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EP0000772A1 - Réactif immunologique sous forme de particules spéciales de latex à base de polymère de vinyle, revêtues d'un matériau immunologiquement actif. Procédé pour sa fabrication. Utilisation de ce réactif. Procédé d'analyse utilisant ce réactif et trousse à réactifs le contenant - Google Patents

Réactif immunologique sous forme de particules spéciales de latex à base de polymère de vinyle, revêtues d'un matériau immunologiquement actif. Procédé pour sa fabrication. Utilisation de ce réactif. Procédé d'analyse utilisant ce réactif et trousse à réactifs le contenant Download PDF

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Publication number
EP0000772A1
EP0000772A1 EP78100581A EP78100581A EP0000772A1 EP 0000772 A1 EP0000772 A1 EP 0000772A1 EP 78100581 A EP78100581 A EP 78100581A EP 78100581 A EP78100581 A EP 78100581A EP 0000772 A1 EP0000772 A1 EP 0000772A1
Authority
EP
European Patent Office
Prior art keywords
reagent
latex
particles
immunologically active
sample
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP78100581A
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German (de)
English (en)
Inventor
Roncari Dr. Gaetano
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
HOFFMANN LA ROCHE
F Hoffmann La Roche AG
Original Assignee
HOFFMANN LA ROCHE
F Hoffmann La Roche AG
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by HOFFMANN LA ROCHE, F Hoffmann La Roche AG filed Critical HOFFMANN LA ROCHE
Publication of EP0000772A1 publication Critical patent/EP0000772A1/fr
Withdrawn legal-status Critical Current

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S436/00—Chemistry: analytical and immunological testing
    • Y10S436/823—Immunogenic carrier or carrier per se

