 NOVEL METHODS OF MANUFACTURING AN ADJUVANT
专利摘要:
The present invention relates to compositions and methods for making an adjuvant comprising a saponin using a microfluidic device and their aspects. 公开号:FR3066920A1 申请号:FR1852512 申请日:2018-03-23 公开日:2018-12-07 发明作者:Pol Harvengt；Philippe Jehoulet；Loic Le Gourrierec；Demostene Sifakakis；Laurent STRODIOT 申请人:GlaxoSmithKline Biologicals SA； IPC主号:
专利说明:
NOVEL METHODS OF MANUFACTURING AN ADJUVANT The present invention relates to methods of making an adjuvant comprising a saponin sn using a microfluidic device and related aspects. Adjuvants are included in vaccines to improve humoral and cellular immune responses, particularly in the case of weakly immunogenic subunit vaccines. Similar to natural infections with pathogens, adjuvants rely on the activation of the innate immune system to promote long-term adaptive immunity. Because the simultaneous activation of multiple innate immune pathways is a feature of natural infections, adjuvants can combine multiple immunostimulants to promote adaptive immune responses to vaccination. The adjuvant system 01 (AS01) is a liposome-based adjuvant that contains two immunostimulants, 3-O-deacyl-4'-monophosphoryl-lipid A (3D-MPL) and QS-21 (Garçon and Van Mechelen, 2011 ; Didierlaurent et al., 2017). The TLR4 agonist, 3D-MPL, is a non-toxic derivative of the lipopolysaccharide from Salmonella minnesota. QS-21 is a natural saponin molecule extracted from the bark of the South American tree Quillaja saponaria Molina (Kensil et al., 1991; Ragupathi et al., 2011). ASOl is included in the recently developed RTS, S malaria vaccine (Mosquirix ™) and Herpes zoster HZ / su vaccine (Shingrix ™) and in multiple candidate vaccine in development directed against pathogens such as the human immunodeficiency and Mycobacterium tuberculosis. During the preclinical and clinical evaluation of these candidate vaccines, immunity due to both antigen-specific antibodies and CD4 + T cells was constantly observed. The ability of ASOl to constantly generate cellular immune responses to vaccination distinguishes it from other adjuvants which generally favor mainly humoral responses to vaccination (Black et al., 2015; Garçon and Van Mechelen, 2011). Simultaneously, adjuvanted vaccines with ASOl have been effective in promoting immunogenicity to vaccination in problematic populations, such as infants (with RTS, S) and older adults (with HZ / su). The injection of ASOl produces rapid and transient activation of innate immunity in animal models. Neutrophils and monocytes are quickly recruited to the draining lymph node (GLD) during immunization. In addition, ASOl induces the recruitment and activation of dendritic cells Activation (CD) MHCII high , which are necessary for T lymphocytes (Didierlaurent AM et al., 2014) . Some data are also available on the mechanism of action of the components of ASOl. The 3D-MPL sends signals by Through TLR4, stimulant The transcriptional activity of NF-kB and cytokine production and directly activates antigen presenting cells (APC) both in humans and in mice (De Becker et al., 2000; Ismaili et al., 2002; Martin and al., 2003; Mata-Haro et al., 2007). QS-21 promotes high antigen-specific antibody responses and CD8 + T-cell responses in mice (Kensil and Kammer, 1998; Newman et al., 1992; Soltysik et al., 1995) and responses antibodies specific for the antigen in humans (Livingston et al., 1994). Because of its physical properties, it is believed that QS-21 could act as an in vivo danger signal (Lambrecht et al., 2009; Li et al., 2008). Although QS-21 has been shown to activate the ASC-NLRP3 inflammasome and the subsequent release of IL-ip / IL-18 (Marty-Roix, R. et al., 2016), the exact molecular pathways involved in the adjuvant effect of saponins must still be clearly defined. 3D-MPL and QS-21 have been shown to work synergistically in the induction of immune responses. Furthermore, it has been shown that the way in which the two immunostimulants are supplied is an important factor which influences the quality of the induced responses, with the liposomal presentation in the ASOl providing a higher power than the AS02 emulsion-based. oil in water. (Dendouga et al. 2012) Documents US 2010202928 and US 2015115488 describe the preparation of liposomes using microfluidics. WO 2013/192310 discloses methods for the mass production of nanoparticles using controlled micro-vortexes. The methods are reported to be used in the preparation of polymeric or non-polymeric particles and hybrid particles. Kim et al. Nano Letters 2012 12 (7): 3587-3591 also disclose methods for the mass production of nanoparticles using controlled micro-vortexes. Hood et al. Small 2015 11 43: 5790-5799 describe methods for the production of liposomes using microfluidics. There remains a need for new manufacturing approaches which allow the safe, practical and profitable production of liposomal adjuvants on a commercially viable scale while retaining the immunological performances emanating from conventional manufacturing approaches. It has surprisingly been discovered that a microfluidic device can be used to manufacture a liposomal adjuvant comprising a saponin while retaining immunological performance comparable to conventional manufacturing approaches. Therefore, there is provided a method of making a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. Also provided is a method of making a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) removing the solvent; and (c) adding the saponin. The present invention also provides a method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent , a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin. Also provided is a method of making a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; and (b) adding the saponin. There is further provided a liposomal concentrate for use in the preparation of a liposomal adjuvant, said liposomal concentrate comprising water, a solvent, a phosphatidylcholine lipid, a saponin and cholesterol. Therefore, there is provided a method of making a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the solution device comprising a solvent, DOPC and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. Also provided is a method of making a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) removing the solvent; and (c) adding the saponin. The present invention also provides a method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent , DOPC and a sterol, and a second solution comprising water and saponin. Also provided is a method of making a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; and (b) adding the saponin. There is further provided a liposomal concentrate for use in the preparation of a liposomal adjuvant, said liposomal concentrate comprising water, a solvent, DOPC, a saponin and cholesterol. The present invention further provides a solution comprising a solvent and 100 to 170 mg / ml lipid, in which the solvent comprises 70 to 90% v / v ethanol and 10 to 30% v / v isopropyl alcohol. Such solutions can be used in the manufacture of liposomes, as a liposomal adjuvant. Brief description of the drawings Figure 1 - Diagram of microfluidic microchip of simple mixing chamber Figure 2 - Microfluidic chip of eight mixing chambers Figure 3 - Liquid distribution tubing (one to sixteen) Figure 4 - Operational arrangement of two microfluidic chips from eight mixing chambers with distribution and collection pipes Figure 5 - Impact of the stock preparation process on the size distribution of liposomes Figure 6 - Dependence of stock stability on concentration and temperature Figure 7 - Impact of solvent composition and temperature on the size of the liposomes Figure 8 - Details of the operational conditions and organizational arrangements in example 4 Figure 9 - Visual summary of the test conditions of Example 4 Figure 10 - Results of Example 4 Figure 11 - Relationship between Zav and Pdl Figure 12 - Prediction of the size at 100 mg / ml of DOPC and ratio of 5 (organic / aqueous phases 1/4) Figure 13 - Prediction of the size at 130 mg / ml of DOPC and ratio of 5 (organic / aqueous phases 1/4) Figure 14 - Prediction of the size at 160 mg / ml of DOPC and ratio of 5 (organic / ageuse phase 1/4) Figure 15 - Prediction of the size at 100 mg / ml of DOPC and ratio of 4 (organic / ageuse phases 1/3) Figure 16 - Prediction of the size at 100 mg / ml of DOPC and ratio of 6 (organic / aqueous phases 1/5) Figure 17 - Prediction of the size at 130 mg / ml of DOPC and ratio of 6 (organic / aqueous phases 1/5) Figure 18 - Prediction of the size at 130 mg / ml of DOPC and ratio of 4 (organic / ageuse phases 1/3) Figure 19 - Prediction of the size at 160 mg / ml of DOPC and ratio of 4 (organic / aqueous phases 1/3) Figure 20 - Prediction of the size at 160 mg / ml of DOPC and ratio of 6 (organic / aqueous phases 1/5) Figure 21 - Confirmation conditions tested at 130 mg / ml of DOPC and a ratio of 5 (organic / aqueous phases 1/4) Figure 22 - Title of specific IgE for gE Figure 23 - Percentage of gE-specific CD4 + T cells Figure 24 - Size of liposomes produced by microfluidics and stability of Pdl after storage Figure 25 - Diagram of a multi-chamber process on a commercial scale Figure 26 - Diagram of an initial tubing design Figure 27 - Photograph of the assembly to test the initial tubing ίο Figure 28 - Variation in flow rate observed for the initial tubing at 200 ml / min Figure 29 - Variation in flow rate observed for the initial tubing at 50 ml / min Figure 30 - Diagram comparing the predicted and measured values of the flow distribution Figure 31 - Diagram of the improved tubing design Figure 32 - Flow variation from the average by each channel for the improved tubing unit 1 ("B1") Figure 33 - Flow variation from the average by each channel for the improved tubing unit 2 (“B2”) Figure 34 - Variation in flow rate from the average for each channel for the improved tubing unit 3 (“A2”) Figure 35 - Comparison of the size distribution between the liposomes of the 16 mixing chambers and the single mixing chamber Figure 36 - Comparison of the specific IgG titers of gE between the liposomal adjuvants produced by microfluidics and thin film Figure 37 - Comparison of g4-specific CD4 + T cells between liposomal adjuvants produced by microfluidigue and thin film Figure 38 - Comparison of RMG of IgG-specific IgG between liposomal adjuvants produced by microfluidics and thin films Figure 39 - Comparison of RMG of gE-specific CD4 + T cells between liposomal adjuvants produced by microfluidics and thin film Figure 40 - Comparison of the size distribution between the liposomes with a saponin (QS21) and a TLR4 agonist (3D-MPL) from the 16 mixing chambers and the simple mixing chamber Description of sequence identifiers SEQ ID NO: 1 - Sequence of the RTS polypeptide SEQ ID NO: 2 - Sequence of the Rv1196 polypeptide of the M. tuberculosis strain H37Rv SEQ ID NO: 3 Sequence of the Rv0125 polypeptide of the M. tuberculosis strain H37Rv SEQ ID NO: 4 - Sequence of polypeptide of fusion M72 SEQ ID NO: 5 - Sequence of polypeptide of fusion M72-2his SEQ ID NO: 6 - Sequence polypeptide of gE truncated of virus ; chickenpox shingles SEQ ID NO: 7 - Sequencepolypeptide of conformally constrained RSV PreF antigen SEQ ID NO: 8 - Sequence polypeptide of the HIV TV1 gpl20 SEQ ID NO: 9 - Sequence polypeptide of the HIV 1086C gpl20 detailed description The present invention provides a method for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. Also provided is a method of making a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) removing the solvent; and (c) adding the saponin. The present invention also provides a method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent , a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin. Also provided is a method of making a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; and (b) adding the saponin. There is further provided a liposomal concentrate for use in the preparation of a liposomal adjuvant, said liposomal concentrate comprising water, a solvent, a phosphatidylcholine lipid, a saponin and cholesterol. The present invention also provides a method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. Also provided is a method of making a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water r (b) removing the solvent; and (c) adding the saponin. The present invention also provides a method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water and saponin. A method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin is also provided, using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; and (b) adding the saponin. There is further provided a liposomal concentrate for use in the preparation of a liposomal adjuvant, said liposomal concentrate comprising water, a solvent, DOPC, a saponin and cholesterol. The present invention further provides a solution comprising a solvent and 100 to 170 mg / ml lipid, in which the solvent comprises 70 to 90% v / v ethanol and 10 to 30% v / v isopropyl alcohol. Such solutions can be used in the manufacture of liposomes, as a liposomal adjuvant. Microfluidic devices A microfluidic device is a device handling liquids in which generally at least one aspect has a dimension on the sub-mm scale and generally the mixing occurs using passive means (i.e., using liquid stream contact and without moving parts within the mixing chamber). The microfluidic device will include a mixing chamber in which the first solution and the second solution are mixed. The mixing chamber will generally have a cross section which is 25.6 mm 2 or less, such as 12.8 mm 2 or less, suitably 6.4 mm 2 or less, especially 3.2 mm 2 or less and particular 1.6 mm 2 or less. The mixing chamber will generally have a cross section which is 0.1 mm 2 or more, suitably 0.2 mm 2 or more, especially 0.3 mm 2 or more and in particular 0.4 mm 2 or more. In some embodiments, the mixing chamber will have a cross section which is from 0.2 to 3.2 mm 2 , such as 0.4 to 1.6 mm 2 , especially 0.6 to 1.2 mm 2 and in particular 0.7 to 1.0 mm 2 (for example, 0.8 mm 2 ). The cross section of the mixing chamber can be of any shape, although it is generally symmetrical. The cross section may be substantially rectangular (as square). The cut can be elongated in kind, with the largest dimension being at least twice that of the perpendicular dimension, such as at least three times or at least four times. The larger dimension can be no more than ten times that of the perpendicular dimension, such as no more than eight times or no more than six times. The larger dimension will usually be two to ten times that of the perpendicular dimension, such as three to eight times, especially four to six times, especially five times. A rectangular cross section can have a side length of 1 to 8 mm, like 1 to 4 mm, for example 1.4 to 3.2 mm, especially 1.6 to 2.4 mm, in particular 1.8 to 2, 2 mm (for example, 2 mm). A rectangular cross section can have a short side of 0.1 to 4 mm, for example, 0.1 to 2 mm, possibly 0.1 to 1.2 mm, like 0.1 to 0.8 mm, especially 0, 2 to 0.6 mm, in particular 0.3 to 0.5 mm (for example, 0.4 mm). The microfluidic device will have at least one inlet (such as an inlet) into the mixing chamber for the administration of the first solution. The device can have a plurality of inlets in the mixing chamber for the administration of the first solution, such as two inlets. Suitably, the microfluidic device will have five or fewer inlets into the mixing chamber for administration of the first solution, such as four or less. The microfluidic device will have at least one inlet into the mixing chamber for the administration of the second solution. The device may have a plurality of inlets in the mixing chamber for the administration of the second solution, such as two inlets. Suitably, the microfluidic device will have five or fewer inlets into the mixing chamber for administration of the second solution, such as four or less. To facilitate adequate mixing, the number of inlets for the first solution and the second solution can be increased for mixing chambers with larger cross sections. The cross section of the entrances can be of any shape, although it is generally symmetrical. The cross section can be rectangular (like square). Each Entrance will have usually cutting transversal who is of 1.28 mm 2 or less, of appropriately 0.64 2 mm or lower, specially 0.32 mm 2 or lower and in particular 0.16 mm 2 or lower. Each entry will generally have a cross section which is 0.01 mm 2 or greater, suitably 0.02 mm 2 or greater, especially 0.03 mm 2 or greater and in particular 0.04 mm 2 or greater. In some embodiments, each entry will have a cross section which is from 0.02 to 0.32 mm 2 , especially 0.06 to 0.12 mm 2 and in particular 0.07 to 0.10 mm 2 (for example , 0.8 mm 2 ). The total cross section of all inlets will generally be less than 70% of the cross section of the mixing chamber, as less than 60% and especially less than 50%. Conveniently, the inlets can cover the full length of one side of the mixing chamber. The shape and size of each entry can vary independently. However, generally the entries for the first solution will be identical in shape and in size, and the entries for the second solution will be identical in shape and in size. In practice, all entries are identical in shape and size. Each entry can be 2 to 20% of the width of the mixing chamber, for example, 5 to 15% as 8 to 12%, especially 10%. A particular inlet design is rectangular, 0.2 mm wide, and covers the full length of the other side of the mixing chamber (for example, 0.4 mm high). The inputs will generally be located in such a way that the direction of flow of the first solution and the second solution in the mixing chamber is substantially parallel (for example, in degrees, such as 10 degrees, in particular 5 degrees), as parallel to the general direction of flow through the mixing chamber. The microfluidic device will include at least one outlet from the mixing chamber for the recovery of the mixed material. The device may include a plurality of outlets from the mixing chamber for recovering the mixed material, such as two or three outlets, which are subsequently combined. Suitably, the device will have a single outlet from the mixing chamber for the recovery of the mixed material. The cross section of the outlets can be of any shape, although it is generally symmetrical. The cross section can be rectangular (as square), generally having an area of 0.2 to 1 mm 2 , such as 0.3 to 0.6 mm 2 , for example 0.4 to 0.5 mm 2 . In other examples, the outlet may be of circular cross section (for example, having a diameter of 0.5 to 1 mm, such as 0.6 to 0.8 mm, for example 0.75 mm) The total cross section of all outlets will generally be less than 70% of the cross section of the mixing chamber, as less than 60% and especially less than 50%. The mixing chamber must have an adequate length to allow the mixture to be substantially complete by the time the liquid reaches the outlet (s). Generally, the chamber will have a length of 1 to 10 cm, like 1.5 to 5 cm, especially 1.8 to 4 cm, in particular 2 to 3 cm, for example 2.5 cm. In one embodiment, the device comprises a mixing chamber which has a rectangular transverse cross section, having from 0.2 to 3.2 mm 2 (for example, 0.6 to a long side section of 1.4 to 3, 2 mm (for example, 1.6 to 2.4 mm), one example side, 0.32 solution and two are arranged proximal to the chamber and one chamber has a short one entries for 0.1 to 1.2 entries for the second one symmetrically with mixing chamber, one mm (by first solution which The end of mixing length 1.5 outlet located at 5 cm (for example, 2 to 3 at the distal end of mixing. Width 0.16 to 0.24 length on the other side of cm) the appropriate, mm entries and cover the full mixing chamber. The microfluidic device can be formed from any suitable material, namely one which is tolerant to the components used in the first solution and the second solution and which lends itself to suitable materials include glass. The appropriate stainless steel. The devices from these materials by devices in plasma ion etching) and one is one can etching, silicon can reactive deep devices in manufacturing. The silicon and the other material to be prepared for example, to be them (DRIE, glass prepared by wet etching (HF etching). prepared by or etching can be The selected materials can be subjected to a surface treatment to improve the characteristics of the surface. To get a batch processing time that is a manageable