 Methods of incorporating diaryltellulose into the gravure layer of anhydrous film immune assay eleme
专利摘要:
(A) a label layer comprising an enzyme labeled ligand or an enzyme labeled receptor; (b) a diffusion layer; (c) a receptor layer comprising a fixed receptor at a fixed concentration for the ligand, and (d) a support with a gravure layer comprising diaryl telluride. Preferred embodiments of the present invention include a vanadyl salt. The invention also relates to a method for performing an assay using an element as described above. 公开号:KR19990030045A 申请号:KR1019980039350 申请日:1998-09-23 公开日:1999-04-26 发明作者:마가렛 엘리자베쓰 로간；자넷 파일레스；스티븐 해셀버그 申请人:스타크 마이클；오르토-클리니칼 디아그노스틱스, 인코포레이티드； IPC主号:
专利说明:
Methods of incorporating diaryltellulose into the gravure layer of anhydrous film immune assay elements to reduce first slide bias and improve enzyme stability This application claims the benefit of priority under 35 USC §117, filed on June 7, 1995 under the name of Margaret E. Logan under the name Reduced first slide bias by incorporation of diaryltellulose in the foil film immunoassay element and improved enzyme stability U.S. Patent Application Serial No. 08 / 476,155. The present invention relates to anhydrous film elements and their uses in immunoassays. In particular, the invention relates to the improved stability of the labeled immunoreactive agent in anhydrous film elements. Immunity assays favoring natural immune responses have been widely used as analytical techniques in clinical chemistry. Due to the specificity of the reactants, they are particularly advantageous for the quantification of biological analytes present in very low concentrations in biological milks. Such analytes include, for example, antigens, antibodies, therapeutic drugs, anesthetics, enzymes, hormones and proteins. The assay that is the target of the assay is referred to herein as a ligand. A compound that specifically recognizes a ligand and reacts with it to form a complex is referred to herein as a receptor. Receptors and ligands form conjugate pairs. A particular number of pairs may act as a receptor or ligand. In the case of competitive assays, labeled assays containing labeled immunocompetent derivatives and homologs of these analytes are common elements of assays, whereas in the case of sandwich assays, labeled assays for analytes are routinely used . These are referred to herein as labeled ligands and labeled receptors, respectively. In competitive binding immunoassays, the labeled ligand competes with an unlabeled ligand to react with a fixed amount of the appropriate receptor. Ligands of unknown concentration can be determined from signal measurements of bound or unbound (i. E., Liberated) labeled ligands. This reaction proceeds as follows: Ligand + labeled ligand + receptor-substrate = Ligand-receptor-substrate + labeled ligand-receptor-substrate. In an alternative immune assay format known as a sandwich immunoassay or immunoassay assay, the ligand contacts two or more receptor molecules that bind to the ligand at different epitope sites. One receptor is normally properly labeled and the other receptor may be immobilized on a solid substrate or immobilized on a solid substrate. The amount of ligand is directly proportional to the amount of ligand and bound complex of the two receptors. This is illustrated as follows: Substrate - receptor 1 + ligand + receptor 2 - label = Substrate - receptor 1 - ligand - receptor 2 - label Common labels include radioactive tags, enzymes, chromophores, fluorophores, stable free radicals and enzyme cofactors, inhibitors and allosteric effectors. Immunoassay assay elements are known from U.S. Pat. Nos. 4,517,288 and 4,258,001. Generally, these elements include receptors such as antibodies for ligands that are immobilized within the particulate layer. In addition, the elements generally contain a reagent system that produces a signal that can be associated with the concentration of the ligand in the sample through the bound or unbound species. In use, the sample can be applied to the element in combination with an enzyme labeled ligand. After the passage of time, a solution containing the substrate for the labeled ligand is applied to the particulate layer. The reaction with the substrate is catalyzed by an enzyme label to form a reaction product that will eventually give rise to a signal, for example, a color. The reflectance density of the color may be correlated with the concentration of the ligand in the sample. Similar signal generation systems are known for other known conventional labels such as radioactive taches, chromophores, fluorophore, stable free radicals and enzyme cofactors, inhibitors and allosteric effectors. The multilayer immunoassay element is an anhydrous film element that measures the analyte in the emulsion sample using the immunoassay principle described above. For a competitive assay element, the hue (or other signal) formation rate is inversely proportional to the amount of analyte present, and for a sandwich assay element, the hue (or other signal) formation rate is directly proportional to the amount of analyte present. In addition, the hue (or other signal) formation rate is directly proportional to the activity of an enzyme-labeled analyte, the enzyme-labeled receptor bound to a drug or immobilized receptor. In immunoassays to maintain stable regulation, enzyme activity (rate of measurement) can not be lost within any part of the slide during a particular adjustment period. Often, an immunoassay element is applied to a user with a plastic cartridge containing 50 distinct elements that can be removed at the time one element is needed. The elements are stacked one on top of the other so that the lower 49 elements in the cartridge all have an upper surface applied by the element. However, the top element in the sediment is not so applied, so its surface is exposed to environmental factors as opposed to the other 49 elements. For example, the upper (or first) element is more exposed to air flow and light than the remaining elements if the cartridge is handled during manufacture or if the cartridge is present in the elemental feeder of the clinical analyzer. During storage prior to use, the cartridge itself is stored in a bag treated with a sealed foil. However, the top element is still more exposed to the residual air and humidity inside the sealed bag than the other 49 elements. If the overnight test milks react with the elements in the cartridge, the color forming rate observed at the top (or first) element is less than the observed color formation rate when the same test fluid is applied to the elements below the top element in the same cartridge It has been found to be low. This is referred to as the first slide bias. One method of dealing with a lower color forming rate of the first slide involves using vanadium ions as described in U.S. Patent No. 5,516,645 to Daniel et al. Incorporated herein by reference. Applicants have developed the use of diaryltelluride (DAT) or multiple diaryltellurides (DATs) to reduce the first slide bias. The use of diaryltoluide to prevent first slide bias can be used in both sandwich assays and competitive immunoassay assays. According to one aspect of the invention, (a) a label layer comprising an enzyme-labeled ligand or an enzyme-labeled receptor; (b) a diffusion layer; (c) a receptor layer comprising a fixed receptor at a fixed concentration for the ligand and and (d) a support having a gravure layer comprising diaryl telluride. In one embodiment, the acceptor is covalently bound to a polymeric bead having a diameter in the range of 0.1 to 5 mu m. In a preferred embodiment, the enzyme labeled ligand and diaryltelluride are present in the same gravure layer or zone wherein the gravure layer or zone abuts or is in contact with the surface of the diffusion layer that first or initially contacts the liquid sample . The defined element substantially reduces the first slide bias, i. E., Changes the color development rate of the top element in the cartridge as compared to other elements in the same cartridge. Moreover, all elements in the cartridge exhibit greater long-term stability and shelf life. The embodiments have established that diaryltelluride compounds provide this advantage. In addition, DAT compounds stabilize conjugates of enzyme compositions, generally in particular of peroxidases, especially horseradish peroxidase (HRP) and enzymes, preferably HRP and small molecules such as therapeutic drugs, abuse drugs and steroids useful. Another embodiment is a conjugate of an enzyme, preferably HRP, with a macromolecule such as a protein and a carbohydrate, wherein such an enzyme is used as a label. For example, the compounds may be used in solution assays and in anhydrous components of the present invention. Another aspect of the present invention is (1) A kit comprising: (a) a label layer comprising an enzyme labeled ligand or an enzyme labeled receptor; (b) a diffusion layer; (c) a receptor layer comprising a fixed receptor at a fixed concentration for the ligand and (d) providing an anhydrous immunoassay analyzing element comprising a support having a gravure layer comprising diaryltelluride; (I) a fixed ligand-receptor complex, (ii) a fixed enzyme-labeled ligand-receptor complex or (iii) a complex of (i) Forming a mixture, or a fixed receptor-ligand-labeled receptor complex; (3) catalyzing the evolution of the signal by contacting the substrate solution to the defined region and the enzyme label, and (4) measuring the concentration of the ligand by detecting the signal and measuring the intensity or amount of the signal. In this method, the enzyme labeled receptor or enzyme labeled ligand is separated from the fixed enzyme labeled ligand or immobilized enzyme labeled receptor. This separation can be carried out by the addition of certain means known in the art, for example, a substrate solution (e.g., hydrogen peroxide solution). One or more layers of the element may comprise one or more zones. The contact layers of the elements can have sharp boundaries with each other, where by optical and / or electron microscopy they show a distinct and separate element layer of the elements and they can also have a contact zone or contact area, And the boundary between the layers is blurred or unclear. In this case, distinct distinct layers may be visible or invisible by optical and / or electron microscopy. The formation of the zone is partially or substantially complete. In the case of partial zone formation, only a portion of the precipitated layer penetrates and mixes with a separate deposited contact layer (in this case, all of the layers may be interpenetrating each other, but in any case, (And all of the individual and phase layer infiltration are considered, all resulting in zone formation as described herein), and zones in separate deposited contact layers (or layers), wherein the layers are distinctly distinct Lt; RTI ID = 0.0 > or < / RTI > On the other hand, most or all of the discrete deposited layers may penetrate the zones (or zones) within the contact layer (or layers) in such a way as to be substantially or entirely contained in the contact layer (or layers) Thus it may not be visible as a distinct or distinct layer separated from the contact layer (or layers) of the element by optical and / or electron microscopy. A partial or complete zone may be formed or necessarily formed as intended. If intentionally formed, zones within the layer may be provided by any means that allows the practitioner to incorporate particular elements or elements or to be included within a zone within one or more layers. For example, specific or specific elements may be enclosed within a polymer or other matrix, which is then incorporated into the layer or layers. On the other hand, another aspect of the present invention includes a layer comprising specific element A (or elements) that can be dissolved in one of the contact layers by being in contact with two adjacent layers that can be made. Initial contact or subsequent successive complete or partial zones are formed in the contact layer in which they are soluble. Thus, the zones within the contact layer comprise a specific element A. The formation of zones may be partial or complete as defined above. Zone formation can be controlled by adjusting conditions such as solvent used to provide the layer, contact time of the pre-drying layer, temperature, etc., as is known to those skilled in the coating arts. Immunoassay elements defined using the terms label layer, receptor layer, gravure layer and / or other layers may be used during deposition or deposition of the indicator layer, receptor layer, gravure layer and / (Or layers) with a distinct distinct layer, or any aspect capable of forming a partially or complete zone (or zones). The gravure layer is a thin layer comprising one or more elements deposited on another layer, e. G., The surface of the diffusing layer. The term gravure layer used refers to a thin layer where the thickness of the layer is used to mean that the thickness of the layer is less than the thickness of the contact layer (or layers) that it is in contact with. Conveniently, the gravure coating method represents a preferred means of coating a thin layer comprising diaryl telluride (and / or other elements). The use of this term is not intended to be limited in any way. As defined above, means of coating, depositing or applying a thin layer are contemplated to fall within the scope of the term gravure layer. The gravure layer directly coated on the diffusion layer of the element of the present invention may be distinct on the diffusion layer or may not be present as a separate layer and the gravure layer may be present on the surface of the first contacting diffusion layer Or it is