targets. FIG. ID shows durations of time from initiation of nucleic acid assay until detection of various respiratory diseases. Fig, IE shows time durations from initiation of nucleic acid assay to detection of various sexually transmitted diseases. Fig, SE shows the durations of time from the start of the nucleic acid assay until the detection of various diseases that can be detected in blood,
[00342] [00342] Additional information presented in Table 2A indicates the copy numbers per μl that are detectable with these assays for various diseases.
[00343] [00343] in the above tables and elsewhere in this document, "NA" indicates neuraminidase; "HA" indicates hemagglutinin; "Klebsiella pneumonia
[00344] [00344] The mean detection time for these 54 diseases (at 100 c/μl or less) was less than 2.3 minutes (mean 22.77 minutes). A smaller subset of 35 diseases measured at 10 C/μl or less had a mean time to detection of less than 30 minutes (1 minute mean 29.1). These assays, including assays for these diseases, are suitable for validation for use in clinical laboratories Laboratories Improvement Act (CL1A). For example, these trials for various forms of influenza (influenza A, pandemic influenza B, h1n 1 -Novel, seasonal H1N1 and influenza F13N2) were performed through CLIA Validation.
[00346] [00346] Samples obtained from individuals, including small samples from individuals, can be tested for other diseases in addition to the diseases indicated in the figures and Table 2A. For example, some other diseases that can be tested by these methods are shown in Table 2B. The "Panel" column indicates the type of disease (where FIAT indicates Hospital acquired the infection, and STD indicates sexually transmitted disease).
[00347] [00347] The systems, methods and devices described herein can be used to test, and detect, the presence of markers indicative of one or more of the infectious agents listed above; such tests, and such that it detects, can be performed on a single clinical sample, or on a plurality of aliquots from a single clinical sample. Such a single clinical sample may be a single clinical sample of small volume. This type of testing, and detection, can be performed at a capture site; systems, devices and methods can be POS systems, devices and methods. For example, the clinical sample can be collected at the POS site, and can be analyzed on a device at the POS site. As shown in the results illustrated in the figures, the analysis of the small volume clinical sample can be completed in a short period of time.
[00349] [00349] TABLE 3 Agents and protein Markers Same influenza A Matrix-cause illness influenza H3N2 Influenza H 1N1 seasonal new influenza H 1IM1 Influenza B Streptococcus pyogenes (A) Mycobacterium tuberculosis Staphylococcus aureus (MR) Staphylococcus aureus (RS) Borciete !! pertussis (whooping cough) Streptococcus agalactiae (B)
[00350] [00350] The disease-causing agents listed in Table 3 can be tested for, and can be detected, through the methods and using the systems and devices described herein. For example, markers for the disease-causing agents listed in Table 3 can be tested for, and can be detected by, the methods and using the systems and devices described herein. Such tags can include, for example, nucleic acid tags. Furthermore, such markers may include saccharide markers, or other markers such as, for example, protein markers. Methods of testing for, and detecting, protein markers are discussed in the following example.
[00351] The detection of nucleic acid from 2}l of sample taken from cultured cells infected with seasonal influenza virus (H1N1) is shown in Figures 19A (sample) and 19B (Controle). Nucleic acid obtained from cell cultures was prepared using the Chemagic I magnetic separator module with 24 DWP XL adapters and reagents fro the Chemagic viral DNA/RNA Kit (CMC-No. 1089; CMC-No. 1082 is similar) from Chemagen (PerkinElmer chemagen Technologic GmbH, Baesweller, Germany)). This method uses magnetic bead separation to isolate RNA. and DN A from a sample. Chemagen reagents and disposables were used in sample preparation.
[00352] [00352] RA influenza H1N1 was obtained from cultured infected MDCK cells. Briefly, cell culture samples were prepared by dispensing approximately 1 mL of the sample solution into a lysis buffer containing as well polyfA RNA reagent), and proteinase K solutions, with gentle mixing. The wells were covered and heated to 55 °C for ten minutes. After this 10 minute incubation, binding buffer was added to the wells containing the lysed sample solution. This mixed solution was then processed by the T magnetic separator module. The Chemagic nucleic acids were released by vortexing (spinning probes) into the buffer and then attached to the magnetic beads, which were immobilized by a magnet during the washing steps. Nucleic acids released in the bead-bound sample and were retained during the washing steps; following the washing steps, nucleic acids were eluted in elution buffer (10 mM TR1S-HC1, pH 8.0).