period of time (for example, 240 minutes or less, especially 120 minutes or less), it is necessary that the system reaches a sufficient level of productivity. In addition, to help homogeneity from one batch to another by reducing the impact of the effects necessary as the starting and stopping, there is processing time adequate length (by specially at least 60 is one example, at least minutes, minutes). _____ increase of ____ scale of the microfluidic device In order to facilitate the production of the liposomal adjuvant on an industrial scale (for example, a scale of at least 0.5 g of phosphatidylcholine lipid per minute, such as at least 1 g per minute, in particular at least 2 g per minute and especially at least 4 g per minute, as a scale of at least 0.5 g of DOPC per minute, as at least 1 g per minute, in particular at least 2 g per minute and especially at least 4 g per minute ), large mixing chambers can be used or a plurality of mixing chambers can operate in parallel. For example, 2 or more mixing chambers, especially 4 or more, especially 8 or more, such as 16 or more (for example, 16). The plurality of mixing chambers which operate in parallel can be 128 or less, such as 64 or less, especially 32 or less. Consequently, in certain embodiments, the plurality of mixing chambers is from 2 to 128, such as 4 to 64, for example 8 to 32. Under certain circumstances, each mixing chamber from the plurality of mixing chambers can operate independently, with the supply of the first solution and the second solution to the mixing chamber by independent pumps (i.e., each pump not simultaneously supplying the solution to any other mixing chamber). The first solution and / or the second solution can be stored in independent containers (i.e. containers not simultaneously supplying the first solution and / or the second solution to more than one mixing chamber), or the first solution and / or the second solution can be stored in a container to be used in more than one mixing chamber (like all mixing chambers). The mixed material from each mixing chamber can be collected individually and stored / processed, possibly being combined at a later stage, or it can be combined (e.g. from all mixing chambers) before further processing and / or storage. In practice, all of the mixing chambers in the plurality of mixing chambers are fed by the same pumps and the mixed material from all of the mixing chambers is collected before further processing and / or storage. Suitably, all of the mixing chambers and the flow of liquid within all of the mixing chambers are substantially identical, such that the material obtained from each mixing chamber is substantially identical. Desirably, the flow rates measured in each mixing chamber vary by less than 5% of the desired flow rate. Optimally, the mixing chambers, the inputs and outputs, the supply of the first solution, of the second solution and the collection of the mixed material from the multiple mixing chambers are configured in such a way that, during operation, their performances are substantially identical. Each mixing chamber from the plurality of mixing chambers can be configured as an individual chip or, for practical reasons, a number of mixing chambers can be combined into a single chip (for example, containing 8 mixing chambers). A number of these chips can be used in parallel to provide the plurality of chambers (for example, two chips each containing 8 mixing chambers to provide a total of 16 mixing chambers which operate in parallel). Suitably, the plurality of mixing chambers is capable of producing the mixed material at a total rate of 50 to 2000 ml / min, such as 100 to 1000 ml / min, in particular 200 to 500 ml / min. The microfluidic devices described herein are an aspect of the present invention. First solution The first solution (the "organic" phase) comprises a solvent, a phosphatidylcholine lipid and a sterol. Suitably, the first solution comprises a solvent, DOPC and a sterol. The solvent should dissolve the phosphatidylcholine lipid (such as DOPC), the sterol and any other component present to provide the first solution in the form of a single phase. In addition, the solvent should be miscible with the aqueous solution, so that the mixture of the first solution and the second solution produces a simple liquid phase which includes a suspension of liposomes. The solvent will be an organic solvent or a simple phase mixture comprising at least one organic solvent. The solvent can include a short chain organic alcohol, such as ethanol and / or isopropanol. Suitably, the solvent will include ethanol, such as at a concentration between 70 and% v / v, more suitably between 75 and 85% v / v, or between 78 and 82% v / v. Suitably, the solvent will include isopropanol, such as at a concentration between 20 and 30% v / v, more suitably between 15 and 25% v / v, or between 18 and 22% v / v. Suitably, the solvent will consist essentially of ethanol at a concentration between 70 and 90% v / v and isopropanol at a concentration between 10 and 30% v / v, such as ethanol at a concentration between 75 and 85% v / v and isopropanol at a concentration between 15 and 25% v / v, especially ethanol at a concentration between 78 and 82% v / v and isopropanol at a concentration between 18 and% v / v, in particular ethanol at a concentration of 80% v / v and isopropanol at a concentration of 20% v / v. At higher ethanol concentrations, such as greater than 90% v / v ethanol, the capacity for dissolving the solvent is limited (which ultimately constrains the capacity of the system). At lower ethanol concentrations, such as less than 70% v / v ethanol, the process may be more sensitive to operational parameters, such as temperature. As mentioned, the first solution will include a phosphatidylcholine lipid. The phosphatidylcholine lipid will contain unbranched acyl chains having 12 to 20 carbon atoms, optionally with a double bond, being of particular interest those with acyl chains having 14 to 18 carbon atoms, possibly with a double bond. Generally, each of the two identical strings. acyl in a lipid molecule is The particular phosphatidylcholine lipids of i phosphatidylcholine phosphatidylcholine phosphatidylcholine phosphatidylcholine phosphatidylcholine phosphatidylcholine phosphatidylcholine and understand : the lipids of saturated -dilauroyl- (DLPC),the dimyristoyl (DMPC),the dipalmitoyl (DPPC),the distearoyl (DSPC) and the diarachidoyl- (DAPC); and lipids from unsaturated - the dipalmitoléoyl- and dioleoyl-phosphatidylcholine phosphatidylcholine (DOPC); and their mixtures. Suitably, the lipid of other phosphatidylcholine is substantially purified lipid. Generally the phosphatidylcholine lipid is at least 80% pure, as at least 90% pure, especially at least 95% pure, in particular 98% pure, for example at least 99% pure or even at least 99, 8%. Consequently, the invention provides a solution comprising a solvent and 100 to 170 mg / ml of lipid, in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v of isopropyl alcohol. Suitably the lipid is a phosphatidylcholine lipid, therefore suitably the solution provided is the first solution. As mentioned above, the first solution suitably comprises DOPC (dioleoylphosphatidylcholine). Suitably, DOPC is substantially purified from other lipids, both other types of acyl chain and other types of head group. Generally, the DOPC is at least 90% pure, as at least 95% pure, especially at least 98% pure, in particular 99% pure, for example at least 99.8% pure. Suitably, the first solution comprises 100 to 170 mg / ml of DOPC, like 100 to 160 mg / ml of DOPC, especially 120 to 160 mg / ml. The first solution may include 120 to 150 mg / ml of DOPC, such as 120 to 140 mg / ml of DOPC. In particular, the first solution can comprise around 130 mg / ml of DOPC (for example, 125 to 135 mg / ml of DOPC, especially 130 mg / ml of DOPC). The sterol will usually be cholesterol. Cholesterol is disclosed in the Merck Index, 13th edition, page 381, as an existing naturally occurring sterol found in animal fat. Cholesterol has the formula (C27H46O) and is also known as (3β) -cholest-5-en-3-ol. Suitably, the first solution comprises 20 to 50 mg / ml of sterol (for example, cholesterol), like 25 to 40 mg / ml, especially around 32.5 mg / ml (for example, 30 to 35 mg / ml, in particular 32.5 mg / ml) Suitably, the dry weight of the first solution is 100 to 250 mg / ml, like 140 to 220 mg / ml, especially 150 to 220 mg / ml. Consequently, the invention provides a solution comprising a solvent and 100 to 170 mg / ml of lipid, in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v of isopropyl alcohol. Suitably the lipid is DOPC, therefore suitably the solution provided is the first solution. The lipids to be used in the present invention will generally be membrane forming lipids. Membrane forming lipids include a diverse range of structures including phospholipids (e.g., phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol and phosphatidylserine), ceramides and sphingomyelins. The lipids forming a membrane will generally have a polar head group (which in a membrane aligns with respect to the aqueous phase) and one or more (two) hydrophobic tail groups (which in a membrane combine to form a nucleus hydrophobic). Hydrophobic tail groups will generally be in the form of acyl esters, which can vary both in length (for example, from 8 to 26 carbon atoms) and in degree of unsaturation (for example, one, two or three double bonds). The lipids to be used in the present invention may be of natural or synthetic origin, and they may be a simple pure component (e.g., 90% pure, specially 95% pure and suitably 99% pure on a base by weight), a simple class of lipid components (for example, a mixture of phosphatidylcholines, or alternatively, a mixture of lipids with a conserved type of acyl chain) or they can be a mixture of many different types of lipids. In one embodiment of the invention, the lipid is a simple pure component. Pure lipids are generally of synthetic or semi-synthetic origin. Examples of pure lipids for use in the present invention include phosphatidylcholines (e.g. DLPC, DMPC, DPPC, DSPC and DOPC; in particular DLPC, DMPC, DPPC and DOPC; especially DOPC) and phosphatidylglycerols (e.g. DPPG) , suitably phosphatidylcholines. The use of pure lipids is desirable due to their defined composition, however, they are generally more expensive. In one embodiment of the invention, the lipid is a mixture of components. The lipid mixtures to be used in the present invention can be of natural origin, obtained by extraction and purification by means known to those skilled in the art. Mixtures of naturally occurring lipids are generally significantly less expensive than pure synthetic lipids. Naturally derived lipids include lipid extracts from egg or soy, which extracts will generally contain lipids with a mixture of acyl chain lengths, degrees of unsaturation and types of lead group. Lipid extracts of plant origin can be expected to demonstrate higher rates of unsaturation compared to those of animal origin. It should be noted that, due to the variation in the source, the composition of the lipid extracts may vary from batch to batch. In one embodiment of the invention, the lipid is a lipid extract containing at least 50%, especially at least 75% and suitably at least 90% by weight of phospholipids from a single type of head group (for example example, phosphatidylcholines). In a second embodiment of the invention, particular lipid extracts may be preferred because of their relatively low cost. In a third embodiment of the invention, the lipid is a mixture of lipids having a conserved acyl chain length (for example, at least 50%, especially at least 75% and suitably at least 90% by weight ), for example 12 (for example, lauryle), 14 (for example, myristyle), 16 (for example, palmityl) or 18 (for example, stearyl or oleoyl) carbon atoms in length. Suitably, a lipid extract to be used in the present invention will comprise at least% of phospholipids by weight (for example, phosphatidylcholines and phosphatidylethanolamines), especially at least 55% of phospholipids by weight, in particular at least 60% of phospholipids by weight (like 75% or 90%). Lipid mixtures can also be prepared by combining pure lipids, or by combining a lipid extract with either other lipid extracts or with pure lipids. The ratio of lipid (for example, DOPC) to sterol is usually 3/1 to 5/1 w / w, like 3.5 / 1 to 4.5 / 1 w / w. In certain embodiments, the first solution consists essentially of a solvent and from 100 to 160 mg / ml of lipid and from 30 to 40 mg / ml of cholesterol in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v isopropyl alcohol, desirably, the lipid is a phosphatidylcholine. Suitably, the lipid is DOPC. In order to prepare liposomal adjuvants comprising a TLR4 agonist, the TLR4 agonist may optionally be included in the first solution. The first solution may contain 1 to 25 mg / ml of the TLR4 agonist, such as 2 to 16 mg / ml, especially 3 to 12 mg / ml and in particular 4 to 10 mg / ml (for example, around 6, 5, such as 5.5 to 7.5 mg / ml, especially 6.5 mg / ml). The present invention also provides a process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with the phosphatidylcholine lipid and cholesterol; (iii) adding other solvents; (iv) the mixture. The present invention also provides a process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with DOPC and cholesterol; (iii) adding other solvents; (iv) the mixture. Suitably, mixing is carried out at a temperature of 30 to 50 ° C, especially 35 to 45, such as 40 ° C. Suitably, the at least part of the solvent is at least 25% of the solvent, especially at least 35% and in particular at least 45%. Suitably, the at least part is 90% of the solvent or less, such as 80% or less, especially 70% or less and in particular 60% or less. In some examples, the at least part is 35 to 70% of the solvent, such as 45 to 60%. Suitably, the other solvent is any remaining solvent, although it may be a part of the remaining solvent with additional solvent added later. Consequently, the present invention also provides a method for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said method comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with the phosphatidylcholine lipid and cholesterol; (iii) adding other solvents; (iv) the mixture; (v) adding additional solvent. Suitably, the other solvent is any remaining solvent, although it may be a part of the remaining solvent with additional solvent added later. Consequently, the present invention also provides a method for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said method comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with DOPC and cholesterol; (iii) adding other solvents; (iv) the mixture; (v) l 4 addition of additional solvent. The additional solvent can be from 0 to 30% of the solvent, such as from 0 to 20%. '33 The solution resulting from any of the above methods can then be filtered to remove any particulate material before use in the microfluidic apparatus. Other characteristics of the process can be as described for the first solution, for example, the solution comprises 100 to 160 mg / ml of lipid and 30 to 40 mg / ml of cholesterol and in which the solvent comprises 70 to 90% v / v ethanol and 10 to 30% v / v isopropyl alcohol. Desirably, the lipid is a phosphatidylcholine. Suitably, the lipid is DOPC. Suitably, the solution comprises 4 to 10 mg / ml of TLR4 agonist, in particular 3D-MPL. Suitably, the invention provides a solution consisting essentially of 100 to 160 mg / ml of lipid and 30 to 40 mg / 1 of cholesterol and in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v isopropyl alcohol. Desirably, the lipid is a phosphatidylcholine, more suitably the lipid is DOPC. Suitably, the solution comprises 4 to 10 mg / ml of TLR4 agonist, in particular 3D-MPL. Second solution The second solution (the "aqueous" phase) includes water and in some processes, it can include a saponin. The second solution acts as a counter solvent, causing the formation of liposomes when mixed with the first solution. The faster the precipitation of the components from the first solution, generally the smaller the liposomes obtained. The second solution can be substantially aqueous and it will comprise at least 90% v / v of water, like at least 95% of water, especially at least 98% of water and in particular 100% of water. When present in the second solution, suitably, the saponin is present at a concentration of 0.05 to 25 mg / ml, such as 0.2 to 10 mg / ml, especially 0.5 to 5 mg / ml and in particular 0.8 to 3 mg / ml (for example, about 1.625 mg / ml, such as 1.2 to 2 mg / ml, especially 1.625 mg / ml). When the saponin is not present in the second solution, suitably, the second solution consists essentially (as is) of water. When the saponin is present in the second solution, suitably, the second solution consists essentially of (as is constituted) of water and of the saponin, for example the second solution can be saponin (such as QS-21) in water for injection. The ionic strength of the second solution will suitably be 150 nM or less, such as 100 nM or less, particularly 80 nM or less, especially 60 nM or less, for example 40 nM or less. Conductivity can be a practical substitute for the ionic strength of an aqueous solution. The conductivity of the second solution will suitably be 12 mS / cm or lower, for example mS / cm or lower, 8 mS / cm or lower, 6 mS / cm or lower, or 4 mS / cm or lower. Suitably, the second solution consists essentially of aqueous saponin. Microfluidic operation Optimal operating conditions will depend on the precise configuration of the device and the desired product characteristics. Suitably, the total flow rate in the mixing chamber is 15 to 30 ml / min / mm 2 of cross section of the mixing chamber, like 16 to 28 ml / min / mm 2 , especially 17.5 to 25 ml / min / mm 2 and in particular 19 to 21 (for example, 20 ml / min / mm 2 ). Suitably, the flow ratio for the first and second solutions will be in the range of 1/2 to 1/6, like 1/3 to 1/5, especially 1 / 3.5 to 1 / 4.5 and especially 1/4. High levels of solvent in the mixed material can have an impact on the stability of the liposomes so that the flow rate ratio which will produce high concentrations of solvent is desirably avoided - solvent concentrations of 50% result from a ratio 1/1, 33% for 1/2, 25% for a ratio of 1/3, 20% for a ratio of 1/4 and 16.6% for a ratio of 1/5. Low throughput from the first solution reduces system productivity. Flow reports that produce relatively large volumes of mixed material are less desirable because of the safety protocols associated with the handling and use of compositions containing solvents that exceed certain thresholds (eg, 50 L). Suitably, the flow rate of the first solution into the mixing chamber is in the range of 2 to 7.5 ml / min / mm 2 of cross section of the mixing chamber, such as 2.5 to 7 ml / min / mm 2 , especially 3 to 6.5 ml / min / mm 2 and in particular 3.5 to 6 (for example, 5) ml / min / mm 2 . Suitably, the flow rate of the second solution into the mixing chamber is in the range of 11 to 25 ml / min / mm 2 of cross section of the mixing chamber, such as 12 to 20 ml / min / mm 2 , especially 14 to 18 ml / min / mm 2 and in particular 15 to 17 (for example, 16) ml / min / mm 2 . The first solution and the second solution will generally be provided at a temperature in the region of 10 to 30 ° C, such as 15 to 25 ° C, in particular 18 to 22 ° C, especially 20 ° C, and they can be at temperatures identical or different, suitably at the same temperature and especially at 20 ° C. The mixing chamber can be maintained at a temperature in the region of 10-30 ° C, such as 15-25 ° C, particularly 18-22 ° C, especially 20 ° C. Depending on the design of the device and the environmental conditions, it may only be necessary to actively control the temperature of the first solution and the second solution, and not to actively control the temperature of the mixing chamber. The mixture of the first solution and the second solution may be slightly exothermic. Lower operating temperatures result in the formation of smaller liposomes. The microfluidic device can operate within a temperature-controlled environment, for example, where the temperature is maintained in the range of 10 to 30 ° C, such as 15 to 25 ° C, in particular about 20 ° C (such as 18 to 22 ° C, especially 20 ° C). The operating pressure of the system does not