expected to form a thin contact zone directly underneath. In any case, when in contact with a diffusion layer comprising a gravure layer and / or a zone, the liquid sample is directly coupled to diffusion layer elements, beads, etc. (these terms are known in the art) due to the porosity of the diffusion layer So as to spread and meter the sample. That is, the contact of a liquid sample with an element having a gravure layer and / or zone can simply be described in terms of the contact of the liquid sample with the diffusion layer, since all other operations of the diffusion, metering and diffusion layers are unimpeded. Te5, defined below, represents one of the many species of Formula I as described in this patent. Ar 1 -Te-Ar 2 Furthermore, the present inventors have found that diaryltellurides, Of Te5 results in a significant reduction in the first slide bias when the immune assay element of carbamazepine (CRBM) competitive immuno challenge assay is incorporated into the gravure layer at a final rewet level of 1 mM. The present inventions have found that the first bias is significantly reduced over the course of a year. In fact, the first slide is not distinguishable from the non-first slide. It is not sufficient to prevent the generation of the first slide bias when the same level of Te5 is directly coated in the bead diffusion layer instead of the gravure layer coated in the bead diffusion layer. Incorporation of 0.25 to 0.50 mM Te5 (in re-wetting of the element) in the gravure layer of the anhydrous immune assay element designated for carbamazepine (CRBM), phenobarbital (PHBR), phenytoin (PHYT) and digoxin It was found that the first slide bias was significantly reduced even after the refrigerator was stored for 9 months. Generally, the present invention relates to an improved anhydrous immune assay element comprising a diarylteralide compound coated within a gravure layer arranged in a diffusion layer. However, the via layer can be arranged in any layer. In another aspect, the invention relates to a method of performing immunoassays using an improved dry-film element. Figure 1 if the buffer only, 1mM VOSO 4 and spotting a 1mM variety of solutions, including telluride each with 1mM VOSO 4 in the same described for each diaryl telluride alone or in a third embodiment of 1mM in dry format model A bar graph showing HRP stability. FIG. 2 is a bar graph showing HRP stability over a 24 hour period in a dry format model using diaryltellurides 1 and 4 at various concentrations as described in Example 3. FIG. 3 is a bar graph showing the stability of an anti-CRP-HRP conjugate in a dry format model using different concentrations of diaryltellurides 4 and 5 incorporated in a spotting solution as described in Example 5. FIG. Elements of the invention include a labeled ligand or labeled receptor, a diffusion layer (or zone), a receptor layer (or zone), and a gravure layer (or zone). The zones and layers were defined earlier. Various zones can be present in one coating layer or in a separate coating layer. For example, the diffusion zone and the receiver zone may be a single layer or may be separate layers. The separate layers can be arranged in any order on the support. Separate layers can also be arranged in such a way that the receptor layer is present directly on the support, the diffusion layer is present directly on the receptor, or the labeled ligand or labeled acceptor zone is present directly on the diffusion layer. If the receptor sites form an entirely separate layer, these layers will also include a binder of the type described below. The element may include additional layers such as a gravure layer. All of these layers can be coated using coating techniques known in the art. Representative compounds of diaryltellurides have formula (1). Formula 1 Ar 1 -Te-Ar 2 Diaryltelluride is incorporated into immune assay elements, including horseradish peroxidase-labeled immunoreactive agents, such as human chorionic gonadotropin (hCG) and C-reactive protein (CRP) The first slide bias is significantly reduced. Some of these compounds are tested for conservation of HRP activity in a model system designed to evaluate compounds against their ability to prevent first slide biases. Useful diogan Nortel rides are those wherein Ar 1 and Ar 2 are the same or different aryl or heteroaryl groups of the formula: In this formula, X is O, S, Se or Te, R 11 , R 12 , R 13 , R 21 , R 22 , R 23 , R 31 , R 32 , R 33 , R 41 , R 42 and R 43 are the same or different and each is hydrogen, , OH, OR 1 , SH, NH 2 , NHR 1 , NR 1 2 , NR 1 R 2 , CO 2 H and salts thereof, CO 2 R 1 , SO 3 H and salts thereof, PO 3 H 2 and salts thereof, And SR 1 wherein R 1 and R 2 are different and each is selected from the group consisting of alkyl, phenyl and substituted phenyl having a carbon chain having from 1 to 14 carbon atoms, with or without one or several hydrophilic groups ), ≪ / RTI > R 14, R 15, R 24 and R 25 are the same or different, each represents hydrogen, alkoxy, CO 2 H or a salt thereof having a carbon number of 1 to 5 alkyl carbon atoms, 1 to 5, and CO 2 R 1, SO 3 H and And salts thereof, PO 3 H 2 and salts thereof, wherein R 1 is as defined above. In the above, alkyl means a group such as methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, amyl, isoamyl or neopentyl. The carbon chain having 1 to 5 carbon atoms means a straight or branched carbon chain such as methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, t-butyl, amyl, isoamyl or neopentyl. The carbon chain of 1 to 14 carbon atoms includes methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, tertiary butyl, amyl, isoamyl, neopentyl, hexyl, octyl, decyl, But is not limited thereto. Hydrophilic group means a group such as sulfuric acid, phosphoric acid or carboxylic acid, hydroxyl and amino group. Some of these compounds may form salts with acids or bases. Salts with sodium, potassium, ammonium, calcium and magnesium salts and with organic acids such as hydrochloric acid, bromic acid, phosphoric acid and sulfuric acid and oxalic acid, fumaric acid, tartaric acid, malonic acid, acetic acid, citric acid and succinic acid are preferred. Preferred DATs are those that have substituents on aromatic rings containing hydroxyl groups, amines and salts thereof, and carboxylic acid and salts thereof, such as, but not limited to, groups that impart water solubility. In particular, the water-soluble DAT that preserves HRP activity, either alone or in combination with a vanadyl compound, is (Te1-Te5): Other materials for use in the diffusion layer are well known in the art, for example in U.S. Patent No. 4,258,001, to produce anhydrous analytical elements as described. These layers include macroporous layers made from cloth, paper, and the like. The preferred particulate layer is a bead diffusion layer (BSL). This layer can be readily prepared to receive a test sample (i.e., 1 to 200 μL of sample) by having porosity suitable for use in the elements of the present invention, and can be diluted or undiluted. Preferably, the diffusing layer is isotropic porosity characteristically produced by inter-connected spaces between particles comprising the zone. The isotropic porosity means that the diffusion layer uniformly disperses the fluid applied in all directions through the layer. Useful diffusion layers comprising bead diffusion layers are described in U.S. Patent Nos. 4,670,381, 4,258,001 and 4,430,436. Particularly useful diffusion layers are those having a particulate structure formed by organic-polymeric particles and polymeric adhesives (these particles are described in U.S. Patent No. 4,258,001). Organic-polymeric particles useful in the diffusion layer are generally spherical beads of thermal stability with a particle size in the range of about 10 to 40 占 퐉 or even a smaller diameter. The particles can be composed of a wide range of organic polymers including natural and synthetic polymers with essential properties. Preferably, however, they are composed of one or more addition polymers as described in the foregoing patents. When the receptor layer is a separate layer, it is prepared and coated on a support or on a reagent layer or a lower layer on a support. The receptor is covalently bound to the polymer particle through a surface reactive group on the acceptor (nucleophilic free amino group and sulfhydryl group). A common method for attaching a receptor to an oligomeric bead involves covalently linking the selected receptor to the bead using generally known reactions. Many pendant groups, such as haloalkyl, 2-substituted activated ethylsulfonyl and vinylsulfonyl, can be used to directly bond the acceptor to the beads. Generally, the beads are mixed with the acceptor in a buffered aqueous solution, wherein the pH is generally about 5 to 10, and the concentration of the polymer particles is about 0.1 to about 40 wt%, preferably about 0.1 to about 10 wt%. The amount of receptor is from about 0.1: 1000 to about 1: 10, preferably from about 1: 100 to about 1: 10, for the polymer. The mixing is carried out at a temperature ranging from about 5 to about 50 캜, preferably from about 5 to about 40 캜, for about 0.5 to about 48 hours. Any suitable buffer solution may be used. In some cases, the pendant reactive groups on the outer surface are modified or activated to induce covalent bonding of the ligand. For example, the carboxyl groups are activated by using the known carbodiimide or carbamoylonium chemistry described in EP 308235, published July 22, 1992, and U.S. Patent No. 5,155,166. However, binding of the receptor to the carboxyl group-containing monodisperse polymer beads is carried out in two steps. The first step involves contacting the aqueous suspension of particles with a carbodiimide or a < RTI ID = 0.0 > vucamoylonium < / RTI > compound to produce an intermediate polymer particle having an intermediate reactive group instead of a carboxyl group. This step provides a suitable pH by carrying out using a suitable pH with a suitable acid or buffer. Generally, the pH is less than 6, but this is not critical as long as the reaction can proceed. More preferably, the pH is from about 3.5 to about 7. The molar ratio of carbodiimide or carbamoylonium compound to carboxyl groups on the particle surface is from about 10: 1 to 500: 1. In a second step of the method, the reactive intermediate formed during the first step is contacted with a reactive amine- or sulfhydryl-group containing receptor. Whereby a covalent bond is formed between the particle and the acceptor. The weight ratio of receptor to polymeric particles is generally from about 1: 1000 to about 1: 1, preferably from about 1: 100 to about 1: 10. As another example, the epoxy group on the outer surface can be hydrolyzed to form a diol compound capable of reacting with cyanogen bromide which can act as a coupling agent for the amine group in the immunological species. The aldehyde reacts directly with the amine to form a schiff base which can be covalently bound to form a covalent bond. The aldehyde can be oxidized to the acid and the amide bond can be formed using the chemistry defined above for the carboxyl group. As long as the intermediate contains the reactive amine or sulfhydryl group to be reacted with the intermediate formed by the reaction of the carboxyl group on the particle with the carbodiimide or the carbamoyl compound, if the reactive group or polymer on the polymer has a carboxyl group, Any reactive amine- or sulfhydryl-containing receptor can be conjugated to the monodispersed polymeric beads. Small polymeric beads having reactive groups that readily react directly with amines or sulfhydryl groups on the receptor are sometimes simply mixed and reacted with the receptor in an appropriate buffer solution. Polymers that may be selected as beads for the receptor include: poly (m p-chloromethylstyrene), poly (styrene-co-m p-chloromethylstyrene-co-2-hydroxyethyl acrylate) Poly (styrene-co-m-p-chloroethylsulfonylmethylstyrene) (95.5: 4.5 molar ratio) Methacrylic acid) (95: 5, 98: 2 and 99.8: 0.2 molar ratio), poly (styrene-co-m-methylstyrene-co-methacrylic acid) styrene-co-N- [m p- (2-chloroethylsulfonylmethyl) phenyl] acrylamide-co-methacrylic acid) (93.5: 4.5: Co-methacrylic acid} (97.3: 0.7: 2 molar ratio), poly (styrene-co-m-p-chloromethylstyrene) (70:30 molar ratio) Propionic acid] (97.6: 2.4 molar ratio), poly (styrene-co-vinylbenzyl chloride-co-acrylic acid) (85: 10: 5 molar ratio) (Styrene-co-acrylic acid) (99: 1 molar ratio), poly (styrene-co-methacrylic acid) (90:10 molar ratio) Acrylic acid) (70:30 molar ratio), poly (styrene-co-2-carboxyethyl acrylate) (90:10 molar ratio) (95: 5 molar ratio) and poly (styrene-co-m-p-vinylbenzaldehyde-co-methacrylic acid) (93: 5: 2 molar ratio). The layers of the element are carried on a suitable support. The receptor layer may be coated on the support even though an intermediate layer such as a gelatin / buffer layer may be present between the support and the receptor layer. The support is suitably scale-stable and preferably non-porous and transparent (i.e., radiation-permeable) material that allows electromagnetic radiation of about 200 to about 900 nm to pass. The support for a particular element should be chosen to match the required detection mode (reflection, transmission, luminescence or fluorescence spectroscopy). Useful support materials include polystyrene, polyesters (i.e., poly (ethylene terephthalate)), polycarbonate cellulose esters (i.e., cellulose acetate), and the like. Polymeric binders for the support layer are generally described in Canadian Patent No. 1,240,445, incorporated herein by reference. Useful polymers are polymers comprising from about 30 to about 97 weight percent polymerized N-alkyl substituted acrylamide, such as N-isopropylacrylamide. Other