RU2372951C2 - Polyaniline as sorbents for removing viruses, non-viral proteins and as immunosorbent base, method of removing or attaching viruses using said sorbents, immunoadsorption method using said sorbents, sorption method using said sorbents - Google Patents

Abstract

FIELD: chemistry.

SUBSTANCE: description is given of polyaniline in form of an emaraldine base of formula [(-C₆ H₄-NH)₂-(NH=C₆·H₄=NH-)]_n n=20 to 10000 or interpolymer of a complex of polyaniline with poly-(2-acrylamido-2-methyl-1-propane-sulphonic acid), i.e. salt of emaraldine with poly-2-acryloamido-2-methyl-1-propane sulphonic acid of formula

,

n=50 to 50000 as a sorbent for removing viruses, non-viral proteins and for making an immunoasorbent based on said sorbent for isolating antiviral antibodies.

EFFECT: following methods are also described: method of removing viruses through immobilisation on a sorbent; immunoadsorption method; method of sorption of non-viral proteins from complex mixtures using sorbent.

20 cl, 2 ex, 4 tbl, 2 dwg

Description

The invention relates to medicine and relates to the use of polyaniline compounds as a sorbent for the removal of viruses, non-viral proteins and as a matrix for the production of immunosorbent and can be used for diagnostic purposes, for deactivation and removal of viruses from the environment.

The wide spread of harmful substances, toxic to humans, pathogenic microorganisms, including viruses, raises the question of decontamination and their removal from the environment. It is known that for the removal of viruses from solutions, there are two methodological approaches — ultracentrifugation or their binding using sorbents.

Immunosorption - the extraction of antibodies or antigens from complex mixtures using immunosorbents, based on the antigen-antibody reaction ("Encyclopedic Dictionary of Medical Terms". M., "Soviet Encyclopedia", 1982, v.1, s. 409). With this technology, organic polymers or inorganic materials are used as a matrix for the preparation of immunosorbents, which are complexes of sorbents with antigen or antibodies, which are used to extract antibodies or antigens from complex mixtures, respectively. Matrices for the preparation of cellulose-based immunosorbents are known (Gurvich AE Use of cellulose matrices in immunochemistry. In the book: “Immunosorbents and their use in biotechnology”, M., 1987, pp. 5-22). Immunosorbents based on vermiculite (natural silicon compounds) or a mixture of vermiculite with magnetic powder (RU 2192013, G01N 33/531) are proposed.

Using immunosorption, specific antibodies can be isolated and detected, for example, against viruses. In particular, to identify antibodies to hepatitis C virus, a polystyrene with a fixed (immobilized) virus-specific recombinant protein or synthetic peptide (RU 2095815, G01N 33/543, RU 2138286, G01N 33/53) was proposed as a matrix (carrier). These immunosorbents are limited in use since solve a highly specialized problem, are intended only to improve the diagnosis of viral hepatitis C and are carried out on well-known materials used for diagnostic purposes.

The classical method of obtaining (isolation) of antibodies to influenza virus using immunosorption is the use of chicken red blood cells with the virus adsorbed on them (Rovnova Z. I. Obtaining specific anti-influenza sera “Issues of Virology”, 1959, No. 4, pp. 465-470) Known BaSO ₄ based immunosorbent for isolation of influenza antibodies. For this purpose, a dry BaSO ₄ pistol at 4 ° C is incubated for 1 h with a virus-containing liquid, based on 1 g of BaSO ₄ per initial concentration of influenza virus - a million hemagglutinating units (GE) or 1000 GE per 1 mg (Zakstelskaya L.Ya. Shenderovich SF Use of an immunosorbent to remove anti-influenza antibodies from diagnostic sera. "Laboratory case", 1979, No. 12, pp. 748-749).

Carbon-containing immunosorbents have been proposed for sorption of specific anti-influenza antibodies from immune sera. To this end, ultrafine Z-split graphite with particles of 20-100 nm was incubated with fluids containing human influenza A and B viruses (Y. E. Kurochkina et al. “Method for purifying water and solutions contaminated with influenza viruses using carbon-containing sorbents” // Materials of the IX Congress of the All-Russian Scientific and Practical Society of Epidemiologists, Microbiologists and Parasitologists, M., 2007, vol. 1, pp. 247-248).

Immunosorption can also be used to isolate viruses with a specific antigenic structure. Such approaches require the preliminary immobilization of specific antibodies to the sorbent. A known method of immunosorption for the isolation of influenza viruses with a specific antigenic structure of surface proteins. As a matrix for the preparation of an immunosorbent, a sorbent prepared in the form of granules from a polyvinyl alcohol cryogel with immobilized (fixed) virus-specific antibodies immobilized on it (Plieva F.M., Isaeva E.I., Lozinsky V.I. Application of polyvinyl alcohol cryogels in biotechnologies VI. Bioaffinity sobrents based on an ultra-macroporous carrier for working with viral particles, Biotechnology, 1998, No. 5, pp. 32-37).