need to be controlled. Suitably, the maximum Reynolds number within the mixing chamber is 2100, in particular 1800, like 1500, especially 1000, for example 500. The maximum Reynolds number within the mixing chamber is so suitable within the range of 25 to 1500, more appropriately between 50 and 500, especially 75 to 300 and especially 100 to 200. The methods for calculating the Reynolds number are known to those skilled in the art and are illustrated in the examples here. liposomes When the first solution and the second solution are mixed, liposomes will form. The term "liposome" is well known in the art and defines a general category of vesicles which include one or more lipid bilayers surrounding an aqueous space. Liposomes thus consist of one or more lipid and / or phospholipid bilayers and can contain other molecules, such as proteins or carbohydrates, in their structure. Because two lipid and aqueous phases are present, liposomes can encapsulate or trap water-soluble material, lipid-soluble material, and / or amphiphilic compounds. The size of the liposomes can vary from 30 nm to several pm depending on the phospholipid composition and the process used for their preparation. The liposomes of the present invention contain a phosphatidylcholine lipid. or, consist essentially of a phosphatidylcholine lipid and sterol (with a saponin and a TLR4 agonist if necessary). Suitably, the liposomes of the present invention contain DOPC, or, consist essentially of sterol DOPC (with a saponin and a TLR4 agonist where appropriate). In the present invention, the size of the liposomes will be in the range of 50 nm to 200 nm, especially 60 nm to 180 nm, such as 70 to 165 nm. Optimally, the liposomes should be stable and have a diameter of ~ 100 nm to allow sterilization by practical filtration. The structural integrity of the liposomes can be estimated by methods such as dynamic light scattering (DLS) measuring the size (mean diameter, Zav) and the polydispersity of the liposomes, or by electron microscopy for Analysis of the structure of liposomes. Suitably, the average particle size is between 95 and (Pdl) is not 120 nm, and / or, the polydispersity index not more than 0.35, in particular not more than 0.3, as not more than 0.25. In one embodiment, the average particle size is between 95 and 120 nm, and / or, the polydispersity index (Pdl) is not more than 0.2. In certain circumstances, the presence of solvents and certain additional components can have an impact on the size of the liposomes. Therefore, the size of the liposomes is appropriately measured after removal of the solvent and incorporation of all additional components. Solvent removal The mixed material recovered will include liposomes in water and solvent. One such material is a liposomal concentrate for use in the preparation of a liposomal adjuvant, said liposomal concentrate comprising water, a solvent, a phosphatidylcholine lipid, a saponin and cholesterol, such as comprising water, a solvent , DOPC, a saponin and cholesterol. The recovered material can be stored for later use or it can be further processed to remove some or all of the solvent. To facilitate the use of liposomes in an adjuvant, it is desirable to remove substantially all of the organic solvent (for example, leaving at least 98% w / w of water, such as at least 99% of water, especially at least 99.5% water, in particular at least 99.9% water, such as at least 99.99%). Suitably, the residual organic solvent is at a level which is equivalent to less than 150 pg per human dose, as less than 100 pg per human dose, as less than 50 pg per human dose and especially less than 20 pg per human dose ( for example, 10 pg or less per human dose). Desirably, the residual organic solvent is at a rate which is in accordance with the directive of the International Council for the Harmonization of Technical Requirements for Pharmaceutical Products for Human Use for Residual Solvents Q3C (R6). Solvent removal can be accomplished by a variety of methods, which can be used individually or in combination. Suitable methods include ultrafiltration and dialysis, especially diafiltration. Removal of at least part of the solvent, like substantially all of the solvent, can be accomplished by dialysis. Dialysis is the use of a semipermeable confinement enclosure which is selectively permeable in such a way that the solvent will pass through the semipermeable part of the enclosure and the liposomes (also the saponin and the TLR4 agonist if present) will be retained when the recovered material is introduced into the semipermeable containment. For example, the semipermeable confinement enclosure used can comprise a single semipermeable membrane and the removal of the solvent can be obtained by immersing the semipermeable confinement enclosure comprising the material recovered in an exchange medium and allowing the liquids separated by the membrane to reach equilibrium by diffusion. Dialysis can be undertaken in closed or continuous operational modes. For example, dialysis can be repeated multiple times with batch replacement of the exchange medium to achieve a desired rate of removal of the solvent. Dialysis can also be in a continuous process where the recovered material and / or the exchange medium is continuously replaced. Examples of dialysis membranes that can be used in the present methods include 7 kDa membranes. Removal of at least part of the solvent, like substantially all of the solvent, can be accomplished by ultrafiltration. Ultrafiltration is the use of a containment enclosure comprising a first compartment and a second separated by a semipermeable membrane. compartments The recovered material can be placed in the first compartment of the confinement enclosure which can then be subjected to a positive pressure with respect to the second compartment so that the liquid is forced by force through the semipermeable part of Diafiltration is a form of ultrafiltration in which at least part of the remaining liquid can be replaced by an exchange medium by adding the exchange medium to the first compartment of the enclosure. Consequently, as the ultrafiltration progresses, the remaining liquid will tend towards the composition of the exchange medium. Diafiltration can be undertaken in a number of ways - continuous (also known as constant volume) in which the exchange medium is added at a rate comparable to filtration of the liquid through the membrane; discontinuous, in which the volume of the remaining liquid varies and exchange medium is added discontinuously (for example, by initial dilution and subsequent concentration to the original volume or by initial concentration and subsequent dilution to the volume d 'origin or the like). The optimal operational mode will depend on a number of factors including: 1) the initial sample volume, concentration and viscosity; 2) the final concentration required of the sample; 3) the stability of the sample at various concentrations; 4) the volume of buffer required for diafiltration; 5) total processing time; 6) the size of tank available; 7) the cost. Examples of the diafiltration membrane include Hydrosart 30 kD. The exchange medium used during the removal of the solvent need not correspond to the medium of the final liposomal adjuvant, for practical reasons, the exchange medium is suitably the medium of the final liposomal adjuvant desired or a concentrate thereof, for example, phosphate buffer solution or other buffered composition, as desired. In some processes, saponin can be added to the mixed material recovered before removal of the solvent. In other methods, saponin can be added after removal of the solvent. saponins A suitable saponin for use in the present invention is Quil A and its derivatives. Quil A is a saponin preparation isolated from the South American tree Quillaja saponaria MoLina and it has been described for the first time as having an adjuvant activity by Dalsgaard et al. in 1974 ("Saponin adjuvants", Archiv. für die gesamte Virusforschung, Vol. 44, Springer Verlag, Berlin, p 243-254). It was isolated by HPLC from the purified fractions of Quil A which retain the adjuvant activity without the toxicity associated with Quil A (see, for example, document EP 0362278). General interest fractions include QS7, QS17, QS18 and QS-21, for example QS7 and QS-21 (also known as QA7 and QA21). QS-21 is a saponin of particular interest. In certain embodiments of the present invention, saponin is a Quil A derivative of Quillaja saponaria Molina, suitably an immunologically active fraction of Quil A, such as QS7, QS17, QS18 or QS-21, in particular QS -21. Generally, saponin, like Quil A and in particular QS-21, is at least 90% pure, like at least 95% pure, especially at least 98% pure, in particular 99% pure. A beneficial feature of the present invention is that the saponin is presented in a less reactogenic composition where it is neutralized with an exogenous sterol, such as cholesterol. In processes where saponin is added after mixing the first and second solutions, the amount of saponin will generally be equivalent to the amounts that would be used if it had been added earlier. TLR4 agonists A suitable example of a TLR4 agonist is a lipopolysaccharide, suitably a non-toxic derivative of lipid A, particularly a monophosphoryl-lipid A and more particularly monophosphoryl-lipid A 3-de-O-acylated (3D-MPL) . The 3D-MPL is sold under the name "MPL" by GlaxoSmithKline Biologicals N.A. and it is named throughout the 3D-MPL document. See, for example, U.S. Patents 4,436,727; 4,877,611; 4,866,034 and 4,912,094. 3D-MPL can be produced according to the methods described in document GB 2 220 211 A. From a chemical point of view, it is a mixture of 3-deacylated monophosphoryllipid A with 4, 5 or 6 acylated chains. In the context of the present invention, small particle 3D-MPL can be used to prepare the aqueous adjuvant composition. The small particle 3D-MPL has a particle size such that it can be sterilized by filtration through a 0.22 µm filter. Such preparations are described in document WO 94/21292. Suitably, powdered 3D-MPL is used to prepare aqueous adjuvant compositions for use in the present invention Other TLR4 agonists that can be used are alkyl glucosaminide phosphates (AGP) such as those described in WO 98/50399 or US Patent No. 6,303,347 (methods for preparing AGP are also described ). Some PGAs are TLR4 agonists, and some are TLR4 antagonists. Other TLR4 agonists which can be used in the present invention include the glucopyranosyl-based lipid adjuvant (GLA) as described in documents WO 2008/153541 or WO 2009/143457 or the articles in the literature Coler RN and al. (2011) Development and Characterization of Synthetic Glucopyranosyl Lipid Adjuvant System as a Vaccine Adjuvant. PLoS ONE 6 (1): el6333. doi: 10.13 71 / journal.pone.00163 33 and Arias MA et al. (2012) Glucopyranosyl Lipid Adjuvant (GLA), a Synthetic TLR4 Agonist, Promûtes Potent Systemic and Mucosal Responses to Intranasal Immunization with HIVgpl40. PLoS ONE 7 (7): e41144. doi: 10.1371 / journal.pone.0041144 Documents WO 2008/153541 or WO 2009/143457 define TLR4 agonists which can be used in the present invention. Generally, the TLR4 agonist, such as the lipopolysaccharide and in particular 3D-MPL, is at least 90% pure, as at least 95% pure, especially at least 98% pure, in particular 99% pure . In some be added to the disposal of the solvent material process. the agonist of TLR4 can mixed recovered before In other processes, TLR4 can be added after the solvent removal agonist (in such circumstances, the amount of TLR4 will generally be equivalent to the amounts that would be used if it had been added earlier). Therefore, there is provided a method of making a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid, a sterol and the TLR4 agonist, and a second solution comprising water and saponin; and (b) removing the solvent. There is also provided a method of making a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the saponin; (c) adding the TLR4 agonist; and (d) removing the solvent; wherein steps (b) and (c) can be in one order or the other, or can be performed in a single step. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising of the water ; (B) 1'élimination solvent; (vs) adding the saponin; and (D) adding the TLR4 agonist; wherein steps (c) and (d) can be in one order or the other, or can be performed in a single step. In addition, there is provided a method of making a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a lipid of phosphatidylcholine and a sterol, and of a second solution comprising of the water ; (B) adding the saponin; (vs) 1'élimination solvent; and (D) adding the agonist of TLR4. II a process is further provided of manufacturing a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the TLR4 agonist; (c) removing the solvent; and (d) adding the saponin. Also provided is a method of making a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid, a sterol and the TLR4 agonist, and a second solution comprising water and saponin. There is also provided a method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) adding the TLR4 agonist; wherein steps (b) and (c) can be in one order or the other, or can be performed in a single step. There is also provided a method of making a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC, a sterol and the agonist of TLR4, and a second solution comprising water and saponin; and (b) removing the solvent. There is also provided a method of making a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; (c) adding the TLR4 agonist; and (d) removing the solvent; wherein steps (b) and (c) can be in one order or the other, or can be performed in a single step. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin and an aqonist of TLR4 using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and D' ' a second solution including some water ; (B) removing the solvent; (vs) 1'ajout saponin; and (D) 1'ajout the TLR4 agonist; in which steps (c) and (d) can to be in a order or in the other, where can to be done in one step. In addition, there is provided a method of making a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; (c) removing the solvent; and (d) adding the TLR4 agonist. There is further provided a method of manufacturing a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the TLR4 agonist; (c) removing the solvent; and (d) adding the saponin. Also provided is a method of making a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, mixing in the device a first solution comprising a solvent , DOPC, a sterol and The TLR4 agonist, and a second solution comprising water and saponin. There is also provided a method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water (b) Adding saponin; and the TLR4 agonist; wherein steps (b) and (c) can be in order or in or can be performed in a single step. By the expression "performed in one step" as used herein, it is meant at the same time or simultaneously. Another aspect of the invention is the liposome-containing solution which can be obtained by (as obtained by) mixing the first solution and the second solution according to any of the methods described herein. Other excipients The liposomal adjuvant resulting from the claimed methods can be further modified. For example, it can be diluted to obtain a particular concentration of the components as desired for later uses and / or additional components added. Such steps can be taken at a number of stages in the processes: before removal of the solvent, during removal of the solvent (for example, using the exchange medium) or after removal of the solvent. In another embodiment, a buffer is added to the composition. The pH of a liquid preparation is adjusted in view of the components of the composition and of a suitability necessary for administration to the subject. Suitably, the pH of a liquid mixture is at least 4, at least 5, at least 5.5, at least 5.8, at least 6. The pH of the mixture liquid may be less than 9, less than 8, less than 7.5 or less than 7. In other embodiments, the pH of the liquid mixture is between 4 and 9, between 5 and 8, as between 5, 5 and 8. Therefore, the pH will suitably be between 6 and 9, such as 6.5 to 8.5. In a particularly preferred embodiment, the pH is between 5, 8 and 6.4. An appropriate buffer can be chosen from acetate, citrate, histidine, maleate, phosphate, succinate, tartrate and TRIS buffers. In one embodiment, the buffer is a phosphate buffer such as Na / Na 2 PO 4 , Na / K 2 PO 4 or K / K 2 PO 4 . The buffer can be present in the liquid mixture in an amount of at least 6 mM, at least 10 mM or at least 40 mM. The buffer can be present in the liquid mixture in an amount less than 100 mM, less than 60 mM or less than 40 mM. It is well known that for parenteral administration, the solutions must have a pharmaceutically acceptable osmolality to avoid distortion or cell lysis. A pharmaceutically acceptable osmolality will generally mean that the solutions will have an osmolality which is approximately isotonic or slightly hypertonic. Suitably, the compositions of the present invention, when reconstituted, will have an osmolality in the range of 250 to 750 mOsm / kg , for example, the osmolality may be in the range of 250 to 550 mOsm / kg, as in the range of 280 to 500 mOsm / kg. In a particularly preferred embodiment, the osmolality can be in the range of 280 to 310 mOsm / kg. Osmolality can be measured using techniques known in the art, such as by using a commercially available osmometer, for example Advanced ™ Model 2020 available from Advanced Instruments Inc. (USA). An "isotonic agent" is a compound which is physiologically tolerated and which imparts an appropriate tone to a formulation to prevent the net flow of water through the cell membranes which are in contact with the formulation. In some embodiments, the isotonicity agent used for the composition is a salt (or mixtures of salts), conveniently, the salt is sodium chloride, suitably a concentration of approximately 150 nM. However, in other embodiments, the composition comprises a nonionic isotonic agent and the concentration of sodium chloride in the composition is less than 100 mM, as less than 80 mM, for example, less than 50 mM, as less than 40 mM, less than 30 mM and especially less than 20 mM. The ionic strength in the composition can be less than 100 mM, as less than 80 mM, for example, less than 50 mM, as less than 40 mM or less than 30 mM. In a particular embodiment, the nonionic isotonicity agent is a polyol, such as sucrose and / or sorbitol. The concentration of sorbitol can be, for example, between about 3% and about 15% (w / v), such as between about 4% and about 10% (w / v). Adjuvants comprising an immunologically active saponin fraction and a TLR4 agonist in which the isotonicity agent is a salt or a polyol have been described in document WO 2012/080369. Suitably, a human dose volume is between 0.05 ml and 1 ml, as between 0.1 and 0.5 ml, in particular a dose volume of approximately 0.5 ml, or 0.7 ml . The volumes of the compositions used may depend on the route and location of administration, with smaller doses being given intradermally. A unit dose container may contain an excess to allow proper handling of materials during administration of the unit dose. Saponin, like QS-21, can be used in amounts between 1 and 100 pg per human dose. QS-21 can be used at a rate of approximately 50 pg. Examples of suitable ranges are 40 to 60 pg, suitably 45 to 55 pg or 49 to 51 pg, such as 50 pg. In another embodiment, the human dose comprises QS-21 at a rate of about 25 pg. Examples of lower ranges include 20 to pg, suitably 22 to 28 pg or 24 to 26 pg, such as 25 pg. Human doses for children may be reduced compared to those for adults (eg, 50% reduction) 5 TLR4 agonist such as lipopolysaccharide, such as 3D-MPL, may be used in amounts between 1 and 100 pg per human dose. 3D-MPL can be used at a rate of approximately 50 pg. Examples of suitable ranges are 40 to 60 pg, suitably 45 to 55 pg or 49 to 51 pg, such as 50 pg. In another embodiment, the human dose comprises 3D-MPL at a rate of about 25 pg. Examples of lower ranges include 20 to 30 pg, suitably 22 to 28 pg or 24 to 26 pg, such as 25 pg. The human doses for children may be reduced compared to those for an adult (for example, 50% reduction). When both a TLR4 agonist and a saponin are present in the adjuvant, then the weight ratio of the TLR4 agonist to the saponin is suitably between 1/5 and 5/1, suitably 1 / 1. For example, where 3D-MPL is present in