[00353] [00353] Following this sample preparation, the prepared sample was placed in a container held in a cartridge, and the cartridge was loaded into an automatic sample analysis device (such cartridges, devices, and their uses are described, for example, in US Patent 8,088,593; US Patent 8,380,541; US Patent Application Serial No. 13 / 769,798, filed February 18, 2013; US Patent Application Serial No. 13 / 769,779, Filed February 18, 2013; United States Patent Application Serial No. 13 / 769,820, filed February 18, 2013; PCT / US2012 / 57155, filed September 25, 2012; Application
[00354] [00354] The 2 IIL aliquot of the prepared sample solution was placed in a vessel containing 20, II, L of MasterMix (contammg buffer, betaine, dNTP, forward (RLX1222) and reverse probes (RLX1223), Syto 59 red dye ), and mixed with 3 µl of the enzyme preparation (containing B. stearothermophilus DNA polymerase (Bst), Avian Myeloblastosis Virus Reverse Transcriptase (AmvRT), NEB4 buffer (New England BioLabs Cat. No. B7004S), and water ) in a reaction vessel in the automatic sample analysis device. Primers specific for the H1N1 influenza virus were included in the mixture in the reaction vessel. The combination of sample, MasterMix, template preparation, and enzyme was incubated at 56°C in the reaction vessel according to the methods discussed above, and fluorescence was measured every minute for 30 minutes (fluorescence was 59 SYTO dye). Fluorescence was read as relative fluorescence.
[00355] [00355] Fig. 19A shows the amplification over time, the increase in fluorescence relative to about 15 to 20 minutes indicating the presence of a seasonal marker of influenza H1N1. The horizontal axis is in "minutes, the vertical axis is shown in rel fluorescence units (relative fluorescence units, RFU).
[00356] [00356] Fig. 19B shows the amplification of "no template control" (no added copies of the target marker; CT). Note that most experiments showed no amplification; the three runs showing final increases in relative fluorescence did so at about 25 minutes or later. The horizontal axis is in "minutes, the vertical axis is shown in relative fluorescence units (relative fluorescence units, RFU).
[00357] [00357] The results of Figures 19A and 19B show that viral nucleic acid can be detected from small volume samples (eg 2, μΙ_ sample) within a short period of time (eg about 15 20 minutes or less).
[00358] [00358] The detection of proteins indicative of influenza A infection and proteins indicative of influenza B infection was performed using devices and systems as described, for example, in the US Patent
[00359] [00359] Assay Design and Purpose: The Influenza A and Influenza B assays are designed to provide qualitative detection of influenza A or influenza B nucleoprotein antigens in a sample obtained with a nasal swab. These assays are useful in diagnosing influenza A viral infections or influenza B viral infections in a subject from which the sample was obtained. The assay was a sandwich assay, in which anti-influenza A or B antibodies were immobilized on a substrate (the inside of a translucent or transparent tip pipette), and the sample, alkaline phosphatase (ALP) - anti-conjugate antibody of Influenza A or B, and the ALP substrate added to produce Chemi luminescence proportional to the amount of Influenza antigen in the sample. Assay results were compared with those of a commercial test (Remel X/pecto influenza A and B; Remel products, Lenexa, KS, USA, a division of Thermo Fisher Scientific, Inc.).
[00360] [00360] Materials and Methods: The interior of a custom polymer tip served as the surface for this sandwich ELISA assay. Pipette tips were typically made from polystyrene or polypropylene, although other polymers or plastic materials are also suitable. The insides of pipette tips were coated with avidin. The capture surface for the ELISA sandwich was prepared by coating biotin-labeled anti-influenza A, or biotin-labeled antibody anti-influenza B virus antibody on the avidin-coated interior surfaces of the pipettes.
[00361] [00361] capture and detection antibodies were obtained from the United States Biological Corporation (Salem, MA, USA) or Souihe nBioiech (SouthemBiotechnology Associates, Inc., Birmingham, AL, USA); capture antibodies were conjugated to biotin using a biotin labeling kit and detection antibodies were conjugated to ALP using an ALP labeling kit, either from Dojindo Molecular Technologies, Inc. (Rockviile, MD, USA). Buffers were obtained from Sigma Aldrich Corporation (St. Louis, MO, USA).