Definitions

  • the present invention relates to valuable diagnostic reagents, a process for their preparation and diagnostic methods in which such reagents are used.
  • An antigen is a foreign substance which, when applied to the living being, causes the formation of certain soluble substances, known as antibodies. Any substance such as a protein which is not normally present in a particular living being can cause the formation of antibodies if it is applied to the living being under suitable conditions.
  • the antibodies After their formation, the antibodies react with the antigens and in this way protect against infections in the case of a bacterial or virus foreign body.
  • Immunological test methods are based on the antigen-antibody reaction, which usually manifests itself through insolubility or agglutination.
  • an antigen or an antibody is confirmed or determined by adding the corresponding antibody or antigen to a body fluid of the living being, usually urine, blood serum or a specially treated blood extract.
  • a body fluid of the living being usually urine, blood serum or a specially treated blood extract.
  • other body fluids can also be used.
  • the presence or absence of the antibody or the antigen in the body fluid of the living being is determined by determining the occurrence or non-occurrence of an antigen-antibody reaction.
  • the antibody or antigen was' by means of a carbodiimide via an amide bond to discrete particles of carboxylated latex polymers, such as e.g. of carboxylated copolymers of butadiene and styrene.
  • the present invention now relates to novel polymeric carriers with which the above disadvantages can be avoided and which with a wide range of immunologically active Materials can form a diagnostically usable reagent that is stable, specific, and sensitive, and that enables an easily detectable visual assessment in a very short time.
  • the present invention relates to a water-insoluble reagent for immunological determination with a specific weight approximately equal to that of water in the form of discrete latex particles to which an immunologically active material is bound, characterized in that the latex consists of a dispersion of particles of vinyl polymers which as end groups are groups of the formula: wear, the particles of a core of vinyl and / or diene polymer which carries carboxyl and / or sulfonate functions, and of an outer layer of vinyl polymer which, as end groups, groups of the formula: carries, are formed and have an average diameter between 0.03 and 5 pm.
  • the invention further relates to a method for producing such a reagent, which is characterized in that the latex is reacted with the immunologically active material after diazotization or in the presence of suitable bifunctional reagents.
  • Immunologically active substances include amines, amino acids, peptides, proteins, lipoproteins, glycoproteides, sterols, steroids, lipoids, nucleic acids, enzymes, hormones, vitamins, polysaccharides and alkaloids.
  • Preferred Immunologically active substances are listed in the following table:
  • immunologically active substances are albumin, rheumatoid factor, human immunoglobulin IgG and antibodies against IgG.
  • Vinyl polymers which form the core of the particles are understood to mean homopolymers of monomers, such as styrene and its derivatives: methylstyrenes, ethylstyrenes, vinyltoluene; Vinyl chloride, vinylidene chloride; Vinyl acetate; Acrylic derivatives, such as alkyl acrylates and methacrylates (alkyl having 1 to 10 carbon atoms) which are optionally hydroxylated, such as 2-hydroxyethyl acrylate and methacrylate and 2-hydroxypropyl acrylate and methacrylate; Acrylonitrile and methacrylonitrile; and copolymers of these monomers with one another and / or with modifying vinyl comonomers such as divinylbenzene, acrylamide and methacrylamide and their N-substituted derivatives, such as e.g. Methylolacrylamide; these comonomers represent up to 5% by weight of the copolymer.
  • monomers
  • Diene polymers that form the core are homopolymers of butadiene and its derivatives: chloroprene, isoprene; and the copolymers of these monomers with one another and / or with vinyl monomers, as mentioned above, in all proportions and / or with modifying vinyl monomers, as enumerated above, the amount of which in the copolymer makes up to 5% by weight.
  • the vinyl polymers which form the outer layer of the particles are homopolymers of monomers, such as styrene and its derivatives, for example methylstyrenes, ethylstyrenes and vinyl toluene; optionally hydroxylated alkyl acrylates and alkyl methacrylates (alkyl with 1 to 10 carbon atoms); Acrylonitrile and methacrylonitrile; and copolymers of these monomers with one another and / or with modifying vinyl comonomers, such as divinylbenzene, acrylamide and methacrylamide, and their N-substituted derivatives, such as methylolacrylamide, which can make up up to 5% by weight of the copolymer.
  • monomers such as styrene and its derivatives, for example methylstyrenes, ethylstyrenes and vinyl toluene
  • the core polymer in the particles constitutes 30 to 99.5% by weight, preferably 60 to 99% by weight, and the polymer in the outer layer 70 to 0.5% by weight, preferably 40 to 1% by weight, represents.
  • the polymer particles the particle size distribution of which may be wide or narrow depending on the desired properties of the latex and the applications under consideration, have an average diameter between 0.03 and 5 ⁇ m, preferably between 0.05 and 1 ⁇ m. They represent up to 60% by weight, preferably up to 45% by weight, of the latex. However, the latex can easily be diluted or concentrated.
  • the core polymer can be prepared by emulsion polymerization of the vinyl monomer (s) and / or diene monomer (s) in the presence of at least one ethylenic mono- or polycarboxylic acid which is copolymerizable with the monomer (s) and / or at least one copolymerizable unsaturated alkali metal organosulfonate; then the polymer of the outer layer is prepared by emulsion polymerizing the vinyl monomer (s) in the presence of the latex of the core polymer obtained above and in the presence of a chain transfer agent.