useful N-alkyl-substituted acrylamides include N-n-butyl acrylamide, N, N-diethylacrylamide and N-n-propylacrylamide. Poly (N-isopropylacrylamide-co-methacrylic acid-co-N, N'-methylenebisacrylamide) is used as an example to list the use of these binders. The polymeric binder also comprises from 3 to 25% by weight of at least one polymerized cross-linking monomer, such as at least two additive-polymerizable groups per molecule. These crosslinking monomers are generally well known in the art. Preferred cross-linking monomers include acrylamido or methacrylamido groups that are easy to polymerize with N-alkyl-substituted acrylamides. Examples of useful crosslinking monomers include: N, N'-methylenebisacrylamide; N, N'-methylenebis methacrylamide; Ethylene dimethacrylate; 2,2-dimethyl-1,3-propylene diacrylate; Divinylbenzene; Mono [2,3-bis (methacryloyloxy) propyl phosphate; N, N'-bis (methacryloyl) urea; Triallyl cyanurate; Allyl acrylate; Allyl methacrylate; N-allyl methacrylamide; 4,4'-isopropylidenediphenylene diacrylate; 1,3-butylene diacrylate; 1,4-cyclohexylenedimethylene dimethacrylate; 2,2'-oxydiethylene dimethacrylate; Divinyloxymethane; Ethylene diacrylate; Ethylidene diacrylate; Propylidene dimethacrylate; 1,6-diacrylamidohexane; 1,6-hexamethylene diacrylate; 1,6-hexamethylene dimethacrylate; Phenylethylenedimethacrylate; Tetramethylene dimethacrylate; 2,2,2-trichloroethylidene dimethacrylate; Ethylene bis (oxyethylene) diacrylate; Ethylene bis (oxyethylene) dimethacrylate; Ethylidene trimethacrylate; Propylidine triacrylate; Vinyl allyloxyacetate; 1-vinyloxy-2-allyloxyethane; 2-crotonoyloxyethyl methacrylate; Diallyl phthalate and 2- (5-phenyl-2,4-pentadienoyloxy) ethyl methacrylate. These polymeric binders also contain from 0 to 60% by weight of polymerized hydrophilic monomers. Amounts of 5 to 35% by weight are also useful. Hydrophilic monomers are described in Canadian Patent 1,240,445. In particular, such monomers have at least one group selected from the group consisting of hydroxy, pyrrolidone, amine, amide, carboxy, sulfo, carboxylate salt, sulfonate salt and sulfate salt groups. In general, the counter ion of the salt group is an alkali metal or ammonium. Useful hydrophilic monomers include acrylic acid and methacrylic acid and salts thereof, sodium 2-acrylamido-2-methylpropanesulfonate, 2-hydroxyethyl acrylate, 2-hydroxyethyl methacrylate, 2- Acrylate, 2-hydroxypropyl methacrylate and glyceryl methacrylate. In addition, the cited binders enable a uniform coating of the receptor layer due to the lower binder viscosity achieved with shear thinning during extrusion hopper coating. A further advantage is achieved with the use of the cited binders, wherein, as soon as a uniform coating is formed, the viscosity of the binder is substantially increased to provide a stable and uniformly present set layer during wet transport and drying of the binder, . The receptor may also be a poly (vinyl alcohol); Bovine serum albumin; - acacia sword; A homopolymer of N-vinylpyrrolidone having a molecular weight of 8000 to 400,000; Alkyl-substituted acrylamides, n-alkyl substituted methacrylamides, 1-vinylimidazole, 2-alkyl substituted -1-vinylimidazole, 2-hydroxyalkyl A water-soluble vinyl-containing copolymer having two or more monomers selected from the group consisting of substituted-1-vinylimidazole, n-vinylpyrrolidone, hydroxyalkyl acrylate, hydroxyalkyl methacrylate, acrylic acid and methacrylic acid (Where the alkyl and hydroxyalkyl in the copolymer have 1 to 6 carbon atoms, such as methyl, ethyl, propyl, and hexyl). The element may comprise a gelatin / buffer layer containing one or more additional layers, i. E. A separate or combined reagent / dispersion layer and other essential additives such as an electron transfer agent. The gelatine / buffer layer or reagent layer or diffusion layer of the urea can comprise one or more synthetic or natural binder materials such as gelatin or other naturally occurring colloids, homopolymers or copolymers such as poly (acrylamide), poly (2-pyrrolidone), poly (N-isopropylacrylamide), poly (acrylamide-co-N- vinyl-2-pyrrolidone) and similar copolymers) ≪ / RTI > The indicator composition may also be dispersed in the receptor layer. Other optional layers may be included, for example, a lower layer, a radiation-blocking layer, and the like. All layers of the element are in contact with one another in the emulsion, which means that the reagents and uncomplexed reaction products in the emulsion and emulsion can pass between the overlapping areas of adjacent layers. The layers of the element may contain various other desirable but optional components including surfactants, thickeners, buffers, hardeners, antioxidants, bonding solvents and other materials known in the art. The amount of these elements is also known to those skilled in the art. Factors can be used to measure low concentrations of immunologically reactive ligands in liquids such as biological milieu (eg, white blood cells, serum, plasma, urine, semen, human or animal tissue suspensions, excreta, saliva, have. This ligand can be measured at a low concentration of about 10 -15 moles, and more usually at a concentration of about 10 -11 to about 10 -4 moles. Ligands that can be measured qualitatively or qualitatively include therapeutic agents such as phenobarbital, digoxin, digethoxin, terofilin, gentamycin, quinidine, phenytoin, propanolol, carbamazepine, tobramycin, lidocaine, Progesterone, estriol), hormones (e.g., thyroid hormone, peptide hormone, insulin, etc.), proteins (e.g., albumin, IgG, IgM , Antibodies (e. G., Monoclonal antibodies), and other species that will naturally react with the receptor. ≪ / RTI > The present invention is particularly useful for the determination of analgesics such as therapeutic drugs such as digoxin, phenytoin, carbamazepine, theophylline or phenobarbital, hormones such as tyoxyne or triiodothyronine, and hCG and C-reactive proteins. This assay can be performed using an enzyme label capable of binding to the ligand to form the labeled ligand. Glucose oxidase, peroxidase such as horseradish peroxidase (HRP), alkaline phosphatase and galactosidase, and the same enzyme are preferable labels. Those skilled in the art of clinical chemistry can measure suitable substrates for a given label. Substrates can be substances that act directly by an enzyme label or by a substance involved in a series of reactions involving an enzymatic reaction of the label. For example, if the enzyme label is a peroxidase, the substrate is hydrogen peroxide and a suitable reducing agent. When glucose oxidase is used, the substrate glucose is generally present in the reactant layer or added as a substrate solution to yield about 0.01 mol / m 2 , preferably about 0.001 to about 0.1 mol / m 2 . Those skilled in the art are aware of how to control the amount of a particular substrate relative to the amount of enzyme label used during the assay. The reagent layer may comprise an indicator agent comprising one or more reagents that provide a detectable species as a result of the reaction catalyzed by the indicator. The detectable species may be color, radioactive, fluorescent or chemiluminescent. For this purpose, the present invention is demonstrated using a colorimetric indicator composition which provides a colorimetrically detectable species as a result of enzymatic reaction of the enzyme labeled ligand analog and substrate. The indicator composition may be a single compound that produces a detectable dye upon an enzymatic reaction or may be a combination of reagents that produce a dye. For example, where glucose is used as a substrate and glucose oxidase is used as an enzyme label, the colorimetric indicator composition may comprise a coupling compound and an oxidizable compound that react to provide a dye. This composition can include leuco dyes and peroxidase or other peroxidase-generating compounds that produce detectable dyes as a result of the formation of hydrogen peroxide produced when glucose oxidase converts glucose to gluconic acid. Useful leuco dyes are known in the art and are described, for example, in U.S. Patent No. 4,089,747 (granted to Bruksey on May 16, 1978) and U.S. Patent No. 4,670,385 (issued June 2, 1987 to Bob et al) Lt; / RTI > The specific amount of colorimetric indicator composition and its various components are within the skill of the art. Labeled ligands can be prepared or purchased using known starting materials and processes. Generally, a ligand is bound to a label (e.g., an enzyme residue) through a covalent bond. The immunoassay can be manual or automated. Generally, the amount of ligand in the liquid is measured by taking the element from the feed roll, chip packet, or other source and physically contacting the limiting site of the diffusion layer with the sample in 1 to 200 μL of liquid. The limited area that is contacted is generally less than about 150 mm 2 . The amount of ligand is determined by passing the element directly through an apparatus suitable for detecting the conjugated ligand analog or by passing a detectable species formed as a result of enzymatic labeling and enzymatic reaction of the substrate. For example, this species can be detected using a suitable spectrophotometric apparatus using generally known processes. In the enzymatic reaction, the obtained product is measured, for example, by measuring the transmittance density or the rate of change of reflection in the confined area in contact with the test sample. The measured area generally has a diameter of about 3 to about 5 mm. The amount of ligand in the liquid sample is inversely proportional to the amount of labeling measured within the confined area for competitive assays, or directly in the case of sandwich assays. In general, labeling is performed after application of the substrate solution. 1. Synthesis of Diaryltelluride There are many synthetic routes to DAT production. The route used for a particular compound is highly dependent on the substituent on the aromatic ring. Some known routes are listed in the Examples below. These and other references to the synthesis of other DATs will be apparent to those skilled in the art. Preparation 1. 4,4'-Di (2-hydroxyethoxy) -1,1 'tellurobisbenzene (Tel) (a) Preparation of [2- (4-bromophenoxy) ethoxy] (1,1-dimethylethyl) dimethylsilane (1a) To a solution (30 mL) of [2- (4-bromophenoxy) ethanol (10.0 g, 46 mmol) in 150 mL of anhydrous dimethylformamide was added tert-butyldimethylsilyl chloride (8.34 g, 55 mmol) and imidazole (7.82 g, 115 mmol). These materials are washed into the reaction flask using additional dimethylformamide (20 mL). The resulting solution is stirred overnight at room temperature using a Newman tube. It is poured into water (200 mL) and extracted with ether (3 X 75 mL). The combined extracts are washed with 0.5N HCl (200 mL), saturated NaHCO 3 (200 mL), water (4 X 150 mL) and saturated NaCl (200 mL), dried over MgSO 4 and filtered. The solvent is removed under reduced pressure on a rotary evaporator to give the crude product (16.5 g; 100%). 1 H NMR (CDCl 3) d 7.34 (2H, d, J = 8.9), 6.78 (2H, d, J = 8.9), 4.00-3.93 (4H, m), 0.89 (9H, s), 0.08 (6H, s). 13 C NMR (CDCl 3) d 158.1, 132.2, 116.4, 112.8, 69.6, 61.9, 25.9, 18.4. FDMS (m / e) 330 (M < + & gt ; , 79 Br). This is used as 100% without purification. (b) Preparation of 4,4'-di (2-hydroxyethoxy) -1,1'-tellurobisbenzene (Tel) A 500 mL three-necked flask is placed under argon and fitted with a condenser and addition funnel. Magnesium turnings (0.89 g, 37 mmol) are added. The bromide (1a) (12.3 g, 37 mmol, used as 100%) is placed in anhydrous tetrahydrofuran (THF) and transferred to the addition funnel. The Grignard reaction is initialized using approximately 10 mL of the bromide solution, 1,2-dibromoethane and iodine. The remaining bromide solution is added and the reaction is refluxed overnight while Mg is no longer remaining. After removing the lower mantle for 30 minutes, the tellurium granules (4.74 g, 37 mmol) are added, and the reaction is heated again for 7 hours to reflux, consuming almost all Te. The reaction is cooled to room temperature, poured into 10% aqueous NH 4 Cl (300 mL) with rapid stirring and stirred for 15 minutes. The precipitated Te is filtered off through celite diatomaceous earth and the filter cake is washed with ether. The filtrate is transferred to a separatory funnel and extracted three times with ether (200 mL, 100 mL, 100 mL). The combined extracts were washed with water and saturated NaCl, and then dried MgSO 4, and filtered on. The solvent was removed under reduced pressure to give crude product (12.4 g) as a red oil. This is added with copper powder (2.5 g) in toluene (50 mL) and heated to reflux for 2.5 hours in a Newman tube, where the color changes from red to gray. The reaction was cooled to room temperature, filtered through celite diatomaceous earth, washed with ether and concentrated to give an amber oil (12.4 g). ≪ 1 > H NMR, this is a mixture of the desired product, quenched Grignard and residual toluene. This is added to methanol (50 mL) and (50 mL), and THF (20 mL) and potassium fluoride (5.0 25, 86 mmol) and refluxed for 24 hours. A large amount of methanol is removed under reduced pressure and partitioned between ether-ethyl acetate and water. The aqueous phase is extracted twice more with ethyl acetate and the combined extracts are washed with water and saturated NaCl, dried over NaSO 4 and filtered. The solvent was removed under reduced pressure to give a crude product (8g) as a brown solid. A white solid is obtained which is separated by filtration with ether grinding, washing with ether and air drying to give the crude product (1.74 g). This is adsorbed onto flash silica gel (20 mL) using dichloromethane. Carefully flash chromatographed over 200 mL of silica gel, eluting with dichloromethane followed by 95: 5 dichloromethane: methanol (product eluted just before a small amount of the corresponding biaryl compound), ether polishing and filtration To give a pure white solid (1.2 g). The ether filtrate from the polishing of the crude product is chromatographed as described above. Ether grinding and recrystallization from ethanol gave additional pure Te1 (0.2 g). Total yield: 1.4 g, 19%, 1 H NMR (DMSO-d 6 ) d 7.52 (4H, d, J = 8.5), 6.80 ), 3.91 (4H, t, J = 5.0), 3.65 (4H, app q, J = 5.1). 