an amount of 50 pg or 25 pg, then suitably QS-21 may also be present in an amount of 50 pg or 25 pg per human dose. The saponin / DOPC ratio will generally be of the order of 1/50 to 1/10 (w / w), suitably between 1/25 and 1/15 (w / w), and preferably 1/22 to 1/18 (w / w), like 1/20 (w / w). Antigens The liposomal adjuvants prepared according to the methods of the invention can be used in conjunction with an immunogen or an antigen. In some embodiments, there is provided a polynucleotide encoding the immunogen or antigen. The liposomal adjuvant can be administered separately from an immunogen or an antigen or can be combined, either during manufacture or extemporaneously, with an immunogen or an antigen in the form of an immunogenic composition for combined administration. Therefore, there is provided a method for the preparation of an immunogenic composition comprising an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen, said method comprising the steps of: (i) preparation of a liposomal adjuvant according to the methods described here; (ii) mixing the liposomal adjuvant with an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen. Also provided is the use of a liposomal adjuvant prepared according to the methods described herein in the manufacture of a medicament. Suitably, the medicament comprises an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen. There is further provided a liposomal adjuvant prepared according to the methods described herein for use as a medicament. Suitably, the medicament comprises an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen. By the term immunogenic, it is meant a polypeptide which is capable of triggering an immune response. Suitably, the immunogen is an antigen which comprises at least one B or T lymphocyte epitope. The triggered immune response may be a response of the antigen-specific B lymphocytes, which produces neutralizing antibodies. The triggered immune response may be an antigen-specific T cell response, which may be a systemic and / or local response. The antigen-specific T cell response may include a CD4 + T cell response, such as a response involving CD4 + T cells expressing a plurality of cytokines, for example, IFN-gamma, TNF-alpha and / or 1 IL2. Alternatively, or in addition, the antigen-specific T cell response includes a CD8 + T cell response, such as a response involving CD8 + T cells expressing a plurality of cytokines, for example, IFN-gamma, TNF -alpha and / or IL2. The antigen may be derived (as obtained from) from a human or non-human pathogen including, for example, bacteria, fungi, parasitic microorganisms or multicellular parasites that infect human and non-human vertebrates, or from a cancer cell or a tumor cell. In one embodiment, the antigen is a recombinant protein, such as a recombinant prokaryotic protein. In a mode of. realization, the antigen is derived from Plasmodium spp. (such as Plasmodium falciparum), Mycobacterium spp. (such as Mycobacterium tuberculosis (TB)), varicella zoster virus (VZV), respiratory syncytial virus, human immunodeficiency virus (HIV), Moraxella spp. (such as Moraxella catarrhalis) or non-typeable Haemophilus influenzae (ntHi). The antigen may include or consist of preparations derived from parasites that cause malaria such as Plasmodium falciparum or Plasmodium vivax. In one embodiment, the antigen may be the circumsporozoite protein (CS) of Plasmodium falciparum or one of its variants. A suitable variant of the CS protein can be a variant in which parts of the CS protein are in the form of a protein hybrid with the surface antigen S originating from hepatitis B (Ag HBs). The CS variant antigen can be, for example, in the form of a hybrid protein comprising substantially all of the portion C-terminal protein CS, four or more tandem repeats of the immunodominant region of the CS protein, and HBsAg. The hybrid protein may comprise a sequence which contains at least 160 amino acids and which is substantially homologous to the C-terminal part of the CS protein, but which lacks the hydrophobic anchoring sequence. The CS protein can be devoid of the last 12 amino acids from the Cterminal end. In addition, it can contain 4 or more, for example, 10 or more repeating units of the Asn-Ala-AsnPro tetrapeptide (NANP). The hybrid protein to be used in the invention may be a protein which comprises a part of the CS protein of P. falciparum corresponding substantially to amino acids 207 to 395 of clone 3D7 of P. falciparum, derived from the strain NF54 fused in the context via a linear linker at the N-terminus of the HBsAg. The linker can comprise a part of preS2 originating from Ag HBs. CS constructs suitable for use in the present invention are presented in WO 93/10152, which is granted in the United States as US Pat. Nos. 5,928,902 and 6,169,171, describing proteins suitable for use in the present invention. A particular hybrid protein to be used in the invention is the hybrid protein known under the name of RTS (SEQ ID NO: 1), also described in documents WO 2015/150568, WO 93/10152 (in which it is indicated by RTS *) and in document WO 98/05355, which consists of: - a methionine residue - three amino acid residues, Met Ala Pro a section of 189 amino acids representing amino acids 207 to 395 of the CS protein of the 3D7 strain of P. falciparum - a residue of glycine four amino acid residues, Pro Val Thr Asn, representing the four carboxy-terminal residues of the preS2 protein of the hepatitis B virus (serotype adw), and - a segment of 226 amino acids, coded by nucleotides 1653 to 2330, and specifying protein S of the hepatitis B virus (serotype adw). The RTS can be in the form of mixed particles RTS, S. The RTS, S particles comprise two polypeptides, RTS and S, which can be synthesized simultaneously and spontaneously form composite particle structures (RTS, S). The antigen can comprise or consist of preparations derived from Mycobacterium spp., Such as Mycobacterium bovis or Mycobacterium tuberculosis, in particular Mycobacterium tuberculosis. Antigens of interest in the field of tuberculosis include Rv1196 and Rv0125. Rv1196 (described, for example, by the name of Mtb39a in Dillon et al. Infection and Immunity 1999 67 (6): 2941-2950) is highly conserved, with 100% sequence identity among the strains H37Rv, C, Haarlem , CDC1551, 94-M4241A, 98-R604INH-RIF-EM, KZN605, KZN1435, KZN4207, KZNR506, the Fil strain with a single point mutation Q30K (most other clinical isolates have more than 90% identity with H37Rv) . Rv0125 (described, for example, by the name of Mtb32a in Skeiky et al. Infection and Immunity 1999 67 (8): 3998-4007) is also highly conserved, with 100% sequence identity among many strains. Full-length Rv0125 includes an N-terminal signal sequence which is cleaved to provide the mature protein. In one embodiment, the antigen is derived from Rv1196, as comprises, as consists of, a sequence having at least 70% identity with SEQ ID NO: 2, as at least 80%, in particular at least 90 %, especially at least 95%, for example at least 98%, as at least 99%. Representative antigens related to Rv1196 will include (as will consist of) a derivative of SEQ ID NO: 2 having a small number of deletions, insertions and / or substitutions. Examples are those with deletions of up to 5 residues at 0 to 5 locations, insertions of up to 5 residues at 0 to 5 locations and a substitution of up to 20 residues. Other derivatives of Rv1196 are those comprising (as consisting of) a fragment of SEQ ID NO: 2 which has a length of at least 200 amino acids, such as a length of at least 250 amino acids, in particular a length of '' at least 300 amino acids, especially a length of at least 350 amino acids. In one embodiment, the antigen is derived from Rv0125, as comprises, as consists of, a sequence having at least 70% identity with SEQ ID NO: 3, as at least 80%, in particular at least 90 %, especially at least 95%, for example at least 98%, as at least 99%. Representative antigens related to Rv0125 will include (as will consist of) a derivative of SEQ ID NO: 3 having a small number of deletions, insertions and / or substitutions. Examples are those with deletions of up to 5 residues at 0 to 5 locations, insertions of up to 5 residues at 0 to 5 locations and a substitution of up to 20 residues. Other derivatives of Rv0125 are those comprising (as consisting of) a fragment of SEQ ID NO: 3 which has a length of at least 150 amino acids, such as a length of at least 200 amino acids, in particular a length of '' at least 250 amino acids, especially a length of at least 300 amino acids. The particular derivatives of Rv0125 are those comprising (as consisting of) the fragment of SEQ ID NO: 3 corresponding to residues 1 to 195 of SEQ ID NO: 3. Other immunogenic derivatives of Rv0125 are those comprising (as consisting of) the fragment of SEQ ID NO: 3 corresponding to residues 192 to 323 of SEQ ID NO: 3. Particularly preferred antigens related to Rv0125 are derivatives of SEQ ID NO: 3 in which at least one (for example, one, two or even all three) of the catalytic triad has been substituted or deleted, so that the protease activity has been reduced and the protein more easily produced - the catalytic serine residue can be deleted or substituted (for example, alanine substituted) and / or the catalytic histidine residue can be deleted and / or the catalytic aspartic acid residue can be deleted or substituted. Of special interest are the derivatives of SEQ ID NO: 3 in which the residue of catalytic serine has been substituted (for example, substituted by alanine). Also of interest are antigens related to Rv0125 which comprise, as consist of, a sequence having at least 70% identity with SEQ ID NO: 3, such as at least 80%, in particular at least 90%, especially at least 95%, for example at least 98%, as at least 99% and in which at least one of the catalytic triad has been substituted or deleted or those comprising, as consisting of, a fragment of SEQ ID NO: 3 which is at least 150 amino acids in length, such as at least 200 amino acids in length, in particular at least 250 amino acids in length, especially at least 300 amino acids in length and in which at least one of the catalytic triad has been substituted or deleted. Other immunogenic derivatives of Rv0125 are those comprising (as consisting of) the fragment of SEQ ID NO: 3 corresponding to residues 192 to 323 of SEQ ID NO: 3 in which at least one (for example, one, two or even even the three) of the catalytic triad has been substituted or deleted. Specific immunogenic derivatives of RvO125 are those comprising (as consisting of) the fragment of SEQ ID NO: 3 corresponding to residues 1 to 195 of SEQ ID NO: in which the residue of catalytic serine (position 176 of SEQ ID NO: 3) has been substituted (for example, substituted by alanine). Suitably, the antigen will include, as will consist of, a sequence having at least 70% identity with SEQ ID NO: 4, such as at least 80%, in particular at least 90%, especially at least 95%, like at least 98%, for example at least 99%. Representative antigens related to M72 will include, as will consist of, a derivative of SEQ ID NO: 4 having a small number of deletions, insertions and / or substitutions. Examples are those with deletions of up to 5 residues at 0 to 5 locations, insertions of up to 5 residues at 0 to 5 locations and a substitution of up to 20 residues. Other derivatives of M72 are those comprising, as consisting of, a fragment of SEQ ID NO: 4 which has a length of at least 450 amino acids, such as a length of at least 500 amino acids, such as a length of at least 550 amino acids, as a length of at least 600 amino acids, as a length of at least 650 amino acids, or a length of at least 700 amino acids. Because M72 is a fusion protein derived from the two individual antigens Rv0125 and Rv1196, any fragment of at least 450 residues will include a plurality of epitopes from the full length sequence (Skeiky et al. J. Immunol. 2004 172: 7618-7628; Skeiky Infect. Immun. 1999 67 (8): 3998-4007; Dillon Infect. Immun. 1999 67 (6): 2941-2950). An antigen related to M72 will comprise, as will consist of, a sequence having at least 70% identity with SEQ ID NO: 4, as at least 80%, in particular at least 90%, especially at least 95%, as in minus 98%, for example at least 99%. Representative antigens related to M72 will include, as will consist of, a derivative of SEQ ID NO: 4 including a small number of deletions, insertions and / or substitutions. Examples are those with deletions of up to 5 residues at 0 to 5 locations, insertions of up to 5 residues at 0 to 5 locations and a substitution of until 20 residues. In of the modes of particular realization, 1'antigène related to M72 will include residues 2 to 723 of SEQ ID NO: 4, for example will include (or will consist of) SEQ ID NO: 4 or will include (or will consist of) SEQ ID NO: 5. Another antigen which can be used in accordance with the present invention is the tuberculosis antigen Rvl753 and its variants, as described in document WO 2010010180, for example a sequence of Rvl753 chosen from SEQ ID NO: 1 and 2 to 7 of document WO 2010010180, in particular SEQ ID NO: 1. Another antigen of interest in the field of tuberculosis is Rv2386 and its variants, as described in document WO 2010010179, for example a sequence of Rv2386 chosen from SEQ ID NO: 1 and 2 to 7 of document WO 2010010179, in particular SEQ ID NO: 1. Other antigens of interest in the field of tuberculosis include Rv3616 and its variants, as described in document WO 2011092253, for example a natural sequence of Rv3616 chosen from SEQ ID NO: 1 and 2 to 7 of document WO 2011092253 or a modified sequence of Rv3616 such as those chosen from SEQ ID NO: 161 to 169, 179 and 180 of document WO 2011092253, in particular ier SEQ ID NO: 167. An additional antigen of interest is HBHA, as described in documents WO 97044463, WO 03044048 and WO 2010149657. The above-mentioned patent applications WO 2010010180, WO 2010010179, WO 2011092253, WO 97044463, WO 03044048 and WO 2010149657 define antigens which can be used in the present invention. Other antigens of interest are those comprising (or consisting of): Rv1174, also known under the name of DPV, as described by SEQ ID NO: 8 of document WO 2010010177; Rvl793, also known as MTI or Mtb9.9, as described by SEQ ID NO: 10 from WO 2010010177; Rv2087, also known as MSL or Mtb9.8, as described by SEQ ID NO: 9 of document WO 2010010177; Rv3616, also known as HTCC1 or Mtb40, as described by SEQ ID NO: 1 and 2 to 7 of document WO 2010010177 or SEQ ID NO: 161 to 169, 179 or 180 of document WO 2011092253; and / or Rv3874, also known as CFP10 or Tb3 8.1, as described by SEQ ID NO: 9 from document WO 2010010177; or one of their immunogenic parts (as at least 20, 50, 75 or 100 residues therefrom) or one of their variants (as having at least 70%, 80%, 90% or 95% therewith ). (WO 2010010177 and WO 2011092253 define antigens which can be used in the present invention). The tuberculosis antigens are most suitably used in the form of a polypeptide, but they can alternatively be provided in the form of a polynucleotide encoding said polypeptide. Another antigen which can be used in accordance with the present invention is derived from the varicella zoster virus (VZV). The VZV antigen to be used in the invention may be any suitable VZV antigen or one of its immunogenic derivatives, suitably being a purified VZV antigen. In one embodiment, the VZV antigen is the VZV gE glycoprotein (also known as gpl) or its immunogenic derivatives. The wild-type or full-length gE protein consists of 623 amino acids comprising a signal peptide, the main part of the protein, a hydrophobic anchoring region (residues 546 to 558) and a C-terminal tail. In one aspect, a truncated gE at the terminal end (also called truncated gE or truncated form of gE) is used whereby truncation removes 4 to 20 percent of the total amino acid residues at the carboxy-terminal end . In another aspect, in truncated gE, the carboxy-terminal anchoring region is lacking (suitably, approximately amino acids 547 to 623 of the wild type sequence). In another aspect, gE is a truncated gE having the sequence of SEQ ID NO: 6. The gE antigen, its anchored derivatives (which are also immunogenic derivatives) and their production are described in document EP 0405867 and the references therein (see also Vafai A., Antibody binding sites on truncated forms of varicallazoster virus gpl (gE) glycoprotein, Vaccine 1994 12: 1265-9). Document EP 192902 also describes gE and its production. Truncated gE is also described by Haumont et al. Virus Research (1996) vol 40, pl99 204. An adjuvanted composition of VZV gE for use in accordance with the present invention is described in WO 2006/094756, i.e., gE of VZV truncated to the carboxy-terminal end in combination with an adjuvant comprising QS-21, 3D-MPL and liposomes additionally containing cholesterol. Leroux-Roels I. et al. (J. Infect. Dis. 2012, 206: 1280-1290) reported a phase I / II clinical trial evaluating the truncated VZV gE trunked subunit vaccine. The antigen may include or consist of preparations derived from the human respiratory syncytial virus (RSV). In certain favorable embodiments, a polypeptide antigen is an F protein polypeptide antigen derived from RSV. Are forced. particularly constraints both 'antigen component background antigens, protein previously have been described both prefusion (PreF) and postfusion (PostF) conformations. Such conformally constrained F proteins generally include a modified ectodomain of the RSV F protein. An ectodomain of the F protein polypeptide is a part of the RSV F protein which comprises all or part of the extracellular domain of the F protein of RSV and which lacks a functional transmembrane domain (for example, by deletion or substitution), which can be expressed, for example, in soluble form (not attached to a membrane) in cell culture. Examples of protein F antigens conformally constrained in the prefusion conformation have been described in the art and are disclosed in detail, for example, in US Patent No. 8,563,002 (WO 2009079796); published US patent applications No. US 2012/0093847 (WO 2010/149745); US 2011/0305727 (WO 2011/008974); US 2014/0141037, WO 2012/158613 and WO 2014/160463, illustrate F polypeptides in prefusion (and nucleic acids), and methods for their production. Generally, the antigen is in the form of a trimer of polypeptides. Other publications providing examples of F proteins in the prefusion conformation include: McLellan et al., Science, Vol. 340: 1113-1117; McLellan et al., Science, Vol 342: 592-598, and Rigter et al., PLOS One, Vol. 8: e71072, each of them can also be used in the context of the immunogenic combinations disclosed here. For example, an F protein polypeptide stabilized in the prefusion conformation generally comprises an ectodomain of an F protein (for example, a soluble F protein polypeptide) comprising at least one modification which has stabilized the prefusion conformation of the F protein For example, the modification can be chosen from an addition of a trimerization domain (generally at the C-terminal end), the deletion of one or more of the furin cleavage sites (at the amino acid level -105 to 109 and -133 to 136), a deletion of the pep27 domain, the substitution or addition of a hydrophilic amino acid in a hydrophobic domain (for example, HRA and / or HRB). In one embodiment, the conformally constrained PreF antigen comprises an F2 domain (for example, amino acids 1 to 105) and a Fl domain (for example, amino acids 137 to 516) of an F protein polypeptide RSV without any furin cleavage site involved in the leguel, the polypeptide further comprises a heterologous trimerization domain in the C-terminal position relative to the Fl domain. Optionally, the PreF antigen also includes a modification which changes the glycosylation (by example, increases glycosylation), as a substitution of one or more amino acids at positions corresponding to amino acids -500 to 502 of an RSV F protein. When an oligomerization sequence is present, it is preferably a trimerization sequence. Appropriate oligomerization sequences are well known in the art and include, for example, the leucine zipper protein helix of the yeast GCN4, the fibritin trimerization sequence of bacteriophage T4 ("fold"), and the field trimer of HA from influenza virus. Additionally or alternatively, the conformally constrained F polypeptide in prefusion may include at least the conformation two introduced cysteine residues, which are in close proximity to each other and form a disulfide bond which stabilizes the F polypeptide of the RSV in pre-merger. For example, the two cysteines can be about 10 Å apart. For example, cysteines can be introduced at positions 165 and 296 or at positions 155 and 290. An example of PreF antigen is represented by SEQ ID NO: 7. The antigen may include or consist of preparations derived from HIV. The antigen can be an HIV protein such as an HIV envelope protein. For example, the antigen may be a gpl20 polypeptide of the HIV envelope or one of its immunogenic fragments. A suitable antigen is the HIV clade B gpl20 polypeptide of SEQ ID NO: 8 of published application WO 2008/107370 (or an immunogenic fragment of this polypeptide). Suitable antigens also include a polypeptide comprising the V1V2 region of SEQ ID NO: 1 of published application WO 2015/036061, or an immunogenic derivative or fragment of the VIV2 region of SEQ ID NO: 1. In addition, a polypeptide comprising V1V2 region of SEQ ID NO: 5 of document WO 2015/036061 or an immunogenic derivative or fragment of the V1V2 region of SEQ ID NO: 5 can be used as a suitable antigen. In another embodiment, the antigen may include two or more different gpl20 HIV envelope polypeptide antigens (or immunogenic fragments of these polypeptides). Suitable antigens include the HIV clade C polypeptide gpl20 antigens including TV1 gpl20 (SEQ ID NO: 8) and 1086C gpl20 (SEQ ID NO: 9). Other suitable HIV antigens include the HIV Nef, Gag and Pol proteins and their immunogenic fragments. The composition can comprise one or more non-typable Haemophilus influenzae antigens, for example chosen from: the fimbrin protein [(US 5766608 - Ohio State Research Foundation)] and fusions comprising peptides derived therefrom [for example, c ies mergers of LB1 (f) peptides; US 5843464 (OSU) or WO 99/64067]; OMP26 [WO 97/01638 (Cortecs)]; P6 [EP 281673 (State University of New York)]; TbpA and or TbpB; Hia; Hsf; Hin47; Hif; Hmwl; Hmw2; Hmw3; Hmw4; Map; D15 (WO 94/12641); protein D (EP 594610); P2; and P5 (WO 94/26304); protein E (WO 07/084053) and or Pila (WO 05/063802). The composition can understand a or many Moraxella catarrhalis protein antigens, for example, selected from: OMP106 [WO 97/41731 (Antex) and WO 96/34960 (PMC)]; OMP21; LbpA and / or LbpB [WO 98/55606 (PMC)]; TbpA and / or TbpB [WO 97/13785 and WO 97/32980 (PMC)]; CopB [Helminen ME, et al. (1993) Infect. Immun. 