[00362] [00362] Samples were obtained from the nasal passages of individuals using nasal swabs. nasal swabs containing sample material were then subjected to an extraction process. ALP-labeled anti-influenza A or B anti-influenza antibodies ALP-labeled anti-influenza antibodies were then mixed with the extracted sample material. This mixture was then incubated with the capture surface for 5 minutes. After incubation, the capture surface was washed and the ALP substrate was incubated on the surface for 5 minutes; the resulting chemiluminescence intensity was then read, with the results presented in relative light units (RLU).
[00363] Buffers: Tris-buffered saline consisted of 138 mM NaCl, 2.7 mM KCl and 0.05 M tris (hydroxymethyl) aminomethane (TRIS), pH 8.
[00364] The extraction buffer was 0.5% Tween 20, 0.1% sodium azide in 20 mM sodium phosphate buffer (pH 7.6).
[00365] [00365] The blocking buffer was 3% BSA blocking buffer, which consists of Tris-buffered saline, 3% BSA, 0.05% NaNs, at pH 8.
[00366] [00366] Alkaline phosphatase (AP) stabilizer was prepared by adding 0. IMM zinc chloride and 5 mM magnesium chloride to 3% BSA blocking buffer,
[00367] [00367] The wash buffer was TRJS-buffered saline, 0.05% Tween 20, 0.05% NaNj, at pH 8.
[00369] [00369] Capture Surface titration The capture surface r w as titrated with the following concentrations: 10 g mL, 5 mL g, and I u / ηι1. Virusys Kit and Microbix Kit controls were used for this selection. The background control was a blank blocking buffer with no sample added. The dAb was maintained at a concentration of 100 ng/ml (final concentration in blocking buffer). The optimal CAB concentration was determined to be 5 ug/ml for influenza A and influenza B.
[00370] [00370] Alkaline Phosphatase Stabilizer: Two alkaline phosphatase stabilizers were tested for use as DAB diluents. In these experiments, 50 µl of the sample was added to 500 µl of extraction buffer. The taxi concentration was 5 μ / ηιΐ while the DAB concentration was maintained at 100 ng / ml (final concentration after running the protocol). Both the custom AP stabilizer solution (cuff ingredients above) and the commercial Stabilzyme ® AP conjugate stabilizer (SurModics, Inc., Eden Prairie, M, USA) worked well. The custom AP stabilizer was used in subsequent experiments.
[00371] [00371] Detection Antibody Titration: The AP-conjugated Dabs were titrated in the AP stabilizer solution. The best modulation between positive and negative controls were observed at 50 ml of final concentration ng. Influenza A positive controls were obtained from Microbix Biosystems, Inc. and ZeptoMetrix Corporation (Buffalo, NY, USA), and influenza B positive controls were obtained from Microbix Biosystems, Inc. and Virusys Corporation.
[00372] [00372] Specificity tests - Influenza A: Specificity and cross-reactivity studies were performed in extraction buffer using the sample processing and analysis devices and systems as disclosed herein. Test controls for cross-reac potential! Aniseed were obtained from Microbix Biosystems, Inc. The potential crossover reagents tested were Respiratory Syncytial Virus, Mycoplasma pneumonia, Adenovirus, Parainfluenza A-III, Parainfluenza A-II and Parainfluenza A-I. Taxi concentration was 5 µg/ Rnl while DAB concentration was 100 Nextel (final concentration after protocol execution) ng /. No cross-reactivity was detected in these experiments. Different strains of influenza A and influenza B were also tested to determine Influenza A specificity in the assay. Both Zeptometrix and Microbix controls (which are pre-diluted controls) were used for this purpose than not. Influenza positive control swabs from the Remei Xpect Flu kit were also used. A sample volume of 200 μί was mixed with 200 μl of extraction buffer and tested for these prediluted samples. Swabs 400 µl of extraction buffer were processed for these experiments. In IHE following ables f, light unit (RLU) relative measurements were made in triplicate; "CV%" is calculated by dividing the standard deviation of three measurements by the average of the three measurements and multiplying by 100.
[00373] [00373] Specificity Tests - Influenza B: Specificity and cross VITY Reacti studies were performed in extraction buffer using the sample processing and analysis devices and systems as disclosed herein. taxi concentration was 5 μ§ τη1 while the DAB concentration was 100 ng/ml (final concentration after protocol execution). No cross-reactivity was detected in these experiments.
[00374] The clinical evaluation of the Influenza A virus assay: The performance of the Influenza A assay using the sample processing and analysis devices and systems as described herein was compared with the results obtained with the Remel FDA kit. CAB concentration was 5 μg/^ml while DAB concentration was maintained at 50 ng/ml (final concentration after protocol was RAN). For the National Institute of Biological Standards and Control for influenza strains (NIBSC, Hertfordshire, UK), 50 µl of sample was added to the swab, and the swab was then treated as a sample rod. For the Zeptometrix panel controls (pre-diluted samples), 200 µl of sample was mixed with 200 µl of extraction buffer. Sample swabs were placed in 500 µl of extraction buffer and incubated for 3-5 minutes. This extracted sample was then analyzed using the devices and systems as disclosed herein. Swabs and specimens were processed in the Remei FDA kit as indicated in the kit instructions.