There is a known development in which for the elution and concentration of the avian influenza virus, magnetic immunosorbents are used, the matrix of which is a solid magnetic base with immobilized (fixed) homologous antibodies. The setup created for these purposes allows monitoring open water bodies for the presence of a pathogen (I.V. Levchenko, V.I. Efremenko, D.K. Lvov, etc. Development of magneto-immunosorbent test systems and a selective concentration unit for detection of avian influenza viruses // Materials of the IX Congress of the All-Russian Scientific and Practical Society of Epidemiologists, Microbiologists and Parasitologists, M., 2007, vol. 1, pp. 248-249).

As you know, the elution of viruses from solutions can be carried out not only by immunosorption. One of the first organic sorbents for the binding (agglutination) of influenza viruses was formalized chicken red blood cells. This classical method requires the maintenance of a specific temperature regime, since otherwise, the erythrocyte complexes with viruses dissociate, and the virions enter the solution. The latter property was actively used in the isolation of influenza virus from solution with the subsequent receipt of purified eluates.

The use of polypyrrole in the form of membranes as a sorbent makes it possible to remove a number of animal viruses from aqueous solutions, including polioviruses, Coxsackie, echo and other enteroviruses, reo, company and hepatitis A viruses (US patent 6156202, application from 1999-03-30, Conducting polymer membrane and a process for the preparation of the same memdrane S. Chandra, R. Singh, H. Singh, AKNarula, S. Broor).

It is known to use for adsorption of viruses various modifications of carbon-containing materials, for example, activated carbon. Clark K.J., Sarr A.B et al. Vet. Microbiol. 1998, v.63 (2-4), p.137-146 clay or charcoal were used as sorbents for rotavirus and coronaviruses of cows that cause gastroenteritis in mammals and birds. Using a carbon sorbent, enteroviruses (type 2 poliovirus) can be removed from solutions (Zadorozhnaia V. I. "Mikrobiol. J"., 1996, v. 58 (3), p. 83-87). In the patent RU 2237022, C02F 1/28, C02F 1/50, publ. 09/27/2004, US priority 05/20/1999 a carbon sorbent is proposed, which is an activated carbon particle capable of removing bacterial viruses from the water, for example, MS-2 bacteriophages having a size of 25 nm or more. It should be noted that, depending on the composition and method of producing activated carbon, a wide range of their sorption properties is observed, in particular, pore size distribution and specific surface area. A known method of removing influenza A and B viruses in humans using ultrafine Z-split graphite obtained from modified graphite by hydrothermal treatment. The authors of the development propose to use this method for cleaning water bodies (Kurochkina Y.E. et al. “Method for the purification of water and solutions contaminated with influenza viruses using carbon-containing sorbents” // Materials of the IX Congress of the All-Russian Scientific and Practical Society of Epidemiologists, Microbiologists and Parasitologists, M., 2007, vol. 1, p. 247-248).

US Pat. No. 7,018,538, published March 28, 2006, provides data on the preparation of DNA preparations from Escherichia coli bacterial lysate. The lysate was applied to a chromatographic column filled with carriers (glass beads or discs coated with polyaniline or its derivatives). Samples containing DNA exited the column in the first fractions after elution with TE buffer. The degree of DNA purification was checked electrophoretically in an agarose gel. No sorption and elution of proteins from the E. coli lysate were reported.

The present invention solves the problem of increasing the sorption ability of the sorbent to substances of a protein nature, to viruses having an outer protein shell with various antigenic properties, to obtain an immunosorbent based on it to isolate specific antiviral antibodies, simplify and expand the conditions for sorption of viruses, and also to be able to have a sorbent , the texture of which can be changed depending on the intended application (powder or film).

This is achieved through the use of polyaniline as a sorbent: or emeraldine base (i.e., the main semi-oxidized form of polyaniline) of the formula

or salt form of the semi-oxidized polyaniline and poly- (2-acrylamido-2-methyl-1-propane-sulfonic acid of the formula

n = 50-50000

The choice of polyaniline was due to the fact that it is a synthetic polymer with a known structure, chemical composition and studied physicochemical properties. The chemical structure of polyaniline includes chemical functional groups of various nature, ensuring their interaction with hydrophobic and hydrophilic sites of antigens. The presence of these groups in the composition of polyaniline ensures their sorption ability to proteins.

The essence of the invention lies in the use of an emeraldine base (i.e., the basic semi-oxidized form of polyaniline) (PAN-O) or the salt form of the semi-oxidized polyaniline and poly- (2-acrylamido-2-methyl-1-propane-sulfonic acid of the formula (PAN-C ) as an sorbent for the removal of viruses, proteins of non-viral nature and as a matrix for the production of immunosorbent for the isolation of specific antiviral antibodies.

PAN-O is prepared according to the procedure described in Cao Y., Andreatta A., Heeger AJ, Smith P. Influence of chemical polymerization conditios on the properties ofpolyaniline, "Polymer", 1989, V.30, N.12, p. 2305-2311. PAN-S - according to the method described in the work of Ivanov V.F., Gribkova O.L., Cheberyako K.V., Nekrasov A.A