61: 2003-2010]; UspAl and / or UspA2 [WO 93/03761 (University of Texas)]; OmpCD; HasR (PCT / EP99 / 03824); PilQ (PCT / EP99 / 03823); OMP85 (PCT / EP00 / 01468); lipo06 (GB 9917977.2); lipolO (GB 9918208.1); lipoll (GB 9918302.2); lipo! 8 (GB 9918038.2); P6 (PCT / EP99 / 03038); D15 (PCT / EP99 / 03822); OmplAl (PCT / EP99 / 06781); Hly3 (PCT / EP99 / 03257); and OmpE. In one embodiment, the composition may include one or more non-typeable H. influenzae protein antigens (NTHi) and / or one or more M. catarrhalis protein antigens. The composition may include protein D (PD) from H. influenzae. Protein D can be as described in document WO 91/18926. The composition can also comprise protein E (PE) and / or pilin A (PilA) from H. Influenzae. Protein E and pilin A can be as described in document WO 2012/139225. Protein E and pilin A can be in the form of a fusion protein; for example, LVL735 as described in document WO 2012/139225. For example, the composition may include three NTHi antigens (PD, PE and PilA, with the latter two combined as a PEPilA fusion protein). The composition can also comprise UspA2 from M. catarrhalis. UspA2 can be as described in document WO 2015125118, for example MC-009 ((M) (UspA2 31564) (HH)) described in document WO 2015125118. For example, the composition can comprise three antigens of NTHi (PD, PE and PilA, with the latter two combined as a fusion protein PEPilA) and an antigen of M. catarrhalis (UspA2). A plurality of antigens can be provided. For example, a plurality of antigens may be provided to enhance the triggered immune response (for example, to provide a plurality of antigens may be provided to enhance the immune response (for example, to provide protection against a variety of pathogenic strains or in a large proportion of a population of subjects) or it may be provided a currently trigger immune responses in relation to a number of disorders (thereby simplifying administration protocols). a plurality of antigens, these may be in the form of separate proteins or they may be in the form of one or more fusion proteins. The antigen can be supplied in an amount of 0.1 to 100 µg per human dose. The present invention can be applied for use in the treatment or prophylaxis of a disease or disorder associated with one or more of the antigens described above. In one embodiment, the disease or disorder is selected from malaria, tuberculosis, COPD, HIV and herpes. The liposomal adjuvant can be administered separately from an immunogen or an antigen, or it can be combined, either during manufacture or extemporaneously, with an immunogen or an antigen in the form of an immunogenic composition for combined administration. Sterilization For parenteral administration in particular, the compositions should be sterile. Sterilization can be carried out by a variety of methods although it is conveniently undertaken by filtration through a sterile quality filter. Sterilization can be carried out a number of times during the preparation of an adjuvant or immunogenic composition, but it is generally carried out at least at the end of manufacture. By "sterile quality filter" is meant a filter which produces a sterile effluent after a test by microorganisms at a test rate greater than or equal to 1 x 10 7 / cm 2 of effective filtration surface. Filters of sterile quality are well known to those skilled in the art of the invention. For the purpose of the present invention, sterile quality filters have a pore size between 0.15 and 0.25 µm, suitably 0.18 to 0.22 µm, such as 0.2 or 0.22 pm. The membranes of the sterile quality filter can be made of any suitable material known to those skilled in the art, for example, but not limited to, cellulose acetate, polyethersulfone (PES), polyvinylidene fluoride ( PVDF), polytetrafluoroethylene (PTFE). In a particular embodiment of the invention, one or more or all of the membranes of the filter of the present invention comprise polyethersulfone (PES), in particular hydrophilic polyethersulfone. In a particular embodiment of the invention, the filters used in the methods described here are a double layer filter, in particular a sterile filter with an integrated prefilter having a pore size greater than the pore size of the final filter. In one embodiment, the sterilization filter is a double layer filter in which the layer of the prefilter membrane has a pore size between 0.3 and 0.5 nm, such as 0.35 or 0.45 nm . According to other embodiments, the filters comprise one or more asymmetric filter membranes, such as one or more asymmetric filter membranes of hydrophilic PES. Alternatively, the layer of the sterilization filter can be made of PVDF, for example, in combination with a layer of hydrophilic PES asymmetric prefilter membrane. In the light of the intended medical uses, the materials must be of pharmaceutical quality (as of parenteral quality). By the term "substantially" with respect to a whole number, it is meant functionally comparable, so that a deviation can be tolerated if the essential nature of the whole number is not changed. For example, with regard to specific values, the term "substantially" will generally mean a value within plus or minus 10 percent of the indicated value. In summary, the present invention provides: 1. A process for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. 2. A method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising (b) adding saponin; and (c) Removal of the solvent. 3. A process for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) removing the solvent; and (c) adding the saponin. 4. A method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent, a lipid of phosphatidylcholine and a sterol, and a second solution comprising water and saponin. 5. A method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; and (b) adding the saponin. 6. A method for manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) adding the TLR4 agonist; wherein steps (b) and (c) can be in one order or the other, or can be performed in a single step. 7. The process for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device according to point 1, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. 8. The method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device according to point 2, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. 9. The method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device according to point 3, comprising the following steps: (a) the mixing in the device of a first solution comprising a solvent, of DOPC and a sterol and a second solution comprising some water ; (B) solvent removal ; and (vs) adding saponin. 10. The manufacturing process of a concentrated liposomal for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device according to point 4, comprising the step of mixing in the device a first solution comprising a solvent, DOPC and a sterol, and of a second solution comprising water and saponin. 11. The process for manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device according to point 5, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; and (b) adding the saponin. 12. The process for the manufacture of a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device according to point 6, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) adding the TLR4 agonist; wherein steps (b) and (c) can be in one order or the other, or can be performed in a single step. 13. The method according to any one of points 1 to 5 or 7 to 11, wherein the first solution further comprises a TLR4 agonist. 14. The method according to any one of points 1 to 3 or 7 to 9, in which a TLR4 agonist is added before removing the solvent. 15. The method according to any one of points 1 to 3 or 7 to 9, wherein a TLR4 agonist is added after removal of the solvent. 16. The method according to any one of points 1 to 15, in which the microfluidic device has an inlet for the first solution to the mixing chamber. 17. The method according to any one of points 1 to 15, in which the microfluidic device has two inlets for the first solution to the mixing chamber. 18. The method according to any one of points 1 to 15, in which the microfluidic device has three or more inlets for the first solution to the mixing chamber. 19. The method according to any one of points 1 to 18, in which the microfluidic device has an inlet for the second solution to the mixing chamber. 20. The method according to any one of points 1 to 18, in which the microfluidic device has two inlets for the second solution‘to the mixing chamber. ' 21. The method according to any one of points 1 to 18, wherein the microfluidic device has three or more inlets for the second solution to the mixing chamber. 22. The method according to any one of points 1 to 21, wherein each inlet has a width of 0.2 mm and covers the full length of the other side of the mixing chamber. 23. The method according to any one of points 1 to 22, wherein the cross section of the mixing chamber is 25.6 mm 2 or less. 24. The method according to any one of points 1 to 23, wherein the cross section of the mixing chamber is 0.1 mm 2 or greater. 25. The method according to any one of points 22 to 24, wherein the cross section of the mixing chamber is 0.2 to 3.2 mm 2 . 26. The method according to point 25, in which the cross section of the mixing chamber is 0.6 to 1.2 mm 2 , as around 0.8 mm 2 . 27. The method according to any one of points 1 to 26, wherein the mixing chamber is substantially rectangular in cross section. 28. The method according to point 27, in which the cross section of the mixing chamber has a side 1 to 8 mm long. 29. The method according to point 28, in which the cross section of the mixing chamber has a long side from 1.6 to 2.4 mm. 30. The method according to point 29, in which the cross section of the mixing chamber has a side 2 mm long. 31. The method according to any one of points 1 to 30, wherein the cross section of the mixing chamber has a short side of 0.1 to 4 mm. 32. The method according to point 31, in which the cross section of the mixing chamber has a short side of 0.2 to 0.6 mm. 33. The method according to point 32, in which the cross section of the mixing chamber has a short side of 0.4 mm. 34. The method according to any one of points 1 to 33, wherein the mixing chamber has a length of 1 to 10 cm. 35. The method according to point 34, in which the mixing chamber has a length of 2 to 3 cm. 36. The method according to point 35, in which the mixing chamber has a length of 2.5 cm. 37. The method according to any one of points 1 to 36, in which the microfluidic device has an outlet from the mixing chamber for the recovery of the mixed material. 38. The method according to any one of points 1 to 37, wherein the microfluidic device has two or more outlets from the mixing chamber for the recovery of the mixed material. 39. The method according to any one of points 1 to 38, wherein the microfluidic device comprises a mixing chamber which is rectangular in cross section, having a cross section of 0.2 to 3.2 mm 2 , one side 1.4 to 3.2 mm long, 0.1 to 1.2 mm short side, an inlet for the first solution and two outlets for the second solution which are arranged symmetrically at the proximal end of the mixing, a mixing chamber length of 1.5 to 5 cm and an outlet located at the distal end of the mixing chamber. 40. The method according to any one of points 1 to 39, in which the total flow rate in the mixing chamber is 12 to 30 ml / min / mm 2 of cross section of the mixing chamber. 41. The method according to point 40, in which the total flow rate in the mixing chamber is 17.5 to 25 ml / min / mm 2 of cross section of the mixing chamber. 42. The method according to point 41, in which the total flow rate in the mixing chamber is 20 ml / min / mm 2 of cross section of the mixing chamber. 43. The method according to any one of points 1 to 42, wherein the ratio of the flow rates for the first and second solutions is in the range of 1/2 to 1/6. 44. The method according to point 43, in which the ratio of the flow rates for the first and second solutions is in the range of 1/3 to 1/5. 45. The method according to point 44, in which the ratio of the flow rates for the first and second solutions is 1/4. 46. The method according to any one of points 1 to 45, in which the flow rate of the first solution in the mixing chamber is from 2 to 7.5 ml / min / mm 2 of cross section of the mixing chamber. 47. The method according to point 46, in which the flow rate of the first solution in the mixing chamber is 3 to 6.5 ml / min / mm 2 of cross section of the mixing chamber. 48. The method according to point 47, in which the flow rate of the first solution in the mixing chamber is 5 ml / min / mm 2 of cross section of the mixing chamber. 49. The method according to any one of points 1 to 48, in which the flow rate of the second solution in the mixing chamber is 11 to 25 ml / min / mm 2 of cross section of the mixing chamber. 50. The method according to point 49, in which the flow rate of the second solution in the mixing chamber is 14 to 20 ml / min / mm 2 of cross section of the mixing chamber. 51. The method according to any one of points 1 to 50, in which the first solution is supplied at a temperature of 10 to 30 ° C. 52. The method according to point 51, in which the temperature of the first solution is supplied at a temperature of 15 to 25 ° C. 53. The method according to any one of points 1 to 52, wherein the temperature of the second solution is supplied at a temperature of 10 to 30 ° C. 54. The method according to point 53, in which the temperature of the second solution is supplied at a temperature of 15 to 25 ° C. 55. The method according to any one of points 1 to 54, wherein the temperature of the mixing chamber is 10 to 30 ° C. 56. The method according to point 55, in which the temperature of the mixing chamber is 15 to 25 ° C. 57. The method according to any one of points 1 to 56, wherein the maximum Reynolds number within the mixing chamber is 1500 or less. 58. The method according to point 57, in which the maximum Reynolds number within the mixing chamber is 75 to 300, like 100 to 200. 59. The method according to any one of points 1 to 58, wherein the microfluidic device comprises a plurality of mixing chambers. 60. The method according to point 59, in which the device comprises 2 to 128 mixing chambers. 61. The method according to point 60, in which the device comprises 4 to 32 mixing chambers. 62. The method according to point 61, in which the device comprises 16 mixing chambers. 63. The method according to any one of points 59 to 62, in which all of the mixing chambers in the plurality of mixing chambers are fed by the same pumps and the mixed material from all of the mixing chambers is collected before a other processing and / or storage. 64. The method according to any of points 59 to 63, wherein the plurality of mixing chambers is capable of producing the mixed material at a rate of 50 to 2000 ml / min. 65. The method according to any one of points 59 to 64, wherein the plurality of mixing chambers is capable of producing the mixed material at a rate of at least 1 g of phosphatidylcholine lipid per minute. 66. The method according to any one of points 59 to 65, wherein the plurality of mixing chambers is capable of producing the mixed material at a rate of at least 1 g of DOPC per minute. 67. The process according to any one of points 1 to 66, wherein the solvent comprises an organic alcohol. 68. The process according to point 67, in which the solvent includes 1 ethanol. 69. The process according to point 68, in which the solvent includes 70 90% v / v ethanol - 70. The process according to point 69, in which the solvent includes 75 85% v / v ethanol • 71. The process according to point 70 in which the solvent includes 80 % v / v ethanol. 72. The process according to any of points 67 to 71, in which the ; solvent includes 1 isopropanol. 73. The process according to point 72, in which the solvent includes 10 at 30% v / v isopropanol. 74. The process according to point 73, in which the solvent includes 15 at 25% v / v isopropanol. 75. The process according to point 74, in which the solvent includes 20 % v / v of isopropanol. 76. The method according to any one of points 1 to 75, in which the first solution comprises 100 to 170 mg / ml phosphatidylcholine lipid. 77. The process according to the point 76 in which the first solution includes 100 to 160 mg / ml of lipid of phosphatidylcholine. 78. The process according to the point 77, in which the first solution comprises 130 mg / ml of lipid of phosphatidylcholine. 79. The method according to any one of points 1 to 78, in which the first solution comprises 100 to 170 mg / ml DOPC. 80. The method according to point 79, in which the first solution comprises 100 to 160 mg / ml of DOPC. 81. The method according to point 80, in which the first solution comprises 130 mg / ml of DOPC. 82. The method according to any one of points 1 to 81, wherein the first solution comprises 20 to 50 mg / ml of sterol. 83. The method according to any one of points 1 to 82, wherein the first solution comprises 30 to 35 mg / ml sterol. 84. The method according to any one of items 1 to 83, wherein the sterol is cholesterol. 85. The method according to any one of points 1 to 84, wherein the dry weight of the first solution is 120 to 250 mg / ml. 86. The method according to any one of points 1 to 85, wherein the second solution comprises at least 90% w / w of water. 87. The method according to point 86, in which the second solution comprises at least 98% w / w of water. 88. The process according to any one of points 1 to 87, in which the saponin is Quil A or one of its derivatives. 89. The method according to point 88, in which the saponin is QS-21. 90. The method according to any one of points 1 to 89, wherein the second solution comprises 0.15 to 15 mg / ml of saponin. 91. The method according to point 90, in which the second solution comprises 1 to 4 mg / ml of saponin. 92. The method according to any one of points 6 or 12 to 91, wherein the TLR4 agonist is a lipopolysaccharide. 