13 C NMR (DMSO-d 6 ) d 159.2, 139.8, 116.5, 104.5, 69.9, 59.9. FDMS (m / e) 404 (M & lt ; + > , 130 Te). 2. Preparation of N, N-dimethyl-2- (phenyltelluro) benzenemethanamine, hydrochloride salt (Te2) (a) Preparation of N, N-dimethyl-2- (phenyltelluro) benzenemethanamine (2a) To a solution of N, N-dimethylbenzenemethanamine (2.70 g, 0.020 mol) in anhydrous ether (50 mL) in a round bottomed flask was dropwise n-BuLi (2.5 M, 10 mL, 0.025 mmol) at room temperature under argon with a syringe. The reaction mixture is stirred at room temperature for 5 hours, and then a solution of phenyltelulenyl bromide in anhydrous tetrahydrofuran (0.5 m) is added dropwise via syringe. After the addition of 38 mL (0.019 mol) of the reaction mixture, the addition stops when the reaction mixture becomes a characteristic orange color of phenyltelluryl bromide. The reaction mixture is poured into ether (100 mL) and the resulting solution is washed with saturated NaCl (1 X 100 mL, 2 x 50 mL), dried over MgSO 4 and concentrated. The residue was dissolved in acetone (100 mL) and iodine (5.08 g, 0.020 mol) was added. The resulting solution is allowed to cool to precipitate the product. The yellow crystals were collected by filtration, washed with cold acetone and dried to give the iodine adduct of 2a (5.95 g, 50%, mp 178-179 [deg.] C). The iodine additive (5.93 g, 0.010 mol) is dissolved in dimethylformamide (100 mL). Sodium sulfite (5.2 g, 0.05 mmol) in water (100 mL) is slowly added and the reaction mixture is stirred for 1 hour at room temperature, during which time the reaction mixture becomes colorless. The reaction mixture is poured into water (500 mL) and washed with ether (2 X 50). The aqueous layer is basified with 10% NaOH and the amine is extracted with ether (3 X 100 mL). The combined extracts were washed with saturated NaCl, then dried MgSO 4, and concentrated to above. The residue was recrystallized from methanol to give pure product (3.14 g, 93%) as a white solid (melting point 51 to 54 캜). 1 H NMR (CDCl 3) d 7.88 (2H, d, J = 6.9), 7.35 (1H, t, J = 7.3), 7.26 (2H, t, J = 7.3), 7.17 (1H, d, J = 7.7 ), 7.10-7.04 (2H, m), 6.94-6.89 (1H, m), 3.53 (2H, s), 2.25 (6H, s). (b) Preparation of N, N-dimethyl-2- (phenyltelluro) benzenemethanamine, hydrochloride salt (Te2) A solution of hydrogen chloride / ether (3.15 mL, 1.0 M, 3.15 mmol) was added to a slurry of 2- (N, N-dimethylaminomethyl) -1-phenyltellobenzene (1.02 g, 3.0 mmol) in ether (50 mL) Drop with a syringe. The viscous slurry is diluted with isopropanol (20 mL) and filtered. The white solid is washed with ether and air dried to give the product (1.13 g, 95%). 1 H NMR (CDCl 3) d 2.6 (1H, br s), 8.13 (1H, d, J = 7.5), 7.80 (1H, d, J = 7.7), 7.52-7.48 (3H, m), 7.26-7.17 (4H, m), 4.44 (2H, d, J = 5.8), 2.68 (6H, d, J = 4.5). Elemental analysis for C 15 H 18 ClNTe: Calculated: C, 48.00; H, 4.83; N, 3.73 Found: C, 47.97; H, 4.87; N, 3.65 3. Preparation of N, N, N ', N'-tetra (2-hydroxyethyl) -4,4'-tellurobisbenzene amine (Te3) (a) Preparation of N, N-bis [2 - [[(1,1-dimethylethyl) dimethylsilyl] oxy] ethyl] 1a is carried out using the following materials: 2,2 '- (phenylimino) diethane (9.0 g, 50 mmol), tert-butyldimethylsilyl chloride (18.1 g, 0.12 mol) Imidazole (17.0 g, 0.25 mol) and dimethylformamide (50 mL). The reaction is stirred overnight for convenience. (21.6 g, 100%) is obtained after work-up as in (1a). 1 H NMR (CDCl 3) d 7.21 (2H, d, J = 8.4), 6.68 (3H, OY member wrapping d, t), 3.77 (4H , d, J = 6.6), 3.52 (4H, d, J = 6.6), 0.92 (18H, s), 0.06 (12H, s). 13 C NMR (CDCl 3) d 147.8, 129.2, 115.7, 111.4, 60.3, 53.5, 25.9, 18.3, -5.3. FDMS (m / e). This is used as 100% without purification. (b) Preparation of N, N, N ', N'-tetra [2 - [[(1,1- dimethylethyldimethylsilyl] oxy] ethyl4,4'-telulobisbenzenamine (3b) Tellurium (IV) chloride (5.93 g, 22 mmol) is added to a 500 mL three-necked flask which is oven-dried and placed under argon. Anhydrous ether (100 mL) was added by syringe to obtain a pale yellow slurry and the flask was placed in a water bath. A solution of (3a) (18 g, 44 mmol) in dry ether (50 mL) is made under argon and the solution is transferred to the reaction mixture with vigorous stirring with a cannula. Initially very viscous yellow precipitate forms, which is less viscous with the addition of subsequent aniline solution. Upon completion of the addition, the greenish yellow slurry is stirred overnight. The reaction mixture is filtered and the filtrate is evaporated under reduced pressure. The resulting yellowish green oil was taken up in dichloromethane (200 mL) and a solution of sodium metabisulfite (8.36 g, 44 mmol) in water (200 mL) was added with stirring. The resulting biphasic mixture is stirred at room temperature for 30 minutes and then filtered through celite diatomaceous earth. Solid NaHCO 3 is added to pH 9 and the reaction mixture is transferred to a separatory funnel. The dichloromethane layer is separated and the aqueous layer is extracted with additional dichloromethane. The combined extracts are washed with water / saturated NaCl (200 mL / 50 mL) followed by sat. NaCl, then dried over NaSO 4 , filtered and the solvent is removed under reduced pressure. The resulting red oil (21.4 g) was placed in toluene (80 mL) and copper powder (4 g) was added. The resulting mixture is heated to reflux overnight using a Newman tube. The resulting gray reaction mixture is cooled to room temperature, filtered through celite diatomaceous earth and the solvent is removed under reduced pressure. The obtained amber oil (18 g) was flash chromatographed with 3: 1 cyclohexane: dichloromethane to separate the product from the recovered starting material to give pure (3b) as a yellow oil (5.0 g, 48% of theory). 1 H NMR (CDCl 3) d 7.54 (4H, d, J = 8.6), 6.53 (4H, d, J = 8.5), 3.72 (4H, t, J = 6.4), 3.47 (4H, t, J = 6.5 ), 0.88 (18H, s), 0.03 (12H, s). 13 C NMR (CDCl 3) d 147.6, 139.7, 112.7, 98.2, 60.2, 53.4, 25.9, 18.3, -5.3, FDMS (m / e) 946 (M +, 130 Te). (c) Preparation of N, N, N ', N'-tetra (2-hydroxyethyl) -4,4'-tellurobisbenzene amine (Te3) A heterogeneous mixture of silyl ether (3b) (2.7 g, 2.86 mmol), potassium fluoride (0.66 g, 11.4 mmol) and methanol (20 mL) is heated to reflux for 20 h. The white slurry was cooled in an ice bath and the product was isolated by filtration, washed with cold methanol and air dried (0.80 g, 57%). A small amount is recrystallized from methanol for analytical sample (melting point: 169-170 < 0 > C). 1 H NMR (DMSO-d 6 ) d 7.54 (4H, d, J = 8.6), 6.53 (4H, d, J = 8.5), 3.45 (4H, app q, J = 5.7), 3.33 (4H, t, J = 5.9) 13 C NMR ( DMSO-d 6) d 148.1, 139.8, 113.1, 97.6, 58.4, 53.5. IR (KBr) 3350, 1585, 1495, 1350 cm < -1 >. FDMS (m / e) 490 (M & lt ; + > , 130 Te). Elemental analysis for C 20 H 28 N 2 O 4 Te: Calculated: C, 49.22; H, 5.78; N, 5.74 Found: C, 48.78; H, 5.72; N, 5.66 4. Preparation of 2,2 '- [telulobis (4,1-phenyleneoxy)] bisacetic acid, disodium salt (Te4) (a) Preparation of 4- (Bromophenoxy) (1,1-dimethylethyl) dimethylsilane (4a) To a solution of p-bromophenol (104 g, 0.6 mol) in anhydrous dimethylformamide (450 mL) was added tert-butyldimethylsilyl chloride (108 g, 0.72 mol) and imidazole (102 g, 1.5 mol). The resulting pale yellow solution is stirred at room temperature for 3 hours using a Newman tube. It is poured into water (1.2 L) and extracted three times with ether (900 mL, 300 mL, 300 mL). The combined extracts are washed four times with water (150 mL), once with 1 N HCl (300 mL), saturated NaHCO 3 (300 mL) and saturated NaCl (300 mL). The solvent was removed under reduced pressure on a rotary evaporator followed by a vacuum pump to give a crude product (183 g, 100%) as a 6: 1 ratio from the hydrolyzed starting material to the silane. 1 H NMR (CDCl 3) d 7.31 (2H, d, J = 8.7), 6.71 (2H, d, J = 8.7), 0.97 (9H, s), 0.18 (6H, s). This is used as 100% without purification. (b) Preparation of 4,4'-di (tert-butyldimethylsilyloxy) -1,1'-tellurobisbenzene (4b) A 2L three-necked flask is placed under argon and fitted with an overhead stirrer and addition funnel. Magnesium turnings (15.2 g, 0.66 mol) are added. Bromide 4a (17.2 g, 0.6 mol, used as 100%) is placed in anhydrous THF (600 mL) and transferred to the addition funnel. The Grignard reaction is initiated using approximately 25 mL of the bromide solution, iodine of some crystals. The remaining bromide solution is added at such a rate that the reaction is continuously refluxed. After the addition is complete, the reaction mixture is refluxed for an additional 30 minutes, at which time a small amount of Mg remains. After the reaction is cooled slightly in a water bath, then tellurium granules (76.8 g, 0.6 mol) are added and the reaction mixture is heated to reflux again for 2.5 hours, at which time almost all Te is consumed. The reaction is cooled to room temperature and poured into 10% aqueous NH 4 Cl (2 L) with rapid stirring and stirring for 30 minutes. The precipitated Te is filtered off through celite diatomaceous earth and the filter cake is washed with ether. The filtrate is transferred to a separatory funnel and extracted three times with ether (1200 mL, 300 mL, 300 mL). The combined extracts are washed with water and saturated NaCl, dried over MgSO 4 and filtered, and both MgSO 4 and Te precipitated during the post-treatment are removed. The solvent was removed under reduced pressure to give a crude product (194 g) as a red oil. This is added with copper powder (38 g) in toluene (900 mL) and heated to reflux for 2.5 hours in a Newman tube, where the color changes from red to gray. The reaction mixture is cooled to room temperature, filtered through celite diatomaceous earth and concentrated to give an amber oil (181 g). 1 H NMR results, it is a mixture of the desired product, quenched Grignard and residual toluene and is calculated to contain 0.23 mol of (4b). (4b). ≪ / RTI > The crude product was added dropwise to acetone (0.6 L) and iodine (58 g, 0.23 mol) with stirring and stirring. After 15 minutes, a very viscous precipitate forms. Ethanol (1.2 L) was added, and the orange solid and dark gray crystalline solid were separated by filtration and air dried. The second product is obtained by re-filtration of the filtrate. The combined yield is 206 g, which is shown by NMR to be pure, and only contaminants that are residual ethanol are present. 1 H NMR (CDCl 3) d 7.95 (4H, d, J = 8.7), 6.86 (4H, d, J = 8.7), 0.98 (18H, s), 0.25 (12H, s). 13 C NMR (CDCl3) d 158.6, 138.4, 121.9, 25.5, 18.2, -4.3. The iodine additive is placed in dioxane (500 mL), dichloromethane (500 mL) and 5% aqueous NaHSO 3 (1.1 L) and the two-phase system is vigorously stirred. After 1 hour, the lower (organic) layer is still red, indicating incomplete reduction. A large amount of a yellow aqueous layer is kept at an angle, and the organic layer is evaporated under reduced pressure. Dioxane (200 mL), dichloromethane (200 mL) and 5% aqueous NaHSO 3 (400 mL) are added to the residue and the mixture is stirred for an additional hour. This is evaporated under reduced pressure to partially remove the organic solvent and the remaining aqueous dioxane is transferred to a separatory funnel and extracted three times with ether. The combined extracts were washed with water, saturated NaHCO 3 and saturated NaCl, then dried NaSO 4, filtered above. The aqueous layer inclined from above is post-treated in the same manner and the two are combined. The solvent was removed under reduced pressure to give pure (4b) (97.3 g, 60%) as an amber oil. 1 H NMR (CDCl 3) d 7.54 (4H, d, J = 8.4), 6.80 (4H, d, J = 8.4), 0.96 (18H, s), 0.17 (12H, s). 13 C NMR (CDCl 3) d 155.8, 139.6, 121.5, 105.2, 25.7, 18.2, -4.4. FDMS 5 (m / e) 544 (M & lt ; + > , 130 Te). (c) Preparation of 4,4'-tellurobisphenol (4c) Silyl ether (4b) (54.2 g, 0.1 mol) is placed in methanol (200 mL) in a 500 mL round bottomed flask. Potassium fluoride (11.6 g, 0.2 mol) is added and the reaction mixture is heated under reflux for 1.5 hours under argon. The reaction mixture is cooled to room temperature and then poured into rapidly stirring water (1.2 L) using additional methanol (20 mL) to facilitate the transfer. The pH is adjusted to 14 using 10% NaOH (about 40 mL) and the reaction mixture is filtered through celite diatomaceous earth. The filtrate is transferred to a separatory funnel and washed twice with dichloromethane. The aqueous layer is transferred into a 2 L Erlenmeyer flask in ice and 25% aqueous acetic acid is added to pH 6 to form a cream colored precipitate. The product is separated by filtration, washed with water and air dried to give the crude product (29g). It is adsorbed onto flash silica gel (150 mL) using ether. Vacuum chromatography on 1 L of flash silica gel, eluting with dichloromethane followed by 1: 4 ethyl acetate: dichloromethane to give the pure product. Dichloro-methane gave pure (4c) (24 g, 76%) as a yellow solid as two products. 