[00375] [00375] In the following tables, the "antibody index" (Index Ab) was used to determine whether or not target influenza antigens were detected in a sample. calculated from normal samples). The cutoff value was set equal to the mean (normal), plus 4.5 x standard deviation (normal). An Ab Index of less than one indicates that a sample was a normal (Ive negat: non-target influenza antigens were detect in the sample and d) normal sample; an Ab Index greater than one indicates that a specimen was a positive specimen (positive: target influenza antigens were detected in the specimen). The column labeled "Remel FDA" displays the results of the Remel FDA kit on samples reported as positive (+): influenza A detected, negative ( ■ ): mfluenza A not detected, or "NT": not tested.
[00376] [00376] The results of these Influenza A clinical evaluation experiments showed that all samples with influenza A antigens tested positive for Influenza A, while normal samples and samples with influenza B antigens did not test positive for Influenza A; These results are in agreement with the results obtained with the Remei FDA kit.
[00377] [00377] The clinical evaluation of the Influenza B assay: The performance of the Influenza B assay using the sample processing and analysis devices and systems as described herein was compared with the Remel FDA kit. taxi concentration was 5 µg/ml, while DAB concentration was maintained at 50 ng ml (final concentration after the protocol was run). For NIBSC influenza strains, 50 µl of sample was added to the swab, and the swab was then treated as a sample swab. For the Zeptometrix panel controls (pre-diluted samples), 200 µl of sample was mixed with 200 µl of extraction buffer. Swabs were placed in 500 µl of extraction buffer and incubated for 3-5 minutes. This extracted sample was then analyzed using the devices and systems as disclosed herein. Swabs and specimens were processed in the Remei FDA kit as indicated in the kit instructions. As discussed above, the cut-off value was defined as equal to the mean (normal) plus 4.5 x standard deviation (normal), and the Ab Index was calculated by dividing the mean RLU by the cut-off value. The column labeled "Remel FDA" displays the results of the Remel FDA kit on samples reported as positive (+): influenza B detected, or negative (-): influenza B not detected.
[00378] [00378] The results of these influenza B clinical evaluation experiments showed that all influenza B antigen samples tested positive for influenza B, whereas normal samples and influenza A antigen samples did not test positive for influenza B; These results are in agreement with the results obtained with the Remei FDA kit.
[00379] [00379] Other examples of markers indicative of infectious diseases that can be detected, identified, and analyzed through the methods, systems and devices described herein are shown in Figs. 22-
[00380] [00380] Fig. 23A shows an influenza panel naming various types of influenza that can be identified by the methods and devices discussed here. This figure relates to detection of various types of influenza by the nucleic acid detection methods discussed herein. In this, and following figures, and other parts of the application, "LOD" indicates "detection limit". Influenza types can be detected at the levels indicated in the figure; For example, influenza A can be detected by the methods, devices and systems disclosed herein when present in a sample at less than 100 copies per microliter (c/^L, where copies refer to copies of the nucleic acid sequence target indicative of influenza A) . As indicated in Fig. 23 A, other types of influenza such as influenza B and influenza H1N1 (seasonal) can be detected at levels of less than 10 copies per microliter.
[00381] [00381] Fig. 2.3B shows the inflection times for various types of influenza that can be identified by the methods and devices discussed herein. Influenza target nucleic acids indicative of the named influenza types were tested at 100 copies per microliter, and the time (from the start of the nucleic acid detection assay) in minutes, until the detection is displayed (as detected by the inflection of the RFU output as shown, for example, in the previous figures).
[00382] Fig. 24A shows various respiratory disease panels naming types of respiratory diseases, which can be identified by the methods and devices discussed herein. The detec tion limit (LOD) is indicated for each of the respiratory diseases in the rightmost column of the figure: LODs were either 10 copies per microliter (c / uL) or 100 C / ul.