93. The method according to point 92, in which the lipopolysaccharide is 3D-MPL. 94. The method according to points 13 or 16 to 93, in which the first solution comprises 4 to 10 mg / ml of the agonist of TLR4. 95. The method according to any one of points 1 to 94, wherein the average size of the liposomes is 95 to 120 nm. 96. The method according to any one of items 1 to 95, wherein the polydispersity of the liposomes is 0.3 or less. 97. The method of item 96, wherein the polydispersity of the liposomes is 0.2 or less. 98. The process according to any one of points 1 to 97, in which the solvent is removed by diafiltration, ultrafiltration and / or dialysis, in particular by diafiltration. 99. The process according to any one of points 1 to 98, in which the removal of the solvent results in a water content of at least 98% w / w of water. 100. The process according to any one of points 1 to 99, including the further dilution step, as to a desired final concentration. 101. The method according to any one of items 1 to 100, comprising the additional step d n adjusting the pH to 5-9. 102. The method according to any one of points 1 to 101, comprising the additional step of adjusting the osmolality to 250 to 750 mOsm / kg. 103. A process for the preparation of an adjuvanted immunogenic composition comprising an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen, said process comprising the steps of: (i) manufacture of a liposomal adjuvant according to the method of any one of points 1 to 102; (ii) mixing the liposomal adjuvant with an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen. 104. A method for manufacturing an adjuvanted immunogenic composition, said method comprising the step of combining an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen, with an adjuvant liposomal produced according to the method of any one of points 1 to 102. 105. The method according to point 103 or 104, in which the antigen is derived from Plasmodium spp. (such as Plasmodium falciparum), Mycobacterium spp. (such as Mycobacterium tuberculosis (TB)), varicella zoster virus (VZV), human respiratory syncytial virus, human immunodeficiency virus (HIV), Moraxella spp. (such as Moraxella catarrhalis) or non-typeable Haemophilus influenzae (ntHi). 106. The method according to any one of points 1 to 105, comprising the additional step of sterilization by filtration. 107. A liposomal adjuvant comprising a saponin, a TLR4 agonist, DOPC and sterol produced according to the method of any one of points 1 to 106. 108. An adjuvanted immunogenic composition produced according to the method of point 104 or 105. 109. The adjuvant or the immunogenic composition according to point 107 or 108 comprising a saponin, such as QS-21, in an amount of 1 to 100 pg per human dose. 110. The adjuvant or the immunogenic composition according to any one of points 107 to 109 comprising a TLR4 agonist, such as 3D-MPL, in an amount of 1 to 100 μg per human dose. 111. A solution comprising a solvent and 100 to 170 mg / ml of lipid, in which the solvent comprises 70 to 90% v / v of ethanol, such as 75 to 85% v / v of ethanol, and 10 to 30% v / v isopropyl alcohol such as 15 to 25% v / v isopropanol. 112. The solution according to point 111, in which the lipid East a phosphatidylcholine. 113. The solution according to point 111, in which the lipid East the DOPC. 114. The solution according to any of points 111 to 113, comprising 100 to 160 mg / ml of lipid. 115. The solution according to point 114, comprising 120 to 140 mg / ml of lipid, such as 130 mg / ml. 116. The solution according to any one of points 111 to 115, further comprising 20 to 50 mg / ml of sterol. 117. The solution according to any one of points 111 to 116, in which the ratio of lipid to sterol is from 3/1 to 5/1. 118. The solution according to point either 111 or 117, in which the sterol is cholesterol. 119. The solution according to any one of points 111 to 118, comprising a TLR4 agonist. 120. The solution according to point 119, in which the TLR4 agonist is present at a concentration of 4 to 12 mg / ml. 121. The solution according to any one of points 111 to 120 which consists essentially of a solvent and from 100 to 160 mg / ml of DOPC and from 30 to 40 mg / ml of cholesterol, from 4 to 10 mg / ml TLR4 agonist, and in laguelle the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v of isopropyl alcohol. 122. The solution according to any one of points 111 to 121 in which the TLR4 agonist is a lipopolysaccharide, such as 3D-MPL. 123. A process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with the phosphatidylcholine lipid and cholesterol; (iii) adding other solvents; (iv) the mixture. 124. A process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with the phosphatidylcholine lipid and cholesterol; (iii) adding other solvents; (iv) the mixture; (v) adding additional solvent. 125. The process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist according to point 123, said process comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with DOPC and cholesterol; (iii) adding other solvents; (iv) the mixture. 126. A process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist according to point 124, said process comprising the steps: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with DOPC and cholesterol; (iii) adding other solvents; (iv) the mixture; (v) adding additional solvent. 127. The process according to any one of points 123 to 126, wherein the mixture is at a temperature of 30 to 50 ° C. 128. The process according to point 127, in which the mixture is at a temperature of 35 to 45 ° C. 129. The process according to point 128, in which the mixture is at a temperature of 40 ° C. 130. The process according to any one of points 123 to 129, wherein the at least part of the solvent is at least 25% of the solvent. 131. The process according to point 130, wherein the at least part of the solvent is at least 35% of the solvent. 132. The process according to point 131, in which the at least part of the solvent is at least 45% of the solvent. 133. The process according to any one of points 123 to 132, wherein the at least part of the solvent is 90% of the solvent or less as 80% or less, especially 70% or less and in particular 60% or less . 134. The process according to any one of points 123 to 133, wherein other solvent is any remaining solvent. 135. The method according to any one of points 123 to 134, in which the solution comprises 100 to 160 mg / ml of lipid and 30 to 40 mg / ml of cholesterol, in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v of isopropyl alcohol, in which the lipid is appropriately DOPC. 136. The method according to any one of points 123 to 135, in which the agonist of TLR4 is 3D-MPL. 137. The method according to point 136, in which the solution comprises 4 to 10 mg / ml of 3D-MPL. 138. The process according to any one of points 124 to 137, wherein the additional solvent is 0 to 30% of the solvent. 139. A solution containing liposomes obtainable by mixing the first solution and the second solution according to the methods of any one of points 1 to 139 before removing the solvent. 140. The process, the adjuvant, the composition or the solution according to any one of points 1 to 139, in which the phosphatidylcholine lipid contains saturated unbranched acyl chains containing 12 to 20 carbon atoms as chains acyl having 14 to 18 carbon atoms. 141. The process, the adjuvant, the composition or the solution according to any one of points 1 to 139, in which the phosphatidylcholine lipid contains unbranched acyl chains containing 12 to 20 carbon atoms and a double bond , such as acyl chains having 14 to 18 carbon atoms and a double bond. 142. The process, the adjuvant, the composition or the solution according to any one of points 1 to 141, in which / which the phosphatidylcholine lipid is chosen from dilauroyl-phosphatidylcholine (DLPC), dimyristoyl-phosphatidylcholine dipalmitoyl- phosphatidylcholine phosphatidylcholine phosphatidylcholine phosphatidylcholine (DOPC); and their mixtures. (DMPC), (DSPC) (DAPC), (DPPC), and la la distéaroyldiarachidoyldipalmitoléoylet dioleoyl-phosphatidylcholine A composition or method or process defined as "comprising" is understood to include a composition, method or process (respectively) made up of these elements. As used herein, "consisting essentially of" means that additional components may be present provided that they do not change the properties or the overall function. The invention will be further described with reference to the following nonlimiting examples. Examples General experimental details Microfluidic device with single mixing chamber and general operation Device FIG. 1 illustrates the design of an example of a microfluidic device comprising a mixing chamber on a single chip. The device comprises a mixing chamber with a length of 2.5 cm and having an elongated cross section of 2 mm by 0.4 mm. The mixing chamber has an inlet located in the center for the supply of the first solution and two inlets for the supply of the second solution. Each of the inlets is 0.2 mm wide and covers the full length of the other side of the mixing chamber. A single outlet is located at the distal end of the mixing chamber. Operation To perform microfluidic experiments, medium pressure Cetoni neMesys syringe pumps, Cetoni glass syringes and a Micronit chip holder containing the device were placed in a temperature controlled area (Sartorius Certomat). Before any experimental series, the system is cleaned and allowed to stabilize at the fixed temperature. Product collection and solvent removal The concentrated liposomes collected were divided into 2 parts: The first part was diluted with phosphate buffer solution (PBS) pH 6.1 to reach a final concentration of 2 mg / ml of DOPC and filtered through a 0.22 µm polyethersulfone (PES) membrane. Analysis of the composition (DOPC, cholesterol, 3D-MPL, QS-21) was carried out on this sample. The second part was dialyzed (device 7000MWCO Thermo with phosphate buffer solution pH 6.1 to remove the organic solvent. The protocol used was: 2 x 15 min, 2 x min and overnight (1 1 of PBS buffer pH 6.1 at each time point). The retentate was then diluted to 2 mg / ml DOPC and filtered through a 0.22 µm PES membrane. Size measurements were undertaken on this sample. The residual alcohol was tested on this sample by gas chromatography. Microfluidic device with multiple mixing chambers and general operation Device FIG. 2 illustrates the design of an example of a microfluidic device with multiple mixing chambers comprising eight mixing chambers on a single chip. The device comprises eight mixing chambers with a length of 2.5 cm and having an elongated cross section of 2 mm by 0.4 mm. Each mixing chamber has an inlet located in the center for the supply of the first solution and two inlets for the supply of the second solution. Each of the inlets is 0.2 mm wide and covers the full length of the other side of the mixing chamber. A single outlet is located at the distal end of each of the mixing chambers. Figure 3 illustrates a tubing design that can be used in conjunction with a microfluidic chip with multiple mixing chambers, to provide the first or second solution to the inlets of sixteen mixing chambers, or to collect mixed material from the outlets sixteen mixing chambers. FIG. 4 is a representation of an example of a microfluidic device with multiple mixing chambers comprising a total of sixteen mixing chambers based on two chips of the style represented in FIG. 2 (occupying the two central supports), together with a manifold distribution supplying the inputs of the first solution of the sixteen chambers (support at the bottom left), a distribution pipe supplying the inputs of the second solution of the sixteen chambers (support at the top left) and a collection tubing which groups the outputs of the sixteen bedrooms (support on the right). Operation The multiple mixing chamber device can operate in a similar fashion to the single device. For example, organic stock (e.g. 4.9 L) can be prepared by containing DOPC 130 mg / ml, cholesterol 32.5 mg / ml and 3D-MPL 6.5 mg / ml in 80/20 ethanol / IPA. The aqueous phase (for example, 19.7 l) can be composed of QS-21 at 1.625 mg / ml diluted in water for injection. Appropriate pumps, such as Isco 500D in tandem for the organic phase and 1000D in tandem for the aqueous phase, can be used in continuous flow to supply the liquid phases in the 100 pipes dividing the streams into 16 streams which enter the 16 mixing chambers arranged in parallel. At the end of the mixing chambers, other tubing can be used to collect the mixed material containing the concentrated liposomes in a container. Diafiltration can be used to remove the organic solvent from the mixed material and replace the water for injection with an appropriate buffer (such as PBS pH 6.1). Another dilution with an appropriate buffer (for example, PBS pH 6.1) makes it possible to obtain the final composition according to the desired concentration of the components. Sterilization by filtration can then be undertaken. Analytical procedures - Size measurement Size measurements make use of the DLS principle with a Zetasizer from Malvern. The samples were diluted in the corresponding buffer (usually PBS pH 6.1) for the measurements. - 3D-MPL content HPLC coupled with fluorescence detection was used to quantify the 3D-MPL component. The separation was carried out on a column C18. The standards were prepared from an equimolar mixture of glucosamine HCl and glucosamine-6phosphate reconstituted in the liposomal matrix (DOPC, cholesterol). 101 The samples and standards are derivatized with acid under strong reducing conditions. - QS-21 content HPLC coupled with UV detection was used to quantify the QS-21 component. The separation is carried out on a column C18. The standards are prepared with a reference of QS-21 diluted in DMSO from 25 to 75 pg / ml The samples are diluted in DSMO for analysis. - Content of DOPC-Chol processes were used: The first method (individual standards) used a U-HPLC coupled to a UV detector. The separation was carried out on a column C18. The standards were prepared with DOPC / Chol diluted in IPA / CHC13 for the stock and diluted in the same buffer from 0 to 700 pg / ml for DOPC and from 0 to 175 pg / ml for cholesterol. The samples are diluted in IPA / CHC13. The second method (relating to an adjuvant composition characterized previously) used an HPLC coupled to a UV detector. The separation is carried out on a column C18. The standards are prepared using an adjuvant composition previously characterized, prepared through of the conventional means, diluted in of methanol from 0 at 500 pg / ml for the DOPC and from 0 at 125 pg / ml for the cholesterol. The samples are diluted in methanol. - Residual solvents 102 Process using gas chromatography coupled to a headspace injector and an FID (flame ionization detector). The separation is carried out on an Agilent CP WAX52-CB column. The standards are prepared for each specific organic solvent (in this case, IPA and ethanol) from 2 to 160 pg / ml. The LOQ is at 2 pg / ml. Samples are diluted to fall within the range of standards. Example 1 - Study of the preparation processes and of the composition of the first solution Example IA - Composition of the solvent Process To study the impact of the composition of the solvent on the production of liposomes, solutions of DOPC, cholesterol and 3D-MPL were prepared in various ratios of ethanol / isopropyl alcohol. DOPC, cholesterol and 3D-MPL were each solubilized individually (60% by volume for DOPC, 20% for cholesterol and 20% for 3D-MPL) for 15 minutes at 55 ° C. The 3DMPL solution was then added to the DOPC solution and this mixture was added to the cholesterol solution and further mixed for another 15 minutes to provide final compositions with 150 mg / ml DOPC (20/5/1 , weight ratio of DOPC / cholesterol / 3D-MPL). The single-chamber microfluidic device operated with a total flow rate of 14 ml / min, ratio 103 flow rates of 20 (19/1) (1/19, organic / aqueous), using water for injection as the aqueous phase, with stock solutions and an environment at room temperature. Results Table 1 Impact of the composition of the solvent on the size of the liposomes Ethanol / IPA ratio Liposome size(Nm) 100/0 - 80/20 124 70/30 139 60/40 154 50/50 174 conclusions Pure ethanol was unable to solubilize the components at the target concentration. Although pure isopropanol was able to dissolve the components at the target concentration, the liposomes produced in a similar experiment (160 mg / ml) were very large with a diameter of 203 nm. Mixtures of ethanol and IPA, particularly in the range of 90/10 to 70/30 provide good solubilization capacity with low viscosity. Example IB - Preparation of the solution 104 Process The order of adding the components was evaluated by comparing the following two methods: DOPC, cholesterol and 3D-MPL each solubilized%, 20% by volume individually have summer (60 o.* 6 f in a minutes ethanol / IPA ratio of 80/20 for 15 to 55 ° C. The 3D-MPL solution was then added to the DOPC solution and further mixed for another few minutes. The 3D-MPL / DOPC mixture was then added to the cholesterol solution and further mixed for another 1 h to provide a final composition with 120 mg / ml of DOPC (20/5/1, weight ratio of DOPC / cholesterol / 3D-MPL). 2. The 3D-MPL was suspended with 50% of the solvent (ethanol / IPA, 80/20) and then added to powders of DOPC and cholesterol. The volume was then adjusted with the rest of the solvent and the mixture was heated to 40 ° C for 15 minutes to provide a final composition with 120 mg / ml of DOPC (20/5/1, weight ratio of DOPC / cholesterol/ 3D-MPL). Method 1 required that the mixture be maintained at 55 ° C for 1 h for complete solubilization of the components. However, if it is not kept under gentle agitation for a few minutes, phase separation can be observed. To avoid this, continuous agitation is necessary until the solubilization is complete. 105 Method 2 allows complete solubilization after less time (15 min) and no separation of the phases can be observed without stirring. The microfluidic device with a single mixing chamber operated with a total flow rate of 18 ml / min, ratio of flow rates of 20 (1/19, organic / aqueous) and at temperatures of 15 to 25 ° C. using the first solution of stock prepared by both processes. The second (aqueous) solution was QS-21 in water for injection. Results The results are shown in Figure 5. conclusions Method 1 is more sensitive to temperature while Method 2 allows microfluidic operation less sensitive to temperature with liposome samples within a size specification of 95 to 120 nm over the range of 15 to 25 ° C . Example IC - Limits of concentrations of solutions The impact of the concentration of DOPC, cholesterol and 3D-MPL on the stability and solubility of stocks was evaluated. DOPC, cholesterol and 3D-MPL stock solutions in 80/20 ethanol / IPA were prepared at DOPC concentrations of 40, 60, 80, 100, 120, 140, 160 and 200 mg / ml (20/5/1, weight ratio of DOPC / cholesterol / 3D-MPL) by following method 2. The measurement was first made at T o (30 ° C), 106 the samples were then stored at 25 ° C while a hour , analyzed and then stored at 20 ° C while a hour , analyzed and then stored at 15 ° C while a hour and analyzed. Results The results are shown in Figure 6. The nephelometric measurement by Nephelostar revealed that the concentrations below 100 mg / ml evolve and present a higher turbidity. Similarly, 200 mg / ml evolves and has higher turbidity. conclusions Concentrations between 100 and 160 mg / ml are stable at temperatures between 15 ° C and 30 ° C. These surprising results could be explained by specific interactions between the lipid (DOPC), the sterol (cholesterol) and the TLR4 agonist (3D-MPL) when mixed in ethanol / IPA within this concentration range. Example 2 Study of the Impact of the Composition of the Solvent and of the Temperature on the Size of the Liposomes Process The stock solutions of DOPC, cholesterol and 3D-MPL were prepared at a concentration of DOPC of 120 mg / ml (20/5/1, weight ratio of DOPC / cholesterol / 3D-MPL) by following the process. 