1 H NMR (CDCl 3 , 5 DMSO) d 8.77 (2H, s), 7.29 (4H, d, J = 8.4), 6.47 (4H, d, J = 8.4). (d) Preparation of 4,4'-di (ethoxycarbonylmethoxy) -1,1'-tellurobisbenzene (4d) Sodium hydride (60%, 1.76 g, 44 mmol) is placed in a 250 mL oven-dried three-necked flask equipped with an overhead stirrer and an addition funnel under argon. This was washed three times with cyclohexane, and then anhydrous (form) dimethylformamide (60 mL) was added. The flask is placed in a water bath and a solution of bisphenol (4c) (6.28 g, 20 mmol) in anhydrous dimethylformamide (15 mL) is added dropwise from the funnel (H 2 release off). Is heated for 20 minutes, the slurry obtained in an oil yoknae to 85 ℃ (additional H 2 emission). The slurry is cooled to room temperature and a solution of ethyl bromoacetate (6.68 g, 40 mmol) in anhydrous dimethylformamide (5 mL) is added dropwise from the addition funnel, during which time the precipitate becomes clear. The reaction mixture is stirred for 45 minutes and then quickly poured into stirred water (320 mL). The brown precipitate obtained is separated by filtration and washed with water. This is partitioned between water (50 mL) and dichloromethane (80 mL). Wash the aqueous phase was extracted two more times with dichloromethane, followed by the combined extracts were washed with water saturated NaCl and dried with NaSO 4 and filtered. The solvent was removed under reduced pressure, and the resulting solid was recrystallized from ethanol (50 mL), quenched and filtered. The product is obtained as a white solid (5.3 g, 55%, melting point: 88.5-89 [deg.] C). 1 H NMR (CDCl 3) d 7.60 (4H, d, J = 8.5), 6.75 (4H, d, J = 8.5), 4.58 (4H, s), 4.25 (4H, q, J = 7.1), 1.28 ( 6H, t, J = 7.1). IR (KBr) 1765, 1720, 1580, 1480 cm < -1 & gt ;. FDMS (m / e) 488 (M & lt ; + > , 130 Te). Elemental analysis for C 20 H 22 O 6 Te: Calculated: C, 49.43; H, 4.56 Found: C, 49.36; H, 4.55 (e) Preparation of 4,4'-di (carboxymethoxy) -1,1'-tellurobisbenzene, disodium salt (Te4) Diester 4d (39.2 g, 81 mmol) is placed in methanol (400 mL) and 10% aqueous NaOH (47 mL, 170 mmol) is added. A very viscous slurry is obtained. The reaction mixture is heated to reflux for 1 hour. The resulting slurry is cooled in an ice bath. The solid was isolated by filtration, washed with ethanol, air dried and triturated with mortar and pestle to give (Te4) as a cream colored solid (37.6 g, 98%). 1 H NMR (DMSO-d 6 , 5 enemy D 2 O) d 7.46 (4H , d, J = 8.4), 6.66 (4H, d, J = 8.5), 4.06 (4H, s). 13 C NMR (D 2 O) d 176.6, 158.0, 139.8, 115.9, 104.8, 66.5, IR (KBr) 3340, 3430, 1595, 1490, 1415, 1230 cm -1 . Elemental analysis for C 16 H 12 Na 2 O 6 Te 1.5 H 2 O: Calculated: C, 38.37; H, 3.02 Found: C, 38.33; H, 3.05 5. Preparation of N, N, N ', N'-tetra (carboxymethyl) -4,4'-tellurobisbenzene amine, sodium salt (Te5) (a) Preparation of N- (2-ethoxy-2-oxoethyl) -N-phenylglycine, ethyl ester (5a) Aniline (9.3 g, 0.1 mmol), ethyl bromoacetate (36.7 g, 0.22 mol) and 2,6-lutidine (23.5 g, 0.22 mol) in acetonitrile (200 mL) Reflux. The reaction is completed by TLC (dichloromethane). It is cooled to room temperature and diluted with ether (200 mL). The precipitated lutidine hydrochloride is removed by filtration, and the filtrate is evaporated under reduced pressure. The residue is partitioned between ether and water, and the aqueous layer is extracted twice more with ether. The combined extracts are washed with 1N HCl, water, saturated NaHCO 3 and saturated NaCl, dried over MgSO 4 and evaporated under reduced pressure to give crude (5a) as a dark liquid. This is taken up in dichloromethane and filtered through silica gel to remove the color to give the product as a pale oil (23.2 g, 88%). 1 H NMR (CDCl 3) d 7.21 (2H, t, J = 7.9), 6.77 (1H, t, J = 7.3), 6.61 (2H, d, J = 8.4), 4.20 (4H, q, J = 7.2 ), 4.13 (4H, s), 1.26 (6H, t, J = 7.1). (b) Preparation of N, N, N ', N'-tetra (ethoxycarbonylmethyl) -4,4'-tellurobisbenzene amine (5b) Add tellurium (IV) chloride (11.9 g, 44 mmol) to an air-dried, 500 mL three-necked flask equipped with an addition funnel and placed under argon. Anhydrous ether (200 mL) was added by syringe to give a light yellow slurry, and the flask was placed in an ice bath. A solution of (5a) (23.2 g, 89 mmol) in anhydrous ether (50 mL) is made under argon and the solution is transferred via cannula to the addition funnel. This is added dropwise to the reaction mixture with vigorous stirring. When the viscous slurry is formed immediately, the stirring bar is stopped. While the remainder of the addition is being carried out, the reaction mixture is vortexed while stirring by hand. The reaction mixture is allowed to stand overnight without stirring and then sonicated until all of the sludge is converted to a solid mass. The reaction mixture is filtered, and the filter cake is washed with ether. The filter cake was added to dichloromethane (200 mL) and a solution of sodium metasulfate (17.0 g, 89 mmol) in water (200 mL) was added with stirring. The resulting biphasic mixture is stirred at room temperature for 30 minutes. Solid NaHCO 3 is added to pH 7 and the reaction mixture is filtered through celite diatomaceous earth. The filtrate is transferred to a separatory funnel, the dichloromethane layer is separated off and the aqueous layer is extracted with additional dichloromethane. The combined extracts were washed with water and saturated NaCl, and Na 2 SO 4 dried, filtered over, and the solvents were removed under reduced pressure. The resulting red oil (17.9 g) was placed in toluene (100 mL) and copper powder (9 g) was added. The reaction mixture is heated to reflux using a Newman tube for 2 hours. The resulting gray reaction mixture is cooled to room temperature, filtered through celite diatomaceous earth and the solvent is removed under reduced pressure. The crude product (18 g amber oil) was flash chromatographed twice with dichloromethane followed by 98: 2 dichloromethane: methanol to separate the product from the recovered starting material and yield pure (5b) as a pale orange viscous oil (7.36 g, 50% of theory). 1 H NMR (CDCl 3) d 7.52 (4H, d, J = 8.6), 6.43 (4H, d, J = 8.7), 4.19 (8H, q, J = 7.1), 4.08 (8H, s), 1.25 ( 12H, t, J = 7.1). 13 C NMR (CDCl 3 ) d 170.7, 147.6, 139.5, 113.6, 101.3, 61.2, 53.3, 14.2 IR (salt plate) 1730, 1580, 1490, 1180 cm -1 . FDMS (m / e) 658 (M & lt ; + > , 130 Te). (c) Preparation of N, N, N ', N'-tetra (carboxymethyl) -4,4'-tellurobisbenzeneamine, sodium salt (Te5) The diester 5b (4.35 g, 6.63 mmol) is placed in methanol (60 mL) and 10% aqueous NaOH (7.4 mL, 26.5 mmol) is added. The reaction mixture is heated to reflux for 1 hour. The resulting slurry is cooled in an ice bath and filtered. The solid was washed with cold methanol and air dried to afford (Te5) as a pale cream colored solid (3.81 g, 91%). 1 H NMR (D 2 O) d 7.56 (4H, d, J = 8.6), 6.38 (4H, d, J = 8.6), 3.83 (8H, s). 13 C NMR (D 2 O) d 179.4, 148.7, 139.6, 113.0, 98.3, 55.5. IR (KBr) 3250 (br), 1575, 1405, 1210 cm < -1 & gt ;. Elemental analysis for C 20 H 16 Na 4 N 2 O 8 Te 3 H 2 O: Calculated: C, 35.02; H, 3.23; N, 4.08 Found: C, 34.82; H, 3.03; N, 4.04 The following examples are illustrative of the present invention and the present invention is not limited thereto. Example 1 Tear off the described coating and place it as a slide. The cartridge containing the slide is made by placing the cartridge in the lab room for 16-20 hours on top of the bench. On the first day of bench top incubation, the slides are tested using a circular automated anhydrous film immunoassay analyzer. 11 μL of human serum matrix containing 10,000 m-IU / mL hCG is applied to each slide. Each of the following slides was incubated at 37 ° C for 5 minutes, and then Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanaline (5 mM), hexadecylpyridinium chloride (0.1% 12 μL of wash solution containing H 2 O 2 (8 mM) and diethylenetriamine-pentaacetic acid (DTPA) (10 mM) is applied to each slide. All of the wash milks wash (remove) the unconjugated horseradish peroxidase labeled ligand or acceptor from the reading area and initiate the HPR-catalyzed dye formation reaction. Following the addition of the wash solution, each slide is incubated at 37 ° C for a second incubation and the reflection density readings are performed at a wavelength of 670 nm at 3 second intervals. The color forming rate for the slides from each coating is calculated from the reflection density readings. For the assay of human chorionic gonadotropin (hCG) in serum samples, anhydrous film coatings of the following formulation are prepared on a poly (ethylene terephthalate) support. The terms used in all coating structures are described in Table 1. Coating 1 layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 70.219 3,5'-dichloro-4-hydroxyacetanilide0.22 Dimedon0.45 Adhesive polymer2.58 Polymer beads130.0 BSA1.0 Glycerol2.0 Mannitol1.0 sign34e-6 Receptor layerTES buffer, pH 7.00.10 TX-1000.02 Polymer binder0.8 Leuco dye0.2 Antibody bead0.1 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 BSVME0.15 Coating 2 layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 70.219 3,5'-dichloro-4-hydroxyacetanilide0.22 Dimedon0.45 Adhesive polymer2.58 Polymer beads130 BSA1.0 Glycerol2 Mannitol1.0 sign34e-6 VOSO 4 0.04 Receptor layerTES buffer, pH 7.00.1 TX-1000.02 Polymer binder0.8 Leuco dye0.2 Antibody bead0.1 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.00.58 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BSVME0.15 Coating 3 layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 70.219 3,5'-dichloro-4-hydroxyacetanilide0.22 Dimedon0.45 Adhesive polymer2.58 Polymer beads130.0 BSA1.0 Mannitol1.0 sign34e-6 Wire0.04 Receptor layerTES buffer, pH 7.00.1 TX-1000.02 Polymer binder0.8 Leuco dye0.2 Antibody bead0.1 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.58 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BSVME0.15 Coating 4 layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 70.219 3,5'-dichloro-4-hydroxyacetanilide0.22 Dimedon0.45 Adhesive polymer2.58 Polymer beads130 BSA1.0 Glycerol2.0 Mannitol1.0 sign34e-6 Te30.012 Receptor layerTES buffer, pH 7.00.1 TX-1000.02 Polymer binder0.8 Leuco dye0.2 Antibody bead0.1 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.58 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BSVME0.15 Coating 5 layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 70.219 3,5'-dichloro-4-hydroxyacetanilide0.22 Dimedon0.45 Adhesive polymer2.58 Polymer beads130 BSA1.0 Glycerol2 Mannitol1.0 sign34e-6 VOSO 4 0.04 Wire0.04 Receptor layerTES buffer, pH 7.00.10 TX-1000.02 Polymer binder0.8 Leuco dye0.2 Antibody bead0.1 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.58 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BSVME0.15 Coating 6 layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 70.219 3,5'-dichloro-4-hydroxyacetanilide0.22 Dimedon0.45 Adhesive polymer2.58 Polymer beads130 BSA1.0 Glycerol2 Mannitol1.0 sign34e-6 VOSO 4 0.04 Te30.012 Receptor layerTES buffer, pH 7.00.1 TX-1000.02 Polymer binder0.8 Leuco dye0.2 Antibody bead0.1 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.58 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BSVME0.15 Comparison of cartridge slides for Coatings 1 to 6 jacketThe upper slide yield (n = 3)Non-top slide yield (n = 15)Top Slide Yield Loss% One0.186 Dt / Min0.231 Dt / min-19.48% 20.316 Dt / min0.317 Dt / Min-0.32% 30.305 Dt / Min0.318 Dt / min-4.09% 40.202 Dt / Min0.211 Dt / Min-4.27% 50.292 Dt / Min0.295 Dt / Min-1.02% 60.211 Dt / Min0.212 Dt / Min-0.47% The cartridge slide comparison clearly shows that a large percent loss occurs in the first slide in the cartridge when the VOSO 4 or DAT compound is not incorporated into the coating. The loss rate of hue formation in the first slide allows for an inaccurate analyte concentration for the sample being tested. Incorporation of VOSO 4 (coating 2), DAT compound (coatings 3 and 4), or combinations of VOSO 4 and DAT compounds (coatings 5 and 6) results in a significant improvement in the yield percentage retained for the top slide. Example 2 To differentiate the effects of the irradiated DAT compounds, the slides from each coating are placed directly against the top of the bench and exposed to the same conditions as the slides remaining in the cartridge (at a room temperature with fluorescent light remaining at a constant temperature of 16 to 20 hours process). After the incubation period, the slides were tested on a circular automated anhydrous-film immunoassay analyzer using the same material protocol as described above. The expected result is greater loss of color formation yield due to increased exposure of the slide to environmental factors (light, air, etc.). As a means to observe the increasing effect of environmental exposure, the color forming yield of the bench top slide is compared to the yield of color formation of the cartridge slide for each condition (except the top slide). The results for the slides processed as described are as follows: Comparison of cartridge slides for Coatings 1 to 6 jacketBench top slide yield (n = 10)Cartridge slide yield (n = 15)Yield loss of bench top slide% One0.161 Dt / Min0.231 Dt / min-30.3% 20.288 Dt / min0.317 Dt / Min-9.15% 30.293 Dt / min0.318 Dt / min-7.86% 40.179 Dt / min0.211 Dt / Min-15.17% 50.291 Dt / Min0.295 Dt / Min-1.36% 60.198 Dt / Min0.212 Dt / Min-6.6% The bench top slide test indicated that incorporation of a combination of VOSO 4 (Coating 2), DAT Compound (Coatings 3 and 4), or VOSO 4 and DAT compounds (Coatings 5 and 6) in the coating protected the loss due to environmental exposure Show me again. This test also shows that co-operation occurs when VOSO 4 and Tel are incorporated into the same coating (coating 5). This results in lower% yield loss when one compound is incorporated (Coatings 2 and 3). Dat, < RTI ID = 0.0 > Te1-Te5 < / RTI > are evaluated within a model system designed to assess loss of HRP activity. Within this system, DAT, Tel, Te4 and Te5 protect HRP activity either alone or in combination with a vanadyl salt as shown in the following examples. Examples 3 to 5 The analytical elements of