[00384] [00384] Fig. 25A shows several hospital-acquired infectious diseases (indicated by the acronym "HAT") of nomenclature diseases that can be identified by the methods and devices discussed here. The limit of detection (LOD) is indicated for each HAI disease in the rightmost column of the figure; all LODs were 10 copies per microliter (C/uL).
[00385] [00385] Fig. 25B shows the inflection times for various hospital-acquired infectious disease panels of nomenclature types of respiratory diseases, which can be identified by the methods and devices discussed here. Target nucleic acids indicative of the named HAI diseases were tested at 100 copies per microliter, and the time (from start of nucleic acid detection assay) in minutes, until detection is displayed (as detected by the inflection of the R output. FU).
[00386] [00386] Fig. 26 shows results of a nucleic acid assay as described herein (see also descriptions of these methods, for example, in US patent applications 61/800,606; 61/908,027; 62,001, 050, and 14/214,850 ) for influenza A matrix protein that is designed to be included in all subtypes of Influenza a. The results are specific. Note that the inflection times for the "template" no control (CT) as well as for the influenza B targets were significantly longer than (and easily distinguishable from) the inflection times for the influenza A targets.
[00388] [00388] Fig. 28 shows that nucleic acid assays as described herein (see also descriptions of these methods, for example, in US Patent Applications 61/800,606; 61/908027; 62001050; and 14/214,850) are specific for seasonal influenza type H1N1 target. Note that the inflection times for influenza A ΗΓΝ1 (seasonal) targets were significantly shorter than (and easily distinguishable) the inflection times for the other influenzas and for the no-template control (NTC).
[00389] [00389] Fig. 29 shows potential interfering substances for nucleic acid assays as applied to the sexually transmitted disease (STD) panel, and concentrations at which these interfering substances were tested for interference with the assays. None of the indicated concentrations of potentially interfering substances interfered with nucleic acid assays.
[00390] [00390] Fig. 30 shows potential interfering substances for nucleic acid assays as applied to the Urine Sexually Transmitted Diseases (STD) panel, and concentrations at which these interfering substances were tested for interference with the assays. None of the indicated concentrations of potentially interfering substances interfered with nucleic acid assays.
[00391] Fig. 31 shows potential interfering substances for the nucleic acid assays, as applied to the blood panel, and that these concentrations of interfering substances were tested for interference with the assays. None of the indicated concentrations of potentially interfering substances interfered with nucleic acid assays.
[00392] [00392] While the above description is a complete description of the preferred embodiment as described herein, it is possible to use various alternatives, modifications and equivalents. Therefore, the scope of the present invention is to be determined not with reference to the foregoing description but should rather be determined with reference to the appended claims, together with the full scope of equivalents. Any feature, whether preferred or not, can be combined with any other feature, whether preferred or not. The appended claims are not to be construed as including means limitations -plus-functions, unless such limitation is explicitly recited in a given claim using the phrase "means to." It is to be understood that, as used in the description and all claims that follow, the meanings of "a", "an," and "the" include plural reference unless the context clearly dictates otherwise. Furthermore, as used in the present description and all claims which follow, the meaning of "on" includes "on" and "on" unless the context clearly dictates otherwise. Finally, as used in this specification and all claims that follow, the meanings of "and" and "or" include both the conjunctiva and disjunctive and may be used interchangeably unless the context expressly indicates otherwise. Thus, in contexts where the terms "and" or "or" are used, the use of such conjunctions does not exclude an "and/or" meaning unless the context expressly indicates otherwise.
[00393] [00393] This document contains material subject to copyright protection. The copyright owner (Applicant here) has no objection to facsimile reproduction by any person of the patent document or patent disclosure as they appear in the Patent and Trademark Office Fife or LIS patent records, but otherwise reserves All copyrights whatsoever. The following notice applies: Copyright 2013-2014 Theranos, Inc.