2 in of 107 ’ethanol / IPA at 80/20 ratios; 70/30 and 60/40 and used with an aqueous stock of QS-21 at 1.5 mg / ml. The microfluidic process was carried out at temperatures of 15 ° C, 20 ° C and 25 ° C at a total flow rate of 18 ml / min and a flow rate ratio of 5 (1/4, organic / aqueous). In this experiment, the sizes of the liposomes were measured before dialysis. Results The results are shown in Figure 7. conclusions The temperature partially directs the solubility of the components (DOPC, cholesterol and 3D-MPL). Stock prepared at 40 ° C can be cooled to 15 ° C without precipitation. However, operation at lower temperatures induces faster precipitation and thereby produces smaller liposomes. The size of the liposomes is impacted by the composition of the first solution at different temperatures with the greatest variation with a ratio of 60/40> 70/30> 80/20. This experience also confirms the choice of a report from 80/20 like exhibiting sensitivity the more weak to the temperature. Example 4 - Detailed analysis conditions of microfluidic operation and of their impact on the liposome size 108 Based on the general limits previously determined, a central compound experiment design (DOE) was constructed to determine the response of the process in terms of size (Zav) and to detect any cross interactions between temperature, total flow , the flow report and the stock concentration. Process Table 2 Summary of the conditions studied Setting Evaluation range (Upper and lower limits) Concentration of DOPC in the first solution 100 pg / ml 160 pg / ml Total flow 14 ml / min 20 ml / min Debits report 4 (1/3, organic / aqueous) 6 (1/5, organic / aqueous) Temperature 15 ° C 25 ° C A table of test conditions is provided in Figure 8 with a visual summary of the test conditions in Figure 9. The experiments were carried out over four days. The first solutions were prepared according to method 2 as described above. In short, DOPC (Lipoid) was weighed, followed by cholesterol (Sigma). In a separate bottle, 3D-MPL (GSK Hamilton) has been 109 weighed. 50% of the organic phase (80% ethanol (Merck) and 20% isopropanol) was added to the 3D-MPL. The suspended MPL was then added to the DOPC / cholesterol powder and placed at 40 ° C with mixing. After the solubilization of the three components (clear solution), the minutes at organic stock are left for a further ° C. The volume is then adjusted to provide the target concentration (20/5/1, weight ratio of DOPC / cholesterol / 3D-MPL). For the second solution, QS-21 concentrated bulk liquid was diluted in water for injection to achieve the required final concentration. Statistical analysis was performed using SAS 9.2 and Design Expert based on a central compound design with centered faces for the estimation of the response surface, with 6 central points and 24 model points. Reynolds numbers have been calculated using the equation: pU 2wh μ H ’+ h P 2Q μ w + h For example., Under the conditions: Organic phase Aqueous phase Density 0.829 g / cm 3 at 22.4 ° C 1.002 g / cm 3 at 21.7 ° C Viscosity 3.345 Cp at 19.8 ° C 1.09 Cp at 20 ° C Debit 3.2 ml / min 12.8 ml / min Working temperature 20 ° C 20 ° C 110 Based on the average densities and viscosities of liquids according to their proportions: Density: (12.8 x 1.002 + 3.2 x 0.829) / 16 - 0.9674 g / cm 3 Viscosity: (12.8 x 1.09 +3.2 x 3.345) / 16 = 1.541 Cp, if 1 Pa = 1 kg. irT 1 . s ~ 2 and 1 Pc = 1 mP a.s then the viscosity = 1.541 gm _1 .s _1 = 0.01541 g. wandered 1 , s 1 The mixing chamber dimensions are: 2000 µm (1) x 400 µm (h). 2Q = 2 x 16 = 32 ml / min = 0.53 cm 3 / s 1 + h = 400 pm (height) + 2000 pm (width) = 2400 pm = 0.24 cm All inside the equation: (0.9674 x 0.53) / (0.01541 x 0.24) = 138.6 An equivalent approach can be taken for all flow rates and flow ratios. Results Figure 10 provides the results of the experiment. - Modeling of the Pdl Table 3 presents the standard deviation (SD) and the coefficient of variance (CV) for Zav and Pdl. Table 3 DOE central point reproducibility analysis Zav Pdl Reproducibility of ET 1.17 0.01 111 Intermediate loyalty of the ET 5, 53 0.03 CV reproducibility 1.05% 4.24% CV intermediate loyalty four hundred ninety seven % 16.89% No reliable Pdl prediction model could to be adapted to the data s, except one correlation significant with the Zav (coefficient of correlation 0.75). A Zav <110 nM produces a Pdl <0.2 in 0, 95 of the cases. The relationship between the Zav and the Pdl is illustrated on Figure 11. - Zav modeling Some significant effects of the various factors studied were observed, and a clear co-effect of concentration and temperature as well as temperature and flow. R 2 adjusted = 0.80 R 2 predicted = 0.73 Other effects are considered not significant (p value> 5%) Table 4 P-value for the studied parameters and cross effect Postman P-value Concentration (A) 0.008308 Debit report (B) 0.000998 Temperature (C) <0, 0001 Flow (D) 0.957301 Concentration and temperature (AC) 0.008791 112 Temperature and flow (CD) 0.000151 Squared ratio (Β Λ 2) <0.0001 Figures 12 to 20 show the prediction of the response of Zav to different fixed factors using the model created. - Confirmation of the model As shown in Figure 21, eight series of confirmations were undertaken to test the capacity of the model (all series at 130 mg / ml of DOPC and a flow rate ratio of 5 (1/4, organic / aqueous). The results obtained are in good agreement with the model, with all the results being within the expected range of 90 to 110 nm. Table 5 Confirmation results Series Temperatureo( VS) Total flow (ml / min) Zav (Nm) Pdl predicts Measured1 23 15 105 101 0.188 2 23 17 108 104 0, 187 3 20 16 100 98 0.185 4 20 16 100 99 0.196 5 17 15 94 97 0.168 6 17 17 91 89 0.160 7 20 16 100 96 0.173 8 20 16 100 95 0.176 Conclusion 113 To optimize the process in terms of temperature, execution time and collection volume, a ratio of 1/4 (organic / aqueous) with a concentration of DOPC around 130 mg / ml, a flow rate between 14 and 17 ml / min and a temperature between 16 and 25 ° C seems best for obtaining liposomes in the region of 95 to 120 nm (i.e., around The specific parameters are 130 mg / ml of DOPC in ethanol / IPA at 80/20, a flow rate ratio of 1/4 (organic / aqueous), a total flow rate of ml / min and a temperature of 20 ° C. Table 3 presents the standard deviation (SD) and the CV for Zav and Pdl. It shows a very low CV representing good reproducibility. Example 5 - Adaptive immune responses Process - Preparation of the adjuvant The liposomes were prepared using 1'appareil to room of mixed unique described before. The sentence organic including the DOPC (120 mg / ml), cholesterol (30 mg / ml) and the 3D-MPL (6 mg / ml) in ethanol / 80/20 IPA was mixed with the aqueous phase containing QS-21 (1.5 mg / ml) in water for injection under full flow conditions of 18 ml / min and flow ratio of 5 (1/4, organic / aqueous). The organic phase was maintained at 20 ° C. The aqueous phase was maintained at ° C. 114 The solvent was removed by dialysis and the resulting concentrate diluted to provide the final preparation of the adjuvant. - Vaccination Female C57B16 mice aged 6 to 8 weeks (22 / group) received twice by injection with an interval of 14 days the gE antigen formulated with liposomes produced by microfluidics with 3D-MPL and QS-21. A control group of 5 mice received gE with buffer alone. The final adjuvant preparation was diluted and mixed with gE as necessary to provide the vaccine mixture. Two doses of adjuvant were evaluated (0.4 and 0.1 pg of both 3D-MPL and QS-21 per animal, corresponding to 1/125 and 1/500 of a conventional human dose (DH) 50 pg, respectively). Each animal received 5 µg of gE. The injection volume was 20 µl. Spleens and sera were removed and analyzed for T and B lymphocyte responses, respectively, 7 days after the second immunization (day 21). - ICS (Staining of intracellular cytokines) The spleens were collected in RPMI medium and dissociated using a Potter tissue mill (homogenizer) using two pushes up and down. The homogenized samples were transferred to 50 ml polypropylene tubes. The fibrous material was removed by filtration through a 100 µM nylon cell sieve. the 115 cells were then washed, counted and resuspended at 10 7 cells per ml. ICS is the technology that allows the quantification of antigen-specific T cells based on the production of cytokines. Lymphoid cells are restimulated overnight. in vitro with the gE peptides or the medium in the presence of a protein transport inhibitor (brefeldine A). These cells are then treated by a conventional procedure using extracellular antibodies: CD4, fluorescent immunofluorescence (staining CD8; intracellular staining TNF-alpha, IFN-gamma and IL2). The results are expressed in frequency of cells positive for cytokines within a population of CD4 cells after substraction of the condition of middle for each mouse. analysis statistic a been carried out on the population who has showed the expression of at least two cytokines (IL2, IFN-alpha or TNF-alpha). - ELISAIgG total anti-gE have summer measured by a ELISA test. Plates of 96 wells have summer sensitized with the antigen during one night at 4 ° C. The plates were then washed and saturated with saturation buffer for 1 hour at 37 ° C. Then, 100 μΐ of diluted mouse serum or standard or control were added and incubated for 1 h 30 at ° C. After washing, the plates were incubated for 1 hour at 37 ° C with 100 μΐ of biotinylated anti-mouse IgG. After washing, the plates were incubated 116 for 30 min at 37 ° C with 100 μΐ of streptavidin-POD conjugate. After washing, 100 µl of TMB per well was added and the plates were stored in the dark at room temperature for 15 minutes. To stop the reaction, 100 μΐ of 0.4 N H2SO4 were added per well. The absorbance was read at a wavelength of 450/630 nm by an ELISA plate reader. The results were calculated using softmax-pro software. Results The liposomes had a diameter of 95.5 nm, a PdL of 0.184. The final adjuvant preparation had a DOPC content of 2.2 mg / ml, a cholesterol content of 0.58 mg / ml, a QS-21 content of 119 pg / ml and a 3D-MPL content. 90 pg / ml. The ELISA results are shown in Figure 22 and the ICS results in Figure 23. conclusions The liposomes produced by microfluidics together with the TLR4 agonist and saponin were able to enhance the cellular and antibody responses against an example of antigen. Example 6 __________ Stability _____ of_____ microfluidic liposomes The liposomes were prepared using the single mixing chamber apparatus described above. The organic phase including DOPC (130 mg / ml), cholesterol (32.5 mg / ml) and 3D-MPL 117 (6.5 mg / ml) in 80/20 ethanol / IPA was mixed with the aqueous phase containing QS-21 (1.625 mg / ml) in water for injection under flow conditions total of 16 ml / min and flow rate ratio of 5 (1/4, organic / aqueous). The organic phase was maintained at 20 ° C. The aqueous phase was maintained at 20 ° C. The solvent was removed by dialysis and the material was sterilized by filtration. The results are shown in Figure 24, indicating that the liposomes produced using microfluidics are substantially unchanged after storage for 2 months at 4 ° C. EXAMPLE 7 Increase in the Scale of the Process The objective of this example was to test the increase in the scale of the process in order to produce batches of adjuvant on a commercial scale using a microfluidic process. A single batch of 320 l of adjuvant was prepared in a production cycle suitable for one working day (Figure 25). The number of mixing chambers used was 16. Tubing assessment In order to distribute the organic and aqueous phases in the 16 mixing chambers, the distribution tubing should ideally provide a uniform flow distribution. Practical limitations mean that perfect distribution is not possible, but that excessive variation should be avoided. A Excessive variation in the flow can lead to inhomogeneity in the final product and potentially to a product which is outside the target specification. A theoretical estimate of the content of components in the final product was calculated depending on variations in the flow distribution. Table 6 shows that variations of less than 5% do not significantly affect the content of components in the final product which remains within the target specification range. Above 6% variation, the 3D-MPL content is close to the upper acceptable limit and exceeds this limit with further increases in the percentage of variation. Table 6 Component content expected in the final product with variation in flow ml / min ml / minml / min Final quantity (mg / ml) % var % (Aq-Org) Aq org Report Total flow DOPC Chol 3D-MPL QS21 0 100-100 12, 8 3.2 5.00 16 2.0 0.50 0.100 0.100 2 102-98 13, 06 3, 14 5, 16 16.2 1, 9 0.48 0.097 0,101 2 98-102 12.54 3.2 6 4.84 15, 8 2.1 0.52 0,103 0.099 5 105-95 13, 44 3, 04 5, 42 16.5 1.8 0.46 0.092 0, 102 5 95-105 12.16 3, 36 4, 62 15.5 2, 2 0.54 0.18 0.098 6 106-94 13, 57 3, 01 5, 51 16, 6 1.8 0.45 0.091 0, 102 6 94-106 12,03 3, 39 4.55 15, 4 2.2 0, 55 0.110 0.098 8 108-92 13, 82 2, 94 5, 70 16, 8 1.8 0.44 0.088 0, 103 8 92-108 11, 78 3.46 4.41 15.2 2.3 0.57 0.13 0.097 10 110-90 14.08 2.88 5, 89 17.0 1.7 0.42 0,085 0.104 10 90-110 11.52 3, 52 4.27 15.0 2, 3 0, 59 0.117 0.096 Procedure applied for the analysis of the tubing 119 The inlet of an initial tubing design (Figure 26) was connected to ISCO pumps filled with water. At each of the 16 exits from the tubing, a 20 cm ETFE (ethylene tetrafluoroethylene) pipe (1.6 mm, 1 mm ID) was connected and placed in a 50 ml Falcon ™ tube (Figure 27). Each Falcon ™ tube was weighed before the experiment. The hoses (inlet and outlet) and tubing were then filled with water to remove air bubbles. The system has operated for 2 min at a total flow rate of -200 ml / min. After the 2 min period, each Falcon ™ tube was weighed to calculate the exact mass of water delivered. The% calculated: weight of the average X channel measured. The initial tubing with water and also ethanol and isopropanol. flow variation was (1 to 16) divided by weight was tested at 200 ml / min to 50 ml / min with a mixture The results (Figure 28 and Figure 29) show the variation among the channels: at a high total flow (-200 ml / min), one was above the acceptable variation limit of 5% (channel 1) but d others were close to this limit (channels 2, 3, 4, 9, 10, 11 and 12). For the lowest flow rate (-50 ml / min), the variation is below% but shows the same general trend. Following these results, analysis by MFN (digital fluid mechanics) was carried out to predict the distribution of flows along each segment of the tubing. Analysis has shown that the initial bend induced differences in flow in the subsequent branches. These differences remained 120 along the final branches. The predicted variation was plotted against the observed experimental values (according to the analysis at 200 ml / min) and shows the same general trends (Figure 30). The experimental discovery for the first tubing that it was not able to distribute with desired homogeneity was confirmed with analysis by MFN. Based on these results, the MFN was used as a tool to support the design of an improved second generation tubing (Figure 31). Studies have led to the elimination of the initial elbow, the shortening of the length of the first channel, the increase in the lengths of the second and third channels. Under these circumstances, the speed profile is more constant and the traffic areas have been almost completely eliminated. Improved tubing test The second tube was tested with a similar procedure (12.8 ml / min / channel = 204.8 of total flow) and reproduced three times on each of the three tubes (designated by B1, B2 and A2). Figures 32 to 34 show the experimental results obtained. In all cases, the variation of the individual channels was less than the desired limit of 5%, in many cases, the variation of the measured flow was less than 2% of the average. Liposome production 121 Using the improved tubing, an experiment was carried out to confirm that liposomes with the same profile as those produced with a single mixing chamber could be produced on a commercial scale. For the experiments, ISCO 1000D and 500D pumps were used in tandem. Only one cylinder from each pump was used, due to the limited run time. The heating jacket of the pumps was connected to a water bath (Julabo F33), a water bath by pump. The temperature control was monitored using certified probes. Two improved tubing was connected to each pump at the inlet and to 2 microchips containing 8 mixing chambers each to provide a total of 16 mixing chambers in parallel. The pipe used was ETFE (1.6 mm, 1 mm ID), with a length of 29.5 cm. A 29.5 cm (1.6 mm, 0.5 mm ID) ETFE pipe was connected to the outlet of each mixing chamber. The temperature of the prototype was not directly controlled, but the device was placed in a temperature-controlled zone (at 20 ° C) and allowed to acclimatize before any experiment. The pumps were rinsed twice and emptied before filling with the organic phases (DOPC at 130 mg / ml, cholesterol at 32.5 mg / ml solubilized in ethanol / isopropanol at 80/20) and aqueous (water for injection) appropriate. The pumps were then primed to remove air from the system before being connected to the distribution pipes. The flow rates were set at 51.2 ml / min for the phase 122 organic and 204.8 ml / min for the aqueous phase. When the system was primed and cleared of air, the first 2 ml from each chamber were discarded and the output from each chamber was then collected in 16 separate containers (run time <30 seconds). 500 μΐ of product from each channel were grouped and subjected to a measurement by DLS after 130x dilution in PBS at pH 6.1. After running the series on the multi-chamber prototype, the hose from the pumps was disconnected from the multi-chamber prototype and connected to a single mixing chamber. The flow rates were adapted to 3.2 ml / min for the organic phase and 12.8 ml / min for the aqueous phase. When the system has been stabilized (absence of air), the first 2 ml leaving the mixing chamber were discarded and the following ~ 2 ml collected. The size measurements were calculated using Malvern ZS Nano series instruments (Figure 35) and show the same trend for the group of 16 mixing chambers compared to liposomes produced with the "single" mixing chamber. The sizes and the polydispersity are also equivalent (table 7). The results show for the first time an equivalence for the production of liposomes (DOPCCholesterol) between a prototype containing 16 mixing chambers (and associated tubing) suitable for use in production on a commercial scale and the single mixing chamber. 123 Table 7 DLS results from multiple chamber and single chamber liposomes Zav (nm) Pdl Unique mixing chamber 141 0.22 Group of 16 mixing chambers 143 0.23 Example 8 Adaptive Immune Responses in Relation to Liposomes Produced by Thin Film Processes Process - Preparation of the adjuvant Three batches of liposomes were prepared using the single mixing chamber apparatus described above. The organic phase comprising DOPC (130 mg / ml), cholesterol (32.5 mg / ml) and 3D-MPL (6.5 mg / ml) in ethanol / IPA at 80/20 was mixed with the aqueous phase containing QS-21 (1.625 mg / ml) in water for injection under conditions of total flow rate of 16 ml / min and ratio of flow rates of 5 (1/4, organic / aqueous). The temperature was maintained at 20 ° C. The solvent was removed by diafiltration using a 30 kDa Hydrosart membrane and six volumes of replacement buffer. The diafiltration time was approximately 40 minutes. The material was then sterilized by filtration using filtration sterilization on a 0.22 µm PES membrane. 