the following formulation are prepared on a poly (ethylene terephthalate) support: layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer, pH 7.00.219 Adhesive polymer2.58 Polymer beads (20-40 mM)130.0 Receptor layerPolymer Binder I0.60 TES buffer, pH 7.00.10 TX-1000.02 Gelgelatin10.0 TES buffer, pH 7.04.58 TX-1000.020 BVSME0.150 The analytical element is used to measure HRP stability according to the following protocol. A 10 μl sample of a solution of 10 mM sodium phosphate buffer (pH 7.0) containing approximately 3 × 10 -8 M HRP and varying only one of the polyarylteralide compounds of the present invention was applied to a separate 1 cm < 2 > pieces (4 samples for each compound and concentration). These elements are dried and placed in a dark drawer. After 30 minutes and again after 24 hours (i.e., the next day), the components were removed from the sulcus and incubated in 1 ml of a solution of 10 mM Sodium Phosphate Buffer, 0.15 M Sodium Chloride, 0.1% Bovine Serum Albumin (pH 7.0) Extract the element by immersing it. After the test tube is vortexed for 1 minute (whereby the analytical elements are removed from the poly (ethylene terephthalate) support), the resulting suspension is centrifuged and the solution is removed. The amount of active HRP in the solution was added to a spectrophotometer cuvette and a reagent solution in an amount of 100 μL to prepare a solution containing 50 mM sodium phosphate buffer (pH 7.0), 5 mM 4'-hydroxyacetanilide, 0.625% Triton X-100 surfactant, 0.005% Is measured by adding 4,5-bis (4-dimethylaminophenyl) -2- (4-hydroxy-3-methoxyphenyl) imidazole leuco dye and 0.85 mM hydrogen peroxide. The assay temperature is 30 ° C. When blue is formed, its yield is measured spectrophotometrically at 655 nm. The color formation yield is directly proportional to the amount of active enzyme in the extract. This data is expressed as the active yield calculated by dividing the activity extracted at 24 hours with the activity extracted at 30 minutes for each sample. Measure deviations in performance data, which will be due to environmental humidity and temperature differences on the test day. This affects the apparent stability of the enzyme since it affects the drying of the HRP on the coating. Thus, the data can be compared within Examples 3 to 5, rather than in any two embodiments. Example 3 To 1mM in the efficacy of Te1, Te2, Te3, and Te4 compound alone or compared in the presence of 1mM VOSO 4 (Figure 1). Compounds Te1 and Te4 protect HRP activity in anhydrous format model compared to no addition. Te2 and Te3 are not effective stabilizers in this format. When both VOSO 4 and Te 1 or Te 4 are included, HRP activity is further maintained in the presence of only one. Example 4 The effect of concentration on the ability of Te1 and Te4 to stabilize HRP is shown (Figure 2). Use concentration to the HRP solution: 1, 0.1 or 0.01mM Te1, and 10, 1 or 0.1mM Te4 addition, spotting the HRP solutions containing buffer only, or containing 1mM VOSO 4. The activity loss is the concentration of the reagent according to both Te1 and Te4. In the presence of 10 mM Te4, the HRP activity is fully protected during the incubation for 24 hours. Example 5 This experiment shows the effect of Te4 and Te5 on the stability of the anti-CRP-HRP enzyme conjugate (2.85 x 10 -9 M). Among the Spotan's solutions, buffer alone, 1, 2, 4 or 8 mM Te4, or 1 or 8 mM Te5 is present. Both Te4 and Te5 protect the activity of conjugated HRP, and this protection is concentration dependent (Table 3). Example 6 The coatings 7 to 10 were prepared as shown in the following coating structures 7 to 10, placed as a slide, and conditioned at 70.F / 33% RH for 2 days. Coating 7 An anhydrous film coating for assay of C-reactive protein (CRP) in a serum sample of the following formulation is prepared on a poly (ethylene terephthalate) support: layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer0.219 Dimedon0.450 CaCl 2 1.0 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Glycerol2 Bovine serum albumin0.5 Magenta dye0.0538 Adhesive polymer2.538 Polymer beads130 Receptor layerTES buffer0.1 TX1000.02 Polymer binder0.8 Leuco dye0.2 PC bead0.3 Anti-CRP antibody0.05 Anti-CRP antibody-HRP conjugate0.0005 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 BSVME0.15 Coating 8 An anhydrous film coating for assay of C-reactive protein (CRP) in a serum sample of the following formulation is prepared on a poly (ethylene terephthalate) support: layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer0.219 Dimedon0.450 CaCl 2 1.0 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Glycerol2 Bovine serum albumin0.5 Magenta dye0.0538 Adhesive polymer2.538 Polymer beads130 Wire0.08 TES buffer0.1 Receptor layerTX 1000.02 Polymer binder0.8 Leuco dye0.2 PC bead0.3 Anti-CRP antibody0.05 Anti-CRP antibody-HRP antibody0.0005 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 BSVME0.15 Coating 9 An anhydrous film coating for assay of C-reactive protein (CRP) in a serum sample of the following formulation is prepared on a poly (ethylene terephthalate) support: layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer0.219 Dimedon0.450 CaCl 2 1.0 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Glycerol2 Bovine serum albumin0.5 Magenta dye0.0538 Adhesive polymer2.538 Polymer beads130 Te20.075 Receptor layerTES buffer0.1 TX 1000.02 Polymer binder0.8 Leuco dye0.2 PC bead0.3 Anti-CRP antibody0.05 Anti-CRP antibody-HRP antibody0.0005 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 BSVME0.15 Coating 10 An anhydrous film coating for assay of C-reactive protein (CRP) in a serum sample of the following formulation is prepared on a poly (ethylene terephthalate) support: layermatterFree range (g / m 2 ) Bead diffusion layerTES buffer0.219 Dimedon0.450 CaCl 2 1.0 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Glycerol2 Bovine serum albumin0.5 Magenta dye0.0538 Adhesive polymer2.538 Polymer beads130 VOSO 4 0.04 Receptor layerTES buffer0.1 TX 1000.02 Polymer binder0.8 Leuco dye0.2 PC bead0.3 Anti-CRP antibody0.05 Anti-CRP antibody-HRP antibody0.0005 TETRONIC T9080.02 OLIN 10G0.01 Gelgelatin10 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 BSVME0.15 The cartridge is thawed and loaded onto the E250 analyzer. Apply 11 μL of human serum sample containing approximately 20 mg / L CRP to each slide. Each slide was incubated at 27 ° C for 5 minutes and then incubated with Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanilide (5 mM), hexadecylpyridinium chloride (0.1%), H 2 O 2 (8 mM) and DTPA (10 [mu] M) was applied to each slide to wash (remove) the unbound antibody-HRP conjugate from the reading region and initiate the HRP-catalyzed dye formation reaction. 2.5 minutes after the reaction at 37 ° C, the reflection density at 670 nm is measured and converted to a CRP concentration through a calibration curve. The predicted concentration of the top slide is compared to the predicted average concentration of the following six slides. The results of seven experiments of this type are shown below: Bias = Top Slide Foresight - The mean value of the predicted values of the second through seventh slides jacketReactantExperiment 11 cartridgesAverage of 22 cartridges of experimentExperiment 31 cartridges 7none-29.9-27.5-34.3 8Wire-6.0-6.1-7.2 9Te2-23.2-19.3-21.8 10VOSO 4 -13.3-16.2-20.6 These results show that, in the absence of any protective agent, the top slide has substantially less predicted value than the continuous slide. Te1, Te2 or VOSO 4 the addition of the reduces the bias between the top slide and the continuous slide. The maximum increment is provided by Te1. Example 7 The coatings 7 to 10 were torn and placed on the slide, conditioned at 70.F / 33% RH for 2 days and frozen. Thaw the cartridge. The top slide of each cartridge is removed and the cartridge is incubated at 70.F / 33% RH for 3 days. This slide is tested using human serum containing approximately 20 mg / L CRP as described in Example 6. [ The predicted values of the upper slides exposed for three days at 70.F / 33% RH are compared with the average predicted values of the second to seventh slides and the results are shown below: Bias = Top Slide Foresight - The mean value of the predicted values of the second through seventh slides jacketReactantAverage of 2 cartridges 7none-26.4 8Wire-4.5 9Te2-8.2 10VOSO 4 -5.6 These results indicate that, in the absence of any protective agent, the newly exposed top slide changes over 3 days, resulting in a substantially lower predicted value than the sequential slide. Te1, Te2 or VOSO 4 protect the slide and, by resulting in a much smaller bias compared with the cartridge in a continuous slide. Tel1 again shows the maximum improvement. Example 8 The coatings 7 to 10 were torn and placed on the slide, conditioned at 70.F / 33% RH for 2 days and frozen. The cartridge is defrosted and incubated at 70.F / 33% RH for 7 days. The incubated and freshly thawed cartridges are analyzed as described in Example 6 using three human serum samples containing CRP in the range of 10-30 mg / L. Compare the predicted values of the thermostated slides with those of the freshly thawed slides. The mean biases for the three liquids of the incubated slide versus the freshly thawed slide were measured and the results are shown below: jacketReactantAverage bias 7none-7.8 8Wire0.8 9Te2-2.62 10VOSO 4 -0.26 These results show that, in the absence of a protective agent, the slides exposed for 7 days at 70. F / 33% RH produce more biases on the freshly thawed slides. In the presence of Te1 or Te2 or VOSO 4 the bias resulting from these exposure is substantially reduced. The following examples demonstrate the results that DATs coated in a gravure layer are unexpectedly better than when DAT is not used or coated on different layers. Example 9 Te5 is evaluated in the CRBM assay in the following manner. Coating 1, No Te5 An anhydrous film coating for assaying carbamazepine (CRBM) in a serum sample of the following formulation is prepared to have the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS0.0045 BSA0.00215 A-1000.00108 3 ', 5'-dichloro-4'-hydroxyacetanilide0.00995 Magenta dye0.05380 TX-1000.0043 Carboxymethyl-apo-HRP0.010 CRBM-HRP cover0.000012 Bead diffusion layerTES buffer, pH 7.00.219 3 ', 5'-dichloro-4'-hydroxyacetanilide0.220 Dimedon0.450 Adhesive polymer2.583 Polymer beads130.00 BSA1.00 Glycerol2.00 Mannitol1.00 Vanadyl sulfate dihydrate0.04 Receptor layerTES buffer, pH 7.00.10 TX-1000.02 Polymer binder0.50 Leuco dye0.20 TETRONIC T9080.02 OLIN 10G0.01 Antibody bead0.20 Gelgelatin10.00 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BVSME0.15 SupportPoly (ethylene terephthalate) Coating 2 A dry film coating for assay of carbamazepine (CRBM) containing Te5 coated in a diffusion layer at a final rewet level of 1 mM is prepared to have the following composition and layer structure: layermatterFree range (g / m 2 ) Gravure layerMOPS0.0045 BSA0.000215 A-1000.00108 3 ', 5'-dichloro-4'-hydroxyacetanilide0.00995 Magenta dye0.05380 TX-1000.0043 Carboxymethyl-apo-HRP0.010 CRBM-HRP cover0.000012 Bead diffusion layerTES buffer, pH 7.00.219 3 ', 5'-dichloro-4'-hydroxyacetanilide0.220 Dimedon0.450 Adhesive polymer2.583 Polymer beads130.00 BSA1.00 Glycerol2.00 Mannitol1.00 Vanadyl sulfate dihydrate0.04 Te50.0632 Receptor layerTES buffer, pH 7.00.10 TX-1000.02 Polymer binder0.50 Leuco dye0.20 TETRONIC T9080.02 OLIN 10G0.01 Antibody bead0.20 Gelgelatin10.00 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BVSME0.15 SupportPoly (ethylene terephthalate) Example 11 Coating 3, 1 mM Te5 (gravure layer) An anhydrous film coating for assay of carbamazepine (CRBM) containing Te5 coated in gravure layer at a final rewet level of 1 mM is prepared to have the following composition and layer structure: layermatterFree range (g / m 2 ) Gravure layerMOPS0.0045 BSA0.000215 A-1000.00108 3 ', 5'-dichloro-4'-hydroxyacetanilide0.00995 Magenta dye0.05380 TX-1000.0043 Carboxymethyl-apo-HRP0.010 CRBM-HRP cover0.000012 Te50.0632 Bead diffusion layerTES buffer, pH 7.00.219 3 ', 5'-dichloro-4'-hydroxyacetanilide0.220 Dimedon0.450 Adhesive polymer2.583 Polymer beads130.00 BSA1.00 Glycerol2.00 Mannitol1.00 Vanadyl sulfate dihydrate0.04 Receptor layerTES buffer, pH 7.00.10 TX-1000.02 Polymer binder0.50 Leuco dye0.20 TETRONIC T9080.02 OLIN 10G0.01 Antibody bead0.20 Gelgelatin10.00 TES buffer, pH 7.04.580 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.02 BVSME0.15 SupportPoly (ethylene terephthalate) Example 12 Comparative Example 1 The coatings 1 to 3 are torn and placed as a slide. The cartridge containing the slide is tested for reaction yield in two weeks after coating and placed in a freezer at -18 ° C for one year. After one year of freezer storage, the cartridge is tested for reaction yield and first bias. Slides are tested on a VITROS E250 ANALYZER. 