  • the monomers used in the polymerization of the core polymer and in the polymerization of the polymer of the outer layer are the monomers listed above. They are either all used before the polymerization or for Part used before the polymerization, the remaining part being added to the reaction medium in successive fractions or continuously in the course of the polymerization, or all added in successive fractions or continuously in the course of the polymerization.
  • copolymerizable ethylenic mono- or polycarboxylic acids acrylic acid, methacrylic acid, maleic acid, fumaric acid, crotonic acid, sorbic acid, cinnamic acid, itaconic acid, aconitic acid may be mentioned, in amounts between 0.5 and 15% by weight, preferably between 0.5 and 10% by weight. -%, based on the monomer or monomers used.
  • the copolymerizable unsaturated alkali organosulfonates are e.g. Sodium vinyl sulfonate, sodium methallyl sulfonate, sodium 2-sulfoethyl acrylate, sodium 2-sulfoethyl methacrylate, 2-acrylamido-2-methylpropane sulfonate; they are used in amounts between 0.1 and 3% by weight, based on the monomer or monomers.
  • copolymerizable ethylene mono- or polycarboxylic acids and the copolymerizable unsaturated alkali organosulfonates can be used individually or in combination in the amounts specified.
  • the core polymer is prepared in emulsion by any conventional method in the presence of an initiator and an emulsifier.
  • the initiator used is preferably alkali metal sulfates, water-soluble diazo derivatives or redox systems based on hydrogen peroxide, organic peroxides or hydroperoxides in amounts of the order of 0.01 to 5% by weight, preferably 0.03 to 3% by weight, based on the or the monomers.
  • the emulsifier used can be anionic and / or nonionic. These are classic products for emulsion polymerization.
  • Salts of fatty acids may be mentioned as anionic emulsifiers; Alkaline alkyl sulfates, Alkalialkylsulfonate, Alkalialkylarylsulfonate, Alkalialkylsulfosuccinate, Alkalialkylphosphate; Alkyl sulfosuccinate; Sulfonates of alkylphenol polyglycol ethers; Salts of esters of alkylsulfopolycarboxylic acids; Condensation products of fatty acids with oxyalkanesulfonic acids and aminoalkanesulfonic acids; sulfated derivatives of polyglycol ethers; sulfated esters of fatty acids and polyglycols; Alkanolamides of sulfated fatty acids.
  • Suitable nonionic emulsifiers are fatty acid esters of polyalcohols, alkanolamides of fatty acids, polyethylene oxides, copolyethylene oxide / propylene oxide and oxyethylated alkylphenols.
  • the amounts of the emulsifier or emulsifiers to be used are of the order of magnitude of 0.01 to 5% by weight, based on the monomer or monomers, and they are introduced either in total before the polymerization or in part before the polymerization, the remaining part is added in the course of the polymerization in successive fractions or continuously to the reaction medium, or overall in the course of the polymerization in successive fractions or continuously.
  • the amount of water to be used for the polymerization of the core polymer must be such that the concentration of the monomer or monomers does not exceed 60% by weight.
  • any compounds to the reaction medium which are capable of modifying either the ionic strength of the medium and consequently the particle size distribution, such as mineral salts, electrolytes, in an amount of up to 3% by weight, based on the Monomers, or which can modify the pH of the medium, such as buffers, acids, bases.
  • the medium is neutral or acidic.
  • the polymerization temperature which is a function of the initiator used and the polymer to be prepared, is generally between -5 and + 90 ° C.
  • the latices obtained have polymer particles with a diameter between 0.03 and 5 ⁇ m, preferably between 0.05 and 1 ktm. These particles are generally not calibrated, but it is possible to obtain them calibrated using known calibration methods for emulsion polymerization, such as the controlled addition of the emulsifier and / or the monomer (s) and in particular the inoculation. In the latter case, the emulsifier can be contained in the inoculum.
  • the particles are formed from homopolymer or copolymer with a surface of carboxyl and / or sulfonate functions. The presence of these functions can be confirmed by conductometric titration.
  • the preparation of the polymer of the outer layer is carried out in an aqueous emulsion in the presence of the core polymer, chain transfer agent, initiator and, if appropriate, emulsifier.
  • the amount of the core polymer used is between 30 and 99.5% by weight and is preferably 60 to 99% by weight, based on the sum of the core polymer and the monomer or monomers to be polymerized.
  • the chain transfer agent of the aminophenyl disulfide or aminophenyl mercaptan type is in particular o, o'-dithiobisaniline, p, p'-dithiobisaniline, 2-mercaptoaniline, 3-mercaptoaniline, 4-mercaptoaniline.
  • This remedy is generally in solution used in the monomer or monomers, in amounts between 0.1 and 10% by weight, preferably between 0.5 and 5% by weight, based on the monomer or monomers.
  • the initiators required for the polymerization of the outer layer or monomers are diazo initiators, azonitriles such as azo-bis-isobutyronitrile or such as sulfonated azonitriles as described in French Patent No.
  • azobis isobutyronitrile sodium sulfonate
  • azobis a-methylbutyronitrile sodium sulfonate
  • azobis a-methyl- ⁇ -ethoxycarbonylbutyronitrile sodium sulfonate
  • carboxylated azonitriles such as 4,4'-azobis (4-cyanopentanoic acid) and their salts
  • azobis-alkylamidinium salts such as a, a'-azobis-isobutyramidinium chloride, azobis-N, N'-dimethylene-isobutyramidinium chloride.
  • the initiator which is used in an amount of 0.01 to 3% by weight, preferably 0.1 to 2% by weight, based on the monomer or monomers, is used in whole or in part before the polymerization, where the other part is added to the reaction medium in successive fractions or continuously in the course of the polymerization, in particular if the life of the initiator at the polymerization temperature is short.
  • the initiator can also be added continuously to the reaction medium overall in the course of the polymerization.