124 The resulting liposomal adjuvant concentrate can be diluted as necessary to provide the final adjuvant preparations. - Vaccination 6 to 8 week old female C57B16 mice (6 mice per group, total of 186 animals) were twice injected with a 14-day interval with the gE antigen in a liposomal formulation with 3D-MPL and QS -21. A negative control group received gE with buffer alone on the same schedule. The final vaccination mixture was prepared by diluting the adjuvant concentrate and mixing with gE as required. Five, adjuvant doses were evaluated (0.05, 0.1, 0.2, 0.4 and 1 pg of both 3DMPL and QS-21 per animal per injection, corresponding to 1/1000, 1 / 500, 1/250, 1/125 and 1/50 of a conventional human dose (DH) of 50 pg, respectively based on the expected content of immunostimulant). Each animal received 5 µg of gE per injection. The injection volume was 20 µl. Three batches of microfluidic liposomal adjuvant were compared to three batches of liposomal adjuvant produced by thin film methods. Due to space restrictions, the experiment was undertaken in two parts (i.e., 3 mice from each group of 6 received treatment in each part, with the results combined). The rats and the sera were taken analyzes for the responses from T cells and 125 respectively, 7 days after the second immunization (day 21). The ICS and ELISA techniques were undertaken using the methods provided in Example 5. Results Characterization of microfluidic batches Table 8 DLS characterization of microfluidic batches Lot After microfluidic mixing Afterdiafiltrationandsterilization After storage(4 ° C) Zav(Nm) Pdl Zav(Nm) Pdl Zav(Nm) Pdl Time 1 98 0.18 96 0.19 98 0.19 4.5 months 2 100 0.21 100 0.22 99 0.21 4 months 3 103 0.22 102 0.24 102 0.24 4 months Table 9 Composition of microfluidic batches (after dilution of the concentrate) Lot DOPC(Mg / ml) Cholesterol(Mg / ml) QS21(Pg / ml)(100 expected) 3D-MPL(Pg / ml)(100expected) Residual alcohol (pg / dose of500 pl) 1 1, 9 0.46 91 74 55 2 2 0, 51 96 82 125 3 2 0.48 96 79 85 126 Table 10 Characterization of batches by thin film Lot DOPC(Mg / ml) Cholesterol(Mg / ml) QS21(Pg / ml)(100 expected) 3D-MPL(Pg / ml)(100expected) Residual alcohol (pg / dose of -500 pl) Zav(Nm) Pdl 1 2 0.49 99 84 Nodone 104 0.14 2 1.9 0.49 94 88 Nodone 108 0.14 3 2 0.48 101 84 Nodone 105 0.13 The ELISA presented on figure ICS on then recover (Lot 1, (Experience 1 against linear then just on separately for the two processes. The associated ratios of the predicted geometric means between the methods are shown in Figure 38 and Figure 39 (ELISA and ICS, respectively). conclusions The liposomes produced by microfluidics together with a TLR4 agonist and a saponin were able to enhance the cellular and antibody responses to an exemplary antigen in a way 127 generally comparable to liposomes produced by thin film. Example 9 Adaptive Immune Responses in Relation to Liposomes Produced by Thin Film Processes Following the successful scaling up described in Example 7, the 16-channel microfluidic apparatus was used to prepare a batch of liposomal material comprising the immunostimulants of saponin (QS21) and TLR4 (3D-MPL). The organic phase comprising DOPC (130 mg / ml), cholesterol (32.5 mg / ml) and 3D-MPL (6.5 mg / ml) in ethanol / IPA at 80/20 was mixed with the aqueous phase containing QS-21 (1.625 mg / ml) in water for injection under conditions of total flow rate of 16 ml / min (per chamber) and ratio of flow rates of 5 (1/4, organic /aqueous). The temperature was maintained at 20 ° C. An initial reference series with a single chamber was conducted using a single chamber (first 2 ml discarded). Then the system worked with the 16 chambers in parallel and the material from each chamber was collected individually (first 2 ml from each chamber discarded). A group of 16 rooms has been prepared. A third series was carried out using the same single mixing chamber as that used for the reference conditions (not placed in the incubator), again the first 2 ml discarded. 128 Size measurements were undertaken by DLS after the microfluidic process (no removal of solvent). Table 11 DLS results from multiple chamber and single chamber liposomes Zav (nm) Pdl Unique mixing chamber reference 105 0.21 Group of 16 mixing chambers 103 0, 20 Unique mixing chamber 106 0.21 The comparison of the sizes obtained over the three series (FIG. 40) shows comparable profiles, confirming the robust and modular nature of the approaches presented in the present application. Throughout the specification and the claims which follow, unless the context requires otherwise, the term "understand", and variations such as "includes" and "comprising", will be understood to imply the inclusion of an integer, of a step, a group of whole numbers or a group of steps indicated but not the exclusion of any other whole number, step, group of whole numbers or group of steps. The application of which this description and the claims form a part may be used as the basis of priority over any subsequent application. The claims of such subsequent application may be directed to any characteristic or combination of characteristics 129 described here. The embodiments are intended to be independently fully combinable with each other when appropriate to the circumstances to form other embodiments of the invention. They may take the form of product, composition, process, or use claims and may include, by way of example and without limitation, the following claims. Of course, The invention is not limited to the exemplary embodiments described and shown above, from which other modes and other embodiments can be provided, without Bibliography Arias MA et al. (2012) Glucopyranosyl Lipid Adjuvant (GLA), a Synthetic TLR4 Agonist, Promotes Potent Systemic and Mucosal Responses to Intranasal Immunization with HIVgpl40. PLoS ONE 7 (7): e41144. doi: 10.13 71 / journal.pone.0041144 Black, S., E. De Gregorio, and R. Rappuoli. 2015. Developing vaccines for an aging population. 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权利要求:
Claims (9) [1" id="c-fr-0001] 1. Method for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. [2" id="c-fr-0002] 2. Method for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps ·. (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. [3" id="c-fr-0003] 3. Method for manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (at) mixing in the device of a first solution comprising a solvent, a lipid phosphated idylcholine and a sterol, and d 'a second solution comprising of Water;(B) 1'élimination solvent; and(vs) adding the saponin.4. Method of manufacturing ' a concentrated liposomal for use in the preparation of a liposomal adjuvant comprising a saponin using a 135 microfluidic device, comprising the step of mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water and saponin. [4" id="c-fr-0004] 5. Method for manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device, comprising the following steps: (a) the mixing in the device of a first solution including a solvent, a lipid from phosphatidylcholine and a sterol, and a second solution comprising of Water; and (b) adding the saponin. 6. Method of manufacturing a concentrate liposomal for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, a phosphatidylcholine lipid and a sterol, and a second solution comprising water; (b) 1 adding saponin; and(c) 1 adding the TLR4 agonist;in which steps (b) and (c) can to be in one order or in the other, or can to be done in one step. [5" id="c-fr-0005] 7. Method for manufacturing a liposomal adjuvant comprising a saponin using a device 136 microfluidics according to claim 1, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water and saponin; and (b) removing the solvent. [6" id="c-fr-0006] 8. A method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device according to claim 2, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) removing the solvent. [7" id="c-fr-0007] 9. A method of manufacturing a liposomal adjuvant comprising a saponin using a microfluidic device according to claim 3, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (B) 1'élimination solvent; and (vs) adding the saponin. 10. Method of making a concentrated liposomal for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device according to claim 4, comprising the step of mixing in the device a first solution comprising a solvent, DOPC and 137 a sterol, and a second solution comprising 1 1 water and saponin. 11. A method of manufacturing a liposomal concentrate for use in the preparation of a liposomal adjuvant comprising a saponin using a microfluidic device according to claim 5, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; and (B) I'm out of the saponin. 12. Process of manufacturing of a concentrated liposomal for use in the preparation of a liposomal adjuvant comprising a saponin and a TLR4 agonist using a microfluidic device according to claim 6, comprising the following steps: (a) mixing in the device a first solution comprising a solvent, DOPC and a sterol, and a second solution comprising water; (b) adding the saponin; and (c) adding the TLR4 agonist; in which steps (b) and (c) may be in order or in The other, or can be done in one step. 13. Process according to 1 any one of claims 1 to 5 or 7 to 11, wherein the first solution further comprises a TLR4 agonist. 14. Method according to any one of claims 1 to 3 or 7 to 9, in which a TLR4 agonist is added before the removal of the solvent. 138 15. A method according to any one of claims 1 to 3 or 7 to 9, wherein a TLR4 agonist is added after removal of the solvent. 16. Method according to any one of claims 1 to 15, in which the microfluidic device has an inlet for the first solution to the mixing chamber. 17. Method according to any one of claims 1 to 16, in which the microfluidic device has two inlets for the second solution to the mixing chamber. 18. Process according to 1 any one of the microfluidic claims to 17, wherein the device of the inputs such that each input has 0.2 mm and covers the full length of the microfluidic mixing chamber. 19. Process according to 1 any one of claims 1 to 18, wherein the device has a microfluidic chamber with a cross section of the chamber 0.6 to 1.2 mm 2 , as around the mixture having the mixture is mm. 0.8 20. Claims 1 microfluidic a rectangular process 21. Process according to one any of the at 19, in which device a room of mixed sensibly chopped off cross.according to claim 20, in which the cross section of the mixing chamber has a long side from 1.6 to 2.4 mm. 139 22. The method of claim 20 or 21, wherein the cross section of the short side microfluidic claims mixing chamber has a chamber of any mixing device having claims length 1 any microfluidic device has a mixing chamber for mixing. material recovery 25. Method according to one of claims 1 to 24, wherein the microfluidic device has a mixing chamber such that the total flow rate in the mixing chamber is 17.5 to 25 ml / min / mm 2 of cross section of the mixing chamber. 26. Process according to 11 a any of the claims 1 to 25, in which The report of the debits for the first and second solutions in the microfluidic device lies in the range of 1/2 at 1/6. 27. Process according to 1 ' a any of the claims 1 to 26, in which the flow of the first solution in the bedroom mixture of microfluidic device East from 3 to 6.5 ml / min / mm 2of cross section of, mixing chamber. 28. Process according to 1 ' a any of the claims 1 to 27, in which the flow of the second solution in the bedroom mixture of 140 microfluidic device is 14 to 20 ml / min / mm 2 of cross section of the mixing chamber. 29. Method according to any one of claims 1 to 28, in which the temperature of the first solution is supplied at a temperature of 15 to 25 ° C. 30. The method according to any one of claims 1 to 29, in which the temperature of the second solution is supplied at a temperature of 15 to 25 ° C. 31. Method according to one of claims 1 to 30, wherein the temperature of the mixing chamber of the microfluidic device is 15 to 25 ° C. 32. The method according to any one of claims 1 to 31, in which the maximum Reynolds number within the mixing chamber of the microfluidic device is 1500 or less. 33. Method according to any one of claims 1 to 32, in which the microfluidic device comprises a plurality of mixing chambers. 34. The method of claim 33, wherein the device comprises 4 to 32 mixing chambers. 35. The method of claim 33 or 34, wherein all of the mixing chambers in the plurality of mixing chambers are fed by the same pumps and the mixed material from all of the mixing chambers is collected before further treatment and / or storage. 36. Method according to any one of claims 33 to 35, in which the plurality of 141 mixing chambers is capable of producing the mixed material at a rate of 50 to 2000 ml / min. 37. The method according to any one of claims 33 to 36, in which the plurality of mixing chambers is capable of producing the mixed material at a rate of at least 1 g of phosphatidylcholine lipid per minute. 38. The method of any of claims 33 to 37, wherein the plurality of mixing chambers is capable of producing the material mixed at a rate of at least 1 g of DOPC per minute. of the 39. Process according to 1 'a any claims 1 to 38, in which the solvent includes 70 to 90% v / v ethanol. 40. Process according to 1 1 a any of the claims 1 to 39, in which the solvent includes [8" id="c-fr-0008] 10 to 30% v / v isopropanol. 41. Process according to 1 'a any of the claims 1 to 40, in which the first solution includes 100 to 170 mg / ml lipid of idylcholine phosphate 42. Process according to 1 'a any of the claims 1 to 41, in which the first solution includes 100 to 170 mg / ml of DOPC.43. Process according to 1 'a any of the 44. Process according to 1 'a any of the claims 1 at 43, in which the sterol is the Claims 1 to 42, wherein the first solution comprises 20 to 50 mg / ml sterol. cholesterol. 142 45. Method according to claims 1 to 44, in the first solution is 120 46. A method according to claims 1 to 45, in any one of which the dry weight of 250 mg / ml. any one of which the second solution comprises at least 90% w / w of water. 47. Process according to 1 any one of claims 1 46, in which saponin is the QS-21. 48. Process according to 1 any one of claims 1 47, wherein the second solution comprises 1 to 4 mg / ml of saponin. 49. The method according to claims 6, 12 to 48, is 3D-MPL. any of which the TLR4 agonist 50. Method according to claim 6, [9" id="c-fr-0009] 12 to 49, wherein the first solution comprises 4 to 10 mg / ml of the TLR4 agonist. 51. Method according to any one of claims 1 to 50, in which the average size of the liposomes is from 95 to 120 nm. 52. The method of any of claims 1 to 51, wherein the polydispersion of the liposomes is 0.3 or less. 53. The method of claim 52, wherein the polydispersion of the liposomes is 0.2 or less. 54. Method according to any one of claims 1 to 53, in which the solvent is removed by diafiltration, ultrafiltration and / or dialysis, in particular by diafiltration. 143 55. Process according to 1 ' a any of the claims 1 at 54, in which Elimination of solvent results in a content in water '' at least 98 % p / p of water. 56. Process according to 1 ' a any of the claims 1 at 55,including 1 'step additional dilution, until a concentration desired finish. 57. Process according to 1 ' a any of the claims 1 at 56,including 1 'step additional adjustment pH between 5 to 9. 58. Process according to 1 ' a any of the Claims 1 to 57, comprising the additional step of adjusting the osmolality to 250 to 750 mOsm / kg. 59. A method for preparing an adjuvanted immunogenic composition comprising an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen, said method comprising the steps of: (i) manufacture of a liposomal adjuvant according to the method of any one of claims 1 to 58; (ii) mixing the liposomal adjuvant with an immunogen or an antigen, or a polynucleotide encoding the immunogen or the antigen. 60. A method of manufacturing an adjuvanted immunogenic composition, said method comprising the step of combining an immunogen or an antigen, or a polynucleotide coding for the immunogen or the antigen, with a manufactured liposomal adjuvant according to the method of any of claims 1 to 58. 144 61. Method according to any one of claims 1 to 60, comprising the additional step of sterilization by filtration. 62. Liposomal adjuvant comprising a saponin, a TLR4 agonist, DOPC and a sterol, produced according to the method of any one of claims 1 to 61. 63. Adjuvanted immunogenic composition produced according to the method of claim 60 or 61. 64. Solution comprising a solvent and 100 to 170 mg / ml of lipid, in which the solvent comprises 70 to 90% v / v of ethanol, such as 75 to 85% v / v of ethanol, and 10 to 30% v / v isopropyl alcohol such as 15 to 25% v / v isopropanol. 65. Solution according to claim 64, in which lipid is a phosphatidylcholine. 66. Solution according to claim 64, in which lipid is here DOPC. 67. Solution according to 1 1 any 1of the claims 64 at 66, further comprising 20 to 50 mg / ml of sterol. 68. Solution according to One or the other of the claim 64 or 67, wherein the sterol is cholesterol. 69. Solution according to 1 'a any of the claims 64 to 68, comprising an agonist of the TLR4. 70. Solution according to claim 69, in which 1'agoniste of the TLR4 is present at a concentration from 4 to 12 mg / ml. 145 71. A solution according to any one of claims 64 to 70 which consists essentially of a solvent and from 100 to 160 mg / ml of DOPC and from 30 to 40 mg / ml of cholesterol, from 4 to 10 mg / ml of TLR4 agonist, and in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v of isopropyl alcohol. 72. A solution according to any one of claims 64 to 71 in which the TLR4 agonist is a lipopolysaccharide, such as 3D-MPL. 73. Process for preparing a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps of: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with the phosphatidylcholine lipid and cholesterol; (iii) adding solvent; (iv) the mixture. 74. Method for preparing a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said method comprising the steps of: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with the phosphatidylcholine lipid and cholesterol; (iii) adding solvent; (Iv) mixing; (v) adding additional solvent. 75. Process for preparing a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps of: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with DOPC and cholesterol; (iii) adding solvent; (iv) the mixture. 76. Process for the preparation of a solution comprising a solvent, a lipid, cholesterol and a TLR4 agonist, said process comprising the steps of: (i) preparing a suspension of the TLR4 agonist in at least part of the solvent; (ii) the combination of the suspended TLR4 agonist with DOPC and cholesterol; (iii) adding solvent; (iv) the mixture; (v) adding additional solvent. 73 to 77. Process according to 1 1 a demands 7 6, in which mixture is at a temperature of 30 at 50 ° C. 78. Process according to 1 ' any of the claims 73 at 77, in which at least a part of the solvent is at least 35% of the solvent. 79. Process according to 1 ' any of the claims 73 at 78, in which at least a 147 part of the solvent is 90% solvent or less as 80% or less, specially 70% or less and in particular 60% or less. 80. Process according to 1 'a any of the claims 73 at 79, in . which the solvent added is the solvent res As. 81. Process according to 1 'a any of the claims 73 to 80, wherein the solution comprises 100 to 160 mg / ml of lipid and 30 to 40 mg / ml of cholesterol, in which the solvent comprises 70 to 90% v / v of ethanol and 10 to 30% v / v isopropyl alcohol, in which the lipid is suitably DOPC. 82. Method according to any one of claims 73 to 81, in which the agonist of TLR4 is 3D-MPL. 83. The method of claim 82, wherein the solution comprises 4 to 10 mg / ml 3D-MPL. 84. The method according to any one of claims 73 to 83, wherein the additional solvent is 0 to 30% of the solvent. 85. Solution containing liposomes obtainable by mixing the first solution and the second solution according to the methods of any one of claims 1 to 53 before removing the solvent. 86. Method, adjuvant, composition or solution according to any one of claims 1 to 85, in which the phosphatidylcholine lipid contains saturated unbranched acyl chains containing 12 to 20 carbon atoms such as acyl chains comprising 14 to 18 carbon atoms. 148 87. Method, adjuvant, composition or solution according to any one of claims 1 to 85, in which the phosphatidylcholine lipid contains unbranched acyl chains containing 12 5 to 20 carbon atoms and a double bond, such as chains acyl having 14 to 18 carbon atoms and a double bond. 88. Method, adjuvant, composition or solution according to any one of claims 1 to 85, in which / which the phosphatidylcholine lipid is chosen from dilauroyl-phosphatidylcholine (DLPC), dimyristoyl-phosphatidylcholine dipalmitoyl-phosphatidylcholine pho sphat idy1choline phosphatidylcholine phosphatidcholine (DMPC), (DPPC), (DSPC) (DAPC), and la la distearoyldiarachidoyldipalmitoleoylet dioleoylphosphatidylcholine (DOPC); and their mixtures.
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同族专利:
公开号 | 公开日 JP2020522485A|2020-07-30| CA3063954A1|2018-12-06| US20210162042A1|2021-06-03| IE20190084A1|2019-12-25| DE112018002827T5|2020-04-09| BR112019025193A2|2020-06-23| GB201918592D0|2020-01-29| WO2018219521A1|2018-12-06| IE20190085A1|2019-12-25| EP3630176A1|2020-04-08| MX2019014319A|2020-02-07| CN111032080A|2020-04-17| GB2577232A|2020-03-18| IE20180065A1|2019-12-25| IE20190086A1|2019-12-25| GB2577232A8|2021-12-29|
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申请号 | 申请日 | 专利标题 US201762512352P| true| 2017-05-30|2017-05-30| US62512352|2017-05-30| 相关专利
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