11 [mu] l of human serum matrix solution containing 8.8 [mu] g / ml CRBM is applied to the first slide (top slide) from three cartridges and also to the second to seventh slides from the same cartridge. Each slide was incubated at 37 ° C for 5 minutes followed by addition of Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanilide (5 mM), hexadecylpyridinium chloride (0.1%), H 2 O 2 (8 mM) and DPTA (10 [mu] M) are applied to each slide. The wash solution will wash (remove) the unbound drug-horseradish peroxidase label and initiate the HRP-catalyzed dye formation reaction. Following the addition of wash solution, each slide is re-incubated at 37 ° C, during which time the reflection density is read at a wavelength of 670 nm for 2.5 minutes at 3 second intervals. Reflectance measurements are converted to transmittance, Dt, using a Clapper-William receptor layer conversion (Clapper-William transform) and color yield is calculated. The results are as follows. jacketcartridgeThe upper slide yield (n = 1)Non-top slide yield (n = 6)Loss yield of top slide% OneOne0.05199 Dt / min0.05682 Dt / min8.50 One20.05243 Dt / min0.05738 Dt / min8.63 One30.05183 Dt / min0.05687 Dt / min8.86 2One0.05500 Dt / min0.05975 Dt / min7.95 220.05406 Dt / min0.05973 Dt / min9.49 230.05470 Dt / min0.05924 Dt / min7.66 3One0.05968 Dt / min0.05927 Dt / min-0.69 320.05918 Dt / min0.05894 Dt / min-0.41 330.06020 Dt / min0.05948 Dt / min-1.21 The coating comparison clearly shows that the loss yield% on the first slide in the cartridge is large when Te5 is absent from the coating or is located in the bead diffusion layer at a re-wetting level of 1 mM. The loss of hue production in the first slide results in an inaccurate analyte concentration prediction for the tested sample. The inclusion of Te5 in the gravure layer at a re-wetting level of 1 mM significantly improves the yield percent retained for the top slide even after 1 year of freezer storage. Example 13 Te5 is evaluated in the gravure layer of the following phenobarbital anhydrous film element. Coating 4, No Te5 layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.01 Magenta dye0.16 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 PHBR-HRP cover0.00001 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N0.58 Dimedon0.50 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 Leuco dye0.20 TETRONIC T9080.02 OLIN 10G0.25 Polymer beads130 Adhesive polymer2.6 Receptor layerTES buffer0.10 Sodium hydroxide, 1N0.12 TX-1000.020 Polymer binder0.80 Antibody bead0.20 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N1.18 Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 14 Coating 5 (0.25 mM Te5 in gravure) Anhydrous film coating for assay of PHBR containing Te5 coated in gravure layer at a final rewet level of 0.25 mM is prepared to have the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.16 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 PHBR-HRP cover0.00001 Te50.016 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N0.58 Dimedon0.50 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 Leuco dye0.20 TETRONIC T9080.02 OLIN 10G0.25 Polymer beads130 Adhesive polymer2.6 Receptor layerTES buffer0.10 Sodium hydroxide, 1N0.12 TX-1000.020 Polymer binder0.80 Antibody bead0.20 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N1.18 Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 15 Coating 5 (0.25 mM Te5 in gravure) A dry film coating for the assay of PHBR containing Te5 coated in gravure layer at a final rewet level of 0.5 mM is prepared to have the following composition layer structure: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.16 Sodium hydroxide 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 PHBR-HRP cover0.00001 Te50.032 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N0.58 Dimedon0.50 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 Leuco dye0.20 TETRONIC T9080.02 OLIN 10G0.25 Polymer beads130 Adhesive polymer2.6 Receptor layerTES buffer0.10 Sodium hydroxide, 1N0.12 TX-1000.020 Polymer binder0.80 Antibody bead0.20 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N1.18 Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 16 Comparative Example 2 The fourth to sixth coatings are torn and placed as a slide. Store cartridges containing slides in a freezer at -18 ° C. After freezing for 6 months and 9 months, the cartridge is tested for reaction yield and first bias. Slides are tested on a VITROS E250 ANALYZER. 11 [mu] l of human serum matrix solution containing about 27 [mu] g / mL phenobarbital is applied to the first slide (top slide) from three cartridges and also to the second to seventh slides from the same cartridge. Each slide was incubated at 37 ° C for 5 minutes followed by addition of Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanilide (5 mM), hexadecylpyridinium chloride (0.1%), H 2 O 2 (8 mM) and DPTA (10 [mu] M) are applied to each slide. The wash solution is used to wash off (remove) the unbound drug-horseradish peroxidase label and initiate the HRP-catalyzed dye formation reaction. Following the addition of wash solution, each slide is re-incubated at 37 ° C, during which time the reflection density is read at a wavelength of 670 nm for 2.5 minutes at 3 second intervals. Reflectance measurements are converted to transmittance, Dt, using the Clapper-William transform, and the color formation yield is calculated. This is converted to a concentration of phenobarbital through a calibration curve. The results are as follows: Bias = Upper Slide Foresight - Average foresight of the second through seventh slides jacketReactantBias, 6 monthsBias, 9 months 4none3.433.54 50.25 Te52.963.55 60.50 Te51.021.15 Example 17 Te5 is evaluated in the gravure layer of a phenytoin free film element as follows Coating 7, NO Te5 A thin film coating for PHYT assay is prepared having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.054 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 PHYT-HRP cover0.000013 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N1.90 g / kg bath Dimedon0.45 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 OLIN 10G0.24 Polymer beads130 Adhesive polymer2.6 Receptor layerTES buffer0.10 Sulfuric acid, 2N1.15 g / kg bath TX-1000.020 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.010 Dimedon0.050 Polymer binder0.50 Antibody bead0.15 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N13.8 g / kg bath Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 18 Coating 8 (0.25 mM Te5 in gravure) A thin film coating for PHYT assay is prepared containing Te5 coated within a gravure layer at a final rewet level of 0.25 mM, having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.054 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 PHYT-HRP cover0.000013 Te50.016 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N1.90 g / kg bath Dimedon0.45 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 OLIN 10G0.24 Polymer beads130 Adhesive polymer2.6 Receptor layerTES buffer0.10 Sulfuric acid, 2N1.15 g / kg bath TX-1000.020 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.010 Dimedon0.050 Polymer binder0.50 Antibody bead0.15 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N13.8 g / kg bath Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 19 A thin film coating for PHYT assay is prepared containing Te5 coated within a gravure layer at a final rewet level of 0.5 mM, with the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.054 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 PHYT-HRP cover0.000013 Te50.032 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N1.90 g / kg bath Dimedon0.45 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 OLIN 10G0.24 Polymer beads130 Adhesive polymer2.6 Receptor layerTES buffer0.10 Sulfuric acid, 2N1.15 g / kg bath TX-1000.020 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.010 Dimedon0.050 Polymer binder0.50 Antibody bead0.15 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N13.8 g / kg bath Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 20 Comparative Example 3 The seventh to ninth coatings are torn and placed as a slide. Store cartridges containing slides in a freezer at -18 ° C. After freezing for 6 months and 9 months, the cartridge is tested for reaction yield and first bias. Slides are tested on a VITROS 250 ANALYZER. 11 [mu] l of human serum matrix solution containing about 12.5 [mu] g / mL phenytoin is applied to the first slide (top slide) from three cartridges and also to the second to seventh slides from the same cartridge. Each slide was incubated at 37 ° C for 5 minutes and then washed with Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanilide (5 mM), hexadecylpyridinium chloride (0.1%), H 2 12 [mu] l of wash solution containing O2 (8 mM) and DPTA (10 [mu] M) is applied to each slide. The wash liquor is used to wash (remove) the unbound drug-horseradish peroxidase label and initiate the HRP-catalyzed dye formation reaction. Following the addition of wash solution, each slide is re-incubated at 37 ° C, during which time the reflection density is read at a wavelength of 670 nm for 2.5 minutes at 3 second intervals. Reflectance measurements are converted to transmittance, Dt, using the Clapper-William transform, and the color formation yield is calculated. This is converted to the concentration of phenytoin through a calibration curve. The results are as follows: Bias = Upper Slide Foresight - Average foresight of the second through seventh slides jacketReactantBias, 6 monthsBias, 9 months 7none0.891.10 80.25 Te50.10-0.17 90.50 Te50.19-0.41 Example 21 Te5 is evaluated in the gravure layer of digoxin as follows. The coating 10 (No Te5) A dry film coating for DGXN testing is prepared having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.054 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 Digoxin-HRP label0.0000060 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N1.90 g / kg bath Dimedon0.50 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.25 Polymer beads130 Adhesive polymer2.6 Antibody bead0.055 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N13.8 g / kg bath Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 22 Coating 11 (0.25 mM Te5 in gravure) A dry film coating for DGXN assay is prepared comprising Te5 coated within a gravure layer at a final rewet level of 0.25 mM, having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.054 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 Digoxin-HRP label0.0000060 Te50.016 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N1.90 g / kg bath Dimedon0.50 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.25 Polymer beads130 Adhesive polymer2.6 Antibody bead0.055 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N13.8 g / kg bath Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 23 Coating 12 (0.5mM Te5 in gravure) A dry film coating for DGXN assay is prepared comprising Te5 coated within a Gravure layer at a final rewet level of 0.5 mM, having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.054 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.00430 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 Digoxin-HRP label0.0000060 Te50.032 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N1.90 g / kg bath Dimedon0.50 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.25 Polymer beads130 Adhesive polymer2.6 Antibody bead0.055 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N13.8 g / kg bath Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 24 Comparative Example 4 The tenth to twelfth coatings are torn and placed as a slide. Store cartridges containing slides in a freezer at -18 ° C. After freezing for 6 months and 9 months, the cartridge is tested for reaction yield and first bias. Slides are tested on a VITROS 250 ANALYZER. 11 [mu] l of human serum matrix solution containing about 2.0 [mu] g / ml digoxin is applied to the first slide (top slide) from three cartridges and also to the second to seventh slides from the same cartridge. Each slide was incubated at 37 ° C for 5 minutes and then washed with Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanilide (5 mM), hexadecylpyridinium chloride (0.1%), H 2 12 [mu] l of wash solution containing O2 (8 mM) and DPTA (10 [mu] M) is applied to each slide. The wash liquor is used to wash (remove) the unbound drug-horseradish peroxidase label and initiate the HRP-catalyzed dye formation reaction. Following the addition of wash solution, each slide is re-incubated at 37 ° C, during which time the reflection density is read at a wavelength of 670 nm for 2.5 minutes at 3 second intervals. Reflectance measurements are converted to transmittance, Dt, using the Clapper-William transform, and the color formation yield is calculated. This is converted to the concentration of phenytoin through a calibration curve. The results are as follows: Bias = Upper Slide Foresight - Average foresight of the second through seventh slides jacketReactantBias, 6 monthsBias, 9 months 10none0.160.38 110.25 Te50.0630.21 120.50 Te50.0030.03 Example 25 Te5 is evaluated in the gravure layer of carbamazepine as follows. The coating 13 (No Te5) A dry film coating for CRBM assay is prepared having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.16 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.0043 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 CRBM-HRP cover0.0000012 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N0.57 Dimedon0.45 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 OLIN 10G0.24 Polymer beads130 Adhesive polymer2.6 ReceptorTES buffer0.10 Sodium hydroxide, 1N0.40 g / kg bath TX-1000.020 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.010 Dimedon0.050 Polymer binder0.50 Antibody bead0.20 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N1.18 Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 26 Coating 14 (0.25 mM Te5 in gravure) A dry film coating for CRBM assay is prepared which comprises Te5 coated within a gravure layer at a final rewetting level of 0.25 mM, having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.16 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.0043 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 CRBM-HRP cover0.000012 Te50.016 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N0.57 Dimedon0.45 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 OLIN 10G0.24 Polymer beads130 Adhesive polymer2.6 TES buffer0.10 ReceptorSodium hydroxide, 1N0.40 g / kg bath TX-1000.020 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.010 Dimedon0.050 Polymer binder0.50 Antibody bead0.20 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N1.18 Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 27 Coating 15 (0.5mM Te5 in gravure) A dry film coating for CRBM assay is prepared comprising Te5 coated within a Gravure layer at a final rewetting level of 0.5 mM, having the composition and layer structure shown below: layermatterFree range (g / m 2 ) Gravure layerMOPS buffer0.0045 3 ', 5'-dichloro-4'-hydroxyacetanilide0.010 Magenta dye0.16 Sodium hydroxide, 1N(17 g / kg bath) BSA0.00022 A-1000.00108 TX-1000.0043 Sulfuric acid, 2N5.3 g / kg bath Carboxymethyl-apo-HRP0.010 CRBM-HRP cover0.000012 Te50.032 Bead diffusion layerTES buffer0.22 Sodium hydroxide, 10N0.57 Dimedon0.45 3 ', 5'-dichloro-4'-hydroxyacetanilide0.22 Vanadyl sulfate dihydrate0.040 Mannitol1.0 Glycerol2.0 Bovine serum albumin1.0 OLIN 10G0.24 Polymer beads130 Adhesive polymer2.6 TES buffer0.10 ReceptorSodium hydroxide, 1N0.40 g / kg bath TX-1000.020 Leuco dye0.20 TETRONIC T9080.020 OLIN 10G0.010 Dimedon0.050 Polymer binder0.50 Antibody bead0.20 Gelgelatin10 TES buffer4.6 3 ', 5'-dichloro-4'-hydroxyacetanilide0.44 TX-1000.020 Sodium hydroxide, 10N1.18 Crosslinking agent0.15 SupportPoly (ethylene terephthalate) Example 28 Comparative Example 5 The thirteenth to fifteenth coatings are torn and placed as a slide. Store cartridges containing slides in a freezer at -18 ° C. After freezing for 6 months and 9 months, the cartridge is tested for reaction yield and first bias. Slides are tested on a VITROS 250 ANALYZER. 