  • the emulsifier if any, is selected from anionic and / or nonionic emulsifiers which have been specified for the preparation of the core polymer; it can be the same or different from the emulsifier used for the production of the core polymer. It is used in amounts of up to 10% by weight, based on the monomer or monomers, and depending on the average diameter of the latex particles to be obtained, it can be introduced either entirely before the polymerization or partly before the polymerization take place, the remaining part in the course of the polymer tion is added in successive fractions or continuously, or it can be done in the course of the polymerization in successive fractions or continuously.
  • the amount of water to be used in the polymerization of the outer layer must be such that the concentration of core polymer and monomers to be polymerized or polymerized does not exceed 60% by weight, preferably 45% by weight.
  • the polymerization temperature which is a function of the chosen initiator, is generally between 5 and 100 ° C, preferably between 40 and 90 ° C.
  • the latices obtained have polymerization particles whose diameter is between 0.03 and 5 ⁇ m, preferably between 0.05 and 1 ⁇ m; since the amount of the outer layer is not very large, it does not noticeably modify the size of the particles of the core polymer.
  • the particles may or may not be calibrated, but in certain applications it is preferred for reasons of reproducibility that they are calibrated, i.e. that they have a narrow grain size distribution.
  • the latices are mechanical and resistant to storage and electrolytes, i.e. they do not flocculate if you are given mineral salts such as the chlorides, nitrates, borates, phosphates of sodium, calcium, magnesium, potassium are added.
  • the particles are formed from polymers and have a surface with carboxyl and / or sulfonate functions and groups of the formula: on.
  • the outer layer is polymerized on the core polymer, the carboxyl and / or sulfonate functions remain accessible, as can be shown by conductometric titration, and the groups of the formula: are available for further reactions.
  • the immunologically active materials can be bound physically and / or chemically to the latex polymers used according to the invention.
  • the reagent according to the invention is produced by forming an azo bond between the latex and the immunologically active material.
  • the primary aromatic amino groups of the latex are converted into a diazonium salt.
  • an inorganic acid such as hydrochloric acid, sulfuric acid or perchloric acid can be used.
  • Sodium nitrite or potassium nitrite is preferably used as the nitrite.
  • the reaction is preferably carried out at 0-5 C because of the instability of the diazonium salts.
  • the immunologically active material is then reacted with the diazotized carrier in an aqueous medium, preferably between 0.degree. C. and room temperature.
  • the immunologically active material can be bound to the latex used according to the present invention by means of a polyfunctional compound via an intermediate piece.
  • polyfunctional compounds are useful ones which react with the aromatic amino groups of the latex polymer or undergo a substitution reaction at the aromatic ring of the latex polymer and at the same mino- with functional groups of the immunologically active material such as A, mercapto, carbonyl and hydroxyl groups react or undergo a substitution reaction on the aromatic ring of the immunologically active material.
  • Representative representatives of such polyfunctional compounds are azó, isocyano, isothiocyano or aldehyde group-containing compounds such as e.g. Bis-diazobenzidine, bis-diazobenzidine-disulfonic acid, bis-diazo-p-phenyl diamine, phenyl diisocyanate, toluene diisocyanate, glutardialdehyde.
  • the immunologically active material When reacting in the presence of a bifunctional compound, the immunologically active material is reacted with the carrier in an aqueous medium, preferably at room temperature (20 ° C. to 25 ° C.). However, the temperature can also be between 0 ° C and 40 ° C.
  • the amount of the bifunctional compound used depends on the number of amino groups on the latex. A ten- to hundred-fold molar excess of the bifunctional compound compared to the number of amino groups of the latex used is preferably used.
  • the pH of the reaction is important. It should not be chosen to denature a protein reactant. Usually the pH is between 5 and 9. This pH is determined using suitable ones. Buffer systems such as phosphate buffers and the like are maintained.
  • the final product is a water-soluble material which is suspended in an aqueous buffer solution of pH 5.0 to 9, the pH of the solution being different from that in detail used system and depends on the requirements for the stability of the immunologically active material.
  • the specific weight of the product corresponds approximately to that of water (0.97-1.02), whereby a stable suspension of the product is achieved.
  • the products can be isolated, for example, by centrifugation in the form of a white or yellowish precipitate.
  • the amount of immunologically active material which is bound to the immunologically inert latex polymer carrier is usually 0.01 to 15.0% by weight. However, each individual immunologically active material is used in an amount which is most useful in a diagnostic test. For this reason, each material is combined with the carrier in a ratio that best suits the specific requirements.
  • the present invention therefore encompasses the use of such an amount of immunologically active material in combination with an immunologically inert latex polymer carrier which is suitable for providing a reagent useful for such diagnostic purposes.
  • the product can be used in specific diagnostic tests based on immunological principles.
  • the determination of the immunologically active substance can be carried out both in a direct and in an indirect (inhibition) test method.
  • the analytical sample and the latex particles coated with the corresponding immunological reaction partner are mixed to determine an immunologically active substance and the occurrence of agglutination is observed.