权利要求:
Claims (16)
[1]
1. A method for detecting infectious diseases, characterized in that it comprises: a) introducing a cartridge comprising at least two different types of samples into an automatic sample processing device, wherein one of said samples comprises a swab sample carried on a swab , wherein said automatic sample processing device comprises: - a sample handling system configured to transport at least a portion of one of said samples and being configured to transport a mobile testing unit independently; and - an optical detector; b) contacting one of said samples, or a portion thereof, with a mobile testing unit, or a reagent, or both, for carrying out an assay for the detection of a disease marker; c) positioning one of said samples, or part thereof, in a suitable location for detecting an optical signal of the sample or part thereof by said optical detector; d) detect the presence of a disease marker; and e) processing the other of said samples using said automatic sample processing device to perform a second assay to detect a second disease marker.
[2]
2. Method according to claim 1, characterized in that said smear is entirely contained in the cartridge.
[3]
3. Method according to claim 2, characterized in that said sample obtained using said swab is obtained by rubbing a mouth, throat, nasal passage, vaginal area or other body cavity of a subject.
[4]
4. Method according to claim 1, characterized in that one of said samples comprises a sample obtained using said smear, and another of said samples comprises a blood sample.
[5]
5. Method according to claim 1, characterized in that said sample has a volume of less than about 500 microliters.
[6]
6. Method according to claim 4, characterized in that it comprises the performance of two or more assays for the detection of disease markers and the detection of two or more disease markers in said samples, or in one or more portions thereof.
[7]
7. Method according to claim 6, characterized in that it comprises detecting the presence of a nucleic acid disease marker and a protein disease marker.
[8]
8. Method according to claim 1, characterized in that said disease marker is selected from a nucleic acid disease marker, a protein disease marker, a saccharide, a prostaglandin, a cytokine, histamine, a steroid and a marker for inflammation.
[9]
9. Method according to claim 6, characterized in that said two or more disease markers are selected from a nucleic acid disease marker, a protein disease marker, a saccharide, a prostaglandin, a cytokine, histamine, a steroid and an inflammation marker.
[10]
10. Method according to claim 1, characterized in that said disease marker is a marker of inflammation selected from prostaglandins, tumor necrosis factor alpha (TNF-α), interleukin-1 (IL-1 ), interleukin-8 (IL-8), interleukin-12 (IL-12), interferon gamma (IF-γ), bradykinin, complement system molecules, blood clotting factors, C-reactive protein, erythrocyte sedimentation rate (ESR), leukocyte count and morphological changes in blood and other cells.
[11]
11. Method according to claim 1, characterized in that the method is a point-of-service method performed at a point-of-service location.
[12]
12. Method according to claim 5, characterized in that the method is a point-of-service method performed at a point-of-service location.
[13]
13. Method according to claim 1, characterized in that the method can be performed in less than about 40 minutes.
[14]
14. Method according to claim 4, characterized in that the method can be performed in less than about 40 minutes.
[15]
15. Method for detecting infectious diseases, characterized in that it comprises: a) introducing a cartridge comprising at least two different types of samples into an automatic sample processing device, wherein one of said samples comprises a swab sample carried on a swab , wherein said automatic sample processing device comprises: - a sample handling system configured to transport at least a portion of one of said samples and being configured to transport a mobile testing unit independently; and - an optical detector; b) contacting one of said samples, or a portion thereof, with a mobile testing unit, or a reagent, or both, for carrying out an assay for the detection of a disease marker; c) positioning one of said samples, or part thereof, in a suitable location for detecting an optical signal of the sample or part thereof by said optical detector; d) detect the presence of a disease marker; and e) processing the other of said samples using said automatic sample processing device to perform a second assay to detect a second disease marker; h) wherein said one of said samples comprises a blood sample.
[16]
16. Method for detecting infectious diseases, characterized in that it comprises: a) introducing a cartridge comprising at least two different types of samples into an automatic sample processing device, wherein one of said samples comprises a swab sample carried on a swab , wherein said automatic sample processing device comprises: - a sample handling system configured to transport at least a portion of one of said samples and being configured to transport a mobile testing unit independently; and - an optical detector; b) contacting one of said samples, or a portion thereof, with a mobile testing unit, or a reagent, or both, for carrying out an assay for the detection of a disease marker; c) positioning one of said samples, or part thereof, in a suitable location for detecting an optical signal of the sample or part thereof by said optical detector; d) detect the presence of a disease marker; and e) processing the other of said samples using said automatic sample processing device to perform a second assay to detect a second disease marker; f) wherein said disease marker comprises a nucleic acid disease marker and said second disease marker comprises a protein disease marker.