11 [mu] l of human serum matrix solution containing about 9.3 [mu] g / ml carbamazepine is applied to the first slide (top slide) from three cartridges and also to the second to seventh slides from the same cartridge. Each slide was incubated at 37 ° C for 5 minutes and then washed with Na 2 HPO 4 (10 mM, pH 6.8), 4'-hydroxyacetanilide (5 mM), hexadecylpyridinium chloride (0.1%), H 2 12 [mu] l of wash solution containing O2 (8 mM) and DPTA (10 [mu] M) is applied to each slide. The wash liquor is used to wash (remove) the unbound drug-horseradish peroxidase label and initiate the HRP-catalyzed dye formation reaction. Following the addition of wash solution, each slide is re-incubated at 37 ° C, during which time the reflection density is read at a wavelength of 670 nm for 2.5 minutes at 3 second intervals. Reflectance measurements are converted to transmittance, Dt, using the Clapper-William transform, and the color formation yield is calculated. This is converted to the concentration of phenytoin through a calibration curve. The results are as follows: Bias = Upper Slide Foresight - Average foresight of the second through seventh slides jacketReactantBias, 6 monthsBias, 9 months 13none0.350.69 140.25 Te50.530.56 150.50 Te5-0.0130.12 Comparative Examples 2 to 5 show that the presence of Te5 in the gravure layer leads to a significant decrease in the first slide bias which is significantly closer to zero than in the absence of Te5. The level of Te5 required varies in some cases to 0.25 (PHYT) and in other cases to 0.5 (PHBR, DGXN, CRBM). However, in all cases, the first bias decreases to almost zero when Te5 is used in the gravure layer. The present invention uses a lower level of Te5 to significantly reduce the first slide bias when the stabilizer is located in the gravure layer across the diffusion layer, as opposed to being coated in the diffusion layer in competitive immuno yield assays. This has been found to be a general phenomenon according to the embodiments provided herein. Even incorporating low levels of diaryltellulose into the gravure layer of four immuno yield assays significantly reduces the first slide barriers compared to directly coating diaryltelluride in the diffusion layer. Since diaryltelluride may have a negative impact on the assay in a manner that is dose-dependent (e.g., by bleaching the recording dye), it is less effective in the gravure layer in contrast to the high capacity required in the diffusion layer The use of dithilyl telluride at a concentration is an important advantage. The present invention improves HRP stability alone or in combination with other stabilizers such as vanadyl compounds. Preferred embodiments of the present invention include both Te5 and vanadyl salts. Adhesive polymerPoly (methyl acrylate-co-sodium 2-acrylamido-2-methylpropanesulfonate-co-2-acetoacetoxyethyl methacrylate Antibody beadPolymer particles of poly (styrene-co-3- (p-vinylbenzylthio) propionic acid having an antibody bound to a ligand PC beadPolymer particles of poly (styrene-co-3- (p-vinylbenzylthio) propionic acid to which phosphoryl choline is bonded BSABovine serum albumin BVSMEBis (vinylsulfonylmethyl) ether DTPADiethylene triamine pentaacetic acid signA ligand, or a ligand and a conjugate of a horseradish peroxidase receptor Leuco dye4,5-bis (4-dimethylaminophenyl) -2- (3,5-dimethoxy-4-hydroxyphenyl) imidazole Magenta dyeDihydroxy-3- (6,8-disulfo-2-naphthyllano) -2,7-naphthalenedisulfonic acid, sodium salt MOPS3- (N-morpholino) propanesulfonic acid buffer solution OLIN 10GAn isononylphenoxy polyglycidol surfactant having an average of about 10 glycidol units per molecule Polymer beads(Vinyl toluene-co-methacrylic acid) particles having an average diameter of 20 to 40 占 퐉 Polymer binderPoly (N-isopropylacrylamide-co-2-acrylamido-2-methylpropanesulfonic acid sodium salt-co-N, N'- methylenebisacrylamide) Polymer Binder IPoly (N-isopropylacrylamide-co-2-hydroxyethyl methacrylate-co-N, N'-methylenebisacrylamide) TESN- [tris (hydroxymethyl) methyl] -2-aminoethanesulfonic acid buffer TX-100Triton X-100 surfactant-octylphenoxypolyethoxyethanol surfactant (commercially available from Union Carbide) TETRONIC T908Nonionic surfactants (commercially available from BASF Corp.) which are barrier polymers of ethylene oxide and propylene oxide, While the invention has been described in detail with particular reference to preferred embodiments thereof, it will be understood that variations and modifications can be effected within the spirit and scope of the invention. In particular, those skilled in the art will appreciate that the use of one or more diaryltellurides, or the use of certain diaryltellurides, or one or more diaryltellurides combined with a particular diaryltelluride blend or one or more vanadyl compounds, ≪ / RTI > can be selected to optimize the invention of FIG. If the stabilizer is located within the gravure layer across the diffusion layer by the present invention using Te5 or other stabilizing agent such as a vanadyl compound as opposed to being coated in the diffusion layer in competitive immuno yield assays, The first slide bias can be significantly reduced.
权利要求:
Claims (25) [1" claim-type="Currently amended] (a) a label layer comprising an enzyme-labeled ligand or an enzyme-labeled receptor; (b) a diffusion layer; (c) a receptor layer comprising a fixed receptor at a fixed concentration for the ligand and and (d) a support having a gravure layer comprising diaryltelluride. [2" claim-type="Currently amended] 2. The element of claim 1, wherein the diaryltelluride compound comprises formula (1). Formula 1 Ar 1 -Te-Ar 2 In this formula, Ar 1 and Ar 2 are the same or different aryl or heteroaryl groups selected from the group consisting of the following compounds: X is O, S, Se or Te, R 11 , R 12 , R 13 , R 21 , R 22 , R 23 , R 31 , R 32 , R 33 , R 41 , R 42 and R 43 are the same or different and each is hydrogen, , OH, OR 1 , SH, NH 2 , NHR 1 , NR 1 2 , NR 1 R 2 , CO 2 H and salts thereof, CO 2 R 1 , SO 3 H and salts thereof, PO 3 H 2 and salts thereof, And SR 1 , wherein R 1 and R 2 are different and are selected from the group consisting of alkyl, phenyl and substituted phenyl having a carbon chain having from 1 to 14 carbon atoms, ), ≪ / RTI > R 14, R 15, R 24 and R 25 are the same or different, each represents hydrogen, alkoxy having 1 to 5 carbon atoms alkyl having 1 to 5, and CO 2 H and salts thereof, CO 2 R 1, SO 3 H and Its salts, and PO 3 H 2 and its salts. [3" claim-type="Currently amended] The element according to claim 1, wherein the diaryl telluride is selected from the group consisting of the following compounds. [4" claim-type="Currently amended] The element of claim 1, further comprising a vanadyl salt. [5" claim-type="Currently amended] 3. The element of claim 2, further comprising a vanadyl salt. [6" claim-type="Currently amended] 4. The element of claim 3, further comprising a vanadyl salt. [7" claim-type="Currently amended] 3. The element of claim 1, wherein the derivative of peroxidase or peroxidase is a labeled ligand or a label in a labeled receptor. [8" claim-type="Currently amended] 8. The element of claim 7 wherein the peroxidase is horseradish peroxidase or a derivative thereof. [9" claim-type="Currently amended] 9. The element of claim 8, wherein the gravure layer further comprises a labeled ligand or labeled receptor. [10" claim-type="Currently amended] 10. The element of claim 9, wherein the gravure layer is in contact with a surface of a diffusion layer providing contact with a liquid sample. [11" claim-type="Currently amended] 11. An element according to claim 10, wherein the diaryl telluride is selected from the group consisting of the following compounds. [12" claim-type="Currently amended] 12. The element of claim 11, further comprising a vanadyl salt. [13" claim-type="Currently amended] (1) a kit comprising (a) a label layer comprising an enzyme labeled ligand or an enzyme labeled receptor, (b) a diffusion layer, (c) a receptor layer comprising a fixed receptor at a fixed concentration for the ligand and (d) providing an anhydrous immunoassay analyzing element comprising a support having a gravure layer comprising diaryltelluride; (2) a fixed ligand-receptor complex, (b) a fixed enzyme-labeled ligand-receptor complex, or (c) a mixture of (a) and b) or a fixed receptor-ligand-labeled receptor complex; (3) catalyzing the evolution of the signal by contacting the substrate solution to the defined region and the enzyme label, and (4) measuring the concentration of the ligand by detecting a signal, thereby determining an immunologically reactive ligand in the aqueous sample. [14" claim-type="Currently amended] 14. The process of claim 13, wherein the diaryltelluride compound comprises formula (1). Formula 1 Ar 1 -Te-Ar 2 In this formula, Ar 1 and Ar 2 are the same or different aryl or heteroaryl groups selected from the group consisting of the following compounds: X is O, S, Se or Te, R 11 , R 12 , R 13 , R 21 , R 22 , R 23 , R 31 , R 32 , R 33 , R 41 , R 42 and R 43 are the same or different and each is hydrogen, , OH, OR 1 , SH, NH 2 , NHR 1 , NR 1 2 , NR 1 R 2 , CO 2 H and salts thereof, CO 2 R 1 , SO 3 H and salts thereof, PO 3 H 2 and salts thereof, And SR 1 wherein R 1 and R 2 are different and each is selected from the group consisting of alkyl, phenyl and substituted phenyl having a carbon chain having from 1 to 14 carbon atoms, with or without one or several hydrophilic groups ), ≪ / RTI > R 14, R 15, R 24 and R 25 are the same or different, each represents hydrogen, alkoxy having 1 to 5 carbon atoms alkyl having 1 to 5, and CO 2 H and salts thereof, CO 2 R 1, SO 3 H and Its salts, and PO 3 H 2 and its salts. [15" claim-type="Currently amended] 14. The process according to claim 13, wherein the diaryl telluride is selected from the group consisting of the following compounds. [16" claim-type="Currently amended] 14. The method of claim 13, wherein the urea further comprises a vanadyl salt. [17" claim-type="Currently amended] 15. The method of claim 14, wherein the urea further comprises a vanadyl salt. [18" claim-type="Currently amended] 16. The method of claim 15, wherein the urea further comprises a vanadyl salt. [19" claim-type="Currently amended] 14. The method of claim 13, wherein the derivative of peroxidase or peroxidase is a labeled ligand or a label in a labeled receptor. [20" claim-type="Currently amended] 20. The method of claim 19 wherein the peroxidase is horseradish peroxidase or a derivative thereof. [21" claim-type="Currently amended] 21. The method of claim 20, wherein the signal is a colorimetric signal. [22" claim-type="Currently amended] 22. The method of claim 21, wherein the gravure layer further comprises a labeled ligand or labeled receptor. [23" claim-type="Currently amended] 23. The method of claim 22, wherein the gravure layer is in contact with a diffusion layer surface providing contact with a liquid sample. [24" claim-type="Currently amended] 24. The process according to claim 23, wherein the diaryltelluride is selected from the group consisting of the following compounds. [25" claim-type="Currently amended] 25. The method of claim 24, wherein the urea further comprises a vanadyl salt.
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同族专利:
公开号 | 公开日 EP0903583A3|2001-04-11| JPH11160316A|1999-06-18| DK0903583T3|2004-03-08| CA2247723A1|1999-03-23| AT254281T|2003-11-15| DK903583T3| PT903583E|2004-04-30| EP0903583B1|2003-11-12| AU736764B2|2001-08-02| US5928886A|1999-07-27| SI0903583T1|2004-04-30| EP0903583A2|1999-03-24| ES2210679T3|2004-07-01| DE69819646D1|2003-12-18| KR100597818B1|2006-12-01| JP4243372B2|2009-03-25| DE69819646T2|2004-12-02| AU8615498A|1999-04-15| CA2247723C|2008-07-08|
引用文献:
公开号 | 申请日 | 公开日 | 申请人 | 专利标题
法律状态:
1997-09-23|Priority to US08/935,965 1997-09-23|Priority to US08/935,965 1997-09-23|Priority to US8/935,965 1998-09-23|Application filed by 스타크 마이클, 오르토-클리니칼 디아그노스틱스, 인코포레이티드 1999-04-26|Publication of KR19990030045A 2006-12-01|Application granted 2006-12-01|Publication of KR100597818B1
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申请号 | 申请日 | 专利标题 US08/935,965|US5928886A|1995-06-07|1997-09-23|Reduction in first slide bias and improved enzyme stability by incorporation of diaryl tellurides in the gravure layer of dry-film, immunoassay elements| US08/935,965|1997-09-23| US8/935,965|1997-09-23| 相关专利
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