  • the test is positive if agglutination is detected.
  • the analytical sample is mixed with a certain amount of the corresponding immunological reaction partner (e.g. antiserum) and latex particles coated with the immunologically active substance to determine an immunologically active substance and the occurrence of agglutination is observed.
  • the test is positive if no agglutination is found.
  • the reagents which can be used in such immunological test methods can advantageously be packaged in a diagnostic test set for commercial purposes.
  • the reagent set for determining an immunologically active substance in a container contains an aqueous suspension of latex particles coated with the corresponding immunological reaction partner.
  • the reagent set for determining an immunologically active substance contains a solution of the corresponding immunological reaction partner (e.g. antiserum) in a first container and in. in a second container an aqueous suspension of latex particles coated with the immunologically active material.
  • the corresponding immunological reaction partner e.g. antiserum
  • the aqueous suspension of the latex-bound immunologically active material or latex-linked immunological reactant can be present in any concentration. However, a concentration of 0.5 to 5% by weight is preferred.
  • the polymerization is carried out at 75 ° C. under a nitrogen atmosphere, the monomers being introduced continuously over a period of 7 hours and the reaction being continued for 8 hours.
  • Electron microscopy shows that the particles have an average diameter of 0.145 ⁇ m; 90% of the particles have a diameter between 0.14 and 0.15 ⁇ m.
  • composition of the polymer is essentially the same as that of the monomers used.
  • the particles have carboxyl and sulfonate functions on their surface, which are determined by conductometric titration.
  • average diameter of the particles 0.15 m, 90% having a diameter between 0.145 and 0.155 pm.
  • the particles have carboxyl and sulfonate functions on their surface, which are confirmed by conductometric titration, and groups of the formula:
  • washed latex 1 ml of the 10% latex produced above is added to 20 ml of water and the mixture is centrifuged at 35,000 g for 1 1/2 hours. The supernatant is decanted off, the residue is taken up in 20 ml of water and centrifuged again at 35,000 g for 1 1/2 hours. This operation is repeated twice and the latex thus obtained is referred to as "washed latex" in the following examples.
  • the latex is centrifuged off at 35,000 g for 1 1/2 hours, the supernatant is decanted off and the residue is washed twice with 25 ml of 0.1 M glycine-NaOH buffer pH 8.2, by centrifuging and slurrying the residue. So much buffer is now added to the latex that a solution with 30 mg / ml results.
  • the following buffer is used for the tube agglutination test: 7.5 g glycine, 6.0 g CaCl 2 , 3 g bovine albumin, 1 g NaN 3 dissolved in 1 liter water. The pH is adjusted to 8.2 with NaOH. To detect the rheumatoid factors in the serum, 20 ⁇ l of latex is diluted with 3 ml of buffer in a small test tube and 25 ⁇ l of the serum to be examined is added. After mixing, the tubes are kept in a heat block at 37 ° C for 2 hours. A positive serum agglutinates under these conditions, while a negative control system shows no agglutination.
  • 1 ml of washed latex is prepared and diazotized as in Example 1.
  • the diazotized latex residue becomes 5 ml ice-cold 0.1 M glycine-NaOH buffer pH 6.0 was added and 5 mg goat anti-human albumin immunoglobulin G dissolved in 1 ml of the above buffer were added and the mixture was left to stir overnight at 10 ° C. for 1 hour.
  • the latex is centrifuged off at 35,000 g for 1 1/2 hours, the supernatant is discarded and the sediment is washed twice with 25 ml of 0.1 M glycine-NaOH pH 8.2. After washing, the latex is mixed with so much buffer that a 3% solution is obtained.
  • the following buffer is used for the tube agglutination test: 7.5 g glycine, 6.0 g CaCl 2 , 3 g bovine albumin and 1.0 g NaN 3 are dissolved in 1 liter water and the pH is adjusted to 6.0 with hydrochloric acid. A series of concentrations of human albumin in 3 ml of buffer is prepared in small test tubes, 20 ⁇ l of latex are added in each case and, after mixing, kept in a heat block at 37 ° C. for 2 hours.
  • Example 1 1 ml of washed latex from Example 1 is added to 5 ml of 0.1 M phosphate buffer pH 5.0 and 5 mg of sheep anti-human IgG immunoglobulin G in 1 ml of buffer are added and the mixture is stirred well. Then 0.1 ml of a 0.01 M p-phenyldiisothiocyanate solution in dimethylformamide added, stirred for 1 hour and at Allow room temperature to stand overnight. The latex is centrifuged at 35,000 g for 1 1/2 hours, the supernatant is discarded and the residue is washed twice with 25 ml of 0.1 M glycine-NaOH buffer pH 8.2. The latex is used in a concentration of 30 mg / ml for the agglutination test.
  • a 0.1 M phosphate buffer pH 6.0 with 0.1% bovine albumin is used for the tube agglutination test.
  • a series of concentrations of human IgG in 3 ml of buffer are prepared.
  • 20 ⁇ l of latex reagent are added to each tube, mixed and incubated for 2 hours at 37 ° C in a heat block.
  • the table shows that 0.1 ⁇ g / ml human IgG can still be determined with this latex reagent.
  • Example 1 1 ml of washed latex from Example 1 is taken up in 5 ml of 0.1 M phosphate buffer pH 7.0 and 5 mg of human immunoglobulin G in 1 ml of the above buffer is added. The suspension is cooled to 0 ° and 0.01 ml of a 0.02 M with stirring. bisdiazot convinced benzidine solution was added and then at 1 0 0 left to stand overnight. The latex is centrifuged at 30,000 g for 1 1/2 hours, the supernatant is discarded and the sediment is washed twice with 25 ml of 0.1 M glycine-NaOH pH 8.2. After washing, the latex is mixed so much buffer that a 3% solution is obtained.
  • a 0.1 M phosphate buffer pH 6.0 is used to determine IgG in the inhibition test. 3 ml of a 1/500 diluted sheep anti-human IgG serum and increasing amounts of human IgG are placed in small test tubes. After incubation for 15 minutes at 37 ° C, 20 ul latex reagent is added to each tube and incubated for 3 hours at 37 ° C.