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法律状态:
2018-05-08| B25A| Requested transfer of rights approved|Owner name: THERANOS IP COMPANY, LLC (US) |

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2018-11-06| B06F| Objections, documents and/or translations needed after an examination request according [chapter 6.6 patent gazette]|

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2020-07-28| B06U| Preliminary requirement: requests with searches performed by other patent offices: procedure suspended [chapter 6.21 patent gazette]|

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2021-10-13| B07A| Application suspended after technical examination (opinion) [chapter 7.1 patent gazette]|

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2021-10-13| B15K| Others concerning applications: alteration of classification|Free format text: AS CLASSIFICACOES ANTERIORES ERAM: G01N 35/00 , G01N 35/02 , G01N 33/48 , G01N 33/53 , G01N 33/569 , C12M 3/00 , A61B 10/00 , C12Q 1/68 , G06F 19/10 , G01N 33/15 Ipc: G01N 35/00 (2006.01), G01N 35/02 (2006.01), G01N 3 |

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2021-12-28| B25D| Requested change of name of applicant approved|Owner name: LABRADOR DIAGNOSTICS LLC (US) |

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2022-01-25| B25I| Requirement for requested change of headquarter|Owner name: LABRADOR DIAGNOSTICS LLC (US) Free format text: A FIM DE ATENDER A ALTERACAO DE ENDERECO CONFORME DOCUMENTOS APRESENTADOS ATRAVES DA PETICAO NO 870210077309 DE 23/08/2021, E NECESSARIO APRESENTAR UMA GUIA DE RECOLHIMENTO, CODIGO 248, ALEM DE ESCLARECER A DIVERGENCIA ENTRE O ENDERECO NO FORMULARIO E O ENDERECO NA DOCUMENTACAO APRESENTADA. ALEM DISSO, E PRECISO APRESENTAR A GUIA DE CUMPRIMENTO DE EXIGENCIA. |

优先权:

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申请号 | 申请日 | 专利标题

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US201361874976P| true| 2013-09-06|2013-09-06|

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US61/874,976|2013-09-06|

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US201361885462P| true| 2013-10-01|2013-10-01|

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US61/885,462|2013-10-01|

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US201462001039P| true| 2014-05-20|2014-05-20|

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US201462001053P| true| 2014-05-21|2014-05-21|

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US62/001,053|2014-05-21|

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US201462010382P| true| 2014-06-10|2014-06-10|

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US62/010,382|2014-06-10|

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PCT/US2014/054424|WO2015035260A1|2013-09-06|2014-09-05|Systems and methods for detecting infectious diseases|

---