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  • Health & Medical Sciences (AREA)
  • Immunology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Chemical & Material Sciences (AREA)
  • Hematology (AREA)
  • Urology & Nephrology (AREA)
  • Biotechnology (AREA)
  • Biochemistry (AREA)
  • Cell Biology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Microbiology (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Addition Polymer Or Copolymer, Post-Treatments, Or Chemical Modifications (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Polymerisation Methods In General (AREA)
EP78100581A 1977-08-03 1978-08-02 Réactif immunologique sous forme de particules spéciales de latex à base de polymère de vinyle, revêtues d'un matériau immunologiquement actif. Procédé pour sa fabrication. Utilisation de ce réactif. Procédé d'analyse utilisant ce réactif et trousse à réactifs le contenant Withdrawn EP0000772A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CH9542/77 1977-08-03
CH954277A CH628738A5 (de) 1977-08-03 1977-08-03 Immunologisches diagnose-reagenz.

Publications (1)

Publication Number Publication Date
EP0000772A1 true EP0000772A1 (fr) 1979-02-21

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Application Number Title Priority Date Filing Date
EP78100581A Withdrawn EP0000772A1 (fr) 1977-08-03 1978-08-02 Réactif immunologique sous forme de particules spéciales de latex à base de polymère de vinyle, revêtues d'un matériau immunologiquement actif. Procédé pour sa fabrication. Utilisation de ce réactif. Procédé d'analyse utilisant ce réactif et trousse à réactifs le contenant

Country Status (12)

Country Link
US (1) US4226747A (fr)
EP (1) EP0000772A1 (fr)
JP (1) JPS5428816A (fr)
AU (1) AU528150B2 (fr)
CA (1) CA1103153A (fr)
CH (1) CH628738A5 (fr)
DE (1) DE2833886A1 (fr)
DK (1) DK343678A (fr)
FR (1) FR2399665A1 (fr)
GB (1) GB2001996B (fr)
NL (1) NL7808143A (fr)
SE (1) SE7808349L (fr)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0008682A1 (fr) * 1978-08-02 1980-03-19 F. HOFFMANN-LA ROCHE & CO. Aktiengesellschaft Latex recouvert d'un matériau de protéine, procédé de préparation de ce latex, réactif immunologique contenant ce latex, procédé de préparation de ce réactif, application de ce réactif, procédé d'analyse utilisant ce réactif et trousse contenant ce réactif
EP0019741A1 (fr) * 1979-05-07 1980-12-10 BEHRINGWERKE Aktiengesellschaft Réactif de latex, procédé pour sa préparation, son utilisation et préparation contenant ce réactif
FR2595826A1 (fr) * 1986-03-13 1987-09-18 Lurhuma Zirimwabagabo Produit d'immuno-essai, son procede de preparation, son utilisation, complexe immunogene le comportant et utilisation de ce complexe

Families Citing this family (23)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CH628738A5 (de) * 1977-08-03 1982-03-15 Hoffmann La Roche Immunologisches diagnose-reagenz.
DE3048883A1 (de) * 1980-12-23 1982-07-15 Boehringer Mannheim Gmbh, 6800 Mannheim Hydrophile latexpartikel, verfahren zu deren herstellung und deren verwendung
JPS57168163A (en) * 1981-04-10 1982-10-16 Japan Synthetic Rubber Co Ltd Carier for immunoserological inspection reagent
DE3116995A1 (de) * 1981-04-29 1982-11-25 Röhm GmbH, 6100 Darmstadt Latex zur immobilisierung von biologisch wirksamen substanzen
JPS585658A (ja) * 1981-07-02 1983-01-13 Japan Synthetic Rubber Co Ltd 免疫血清学的検査試薬用担体
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DE2204684A1 (de) * 1971-02-01 1972-08-17 Hoffmann La Roche Verfahren zum Nachweis von Antikörpern bzw. Antigenen in einer Körperflüssigkeit
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DE2204684A1 (de) * 1971-02-01 1972-08-17 Hoffmann La Roche Verfahren zum Nachweis von Antikörpern bzw. Antigenen in einer Körperflüssigkeit
US3857931A (en) * 1971-02-01 1974-12-31 Hoffmann La Roche Latex polymer reagents for diagnostic tests
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EP0008682A1 (fr) * 1978-08-02 1980-03-19 F. HOFFMANN-LA ROCHE & CO. Aktiengesellschaft Latex recouvert d'un matériau de protéine, procédé de préparation de ce latex, réactif immunologique contenant ce latex, procédé de préparation de ce réactif, application de ce réactif, procédé d'analyse utilisant ce réactif et trousse contenant ce réactif
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FR2595826A1 (fr) * 1986-03-13 1987-09-18 Lurhuma Zirimwabagabo Produit d'immuno-essai, son procede de preparation, son utilisation, complexe immunogene le comportant et utilisation de ce complexe
EP0238396A1 (fr) * 1986-03-13 1987-09-23 Zirimwabagabo Lurhuma Produit d'immuno-essai, son procédé de préparation, son utilisation, complexe immunogène le comportant et utilisation de ce complexe

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NL7808143A (nl) 1979-02-06
FR2399665B1 (fr) 1982-11-05
CH628738A5 (de) 1982-03-15
AU528150B2 (en) 1983-04-14
US4226747A (en) 1980-10-07
DK343678A (da) 1979-02-04
FR2399665A1 (fr) 1979-03-02
CA1103153A (fr) 1981-06-16
SE7808349L (sv) 1979-02-04
GB2001996B (en) 1982-03-24
AU3840778A (en) 1980-01-31
JPS5428816A (en) 1979-03-03
DE2833886A1 (de) 1979-02-22
GB2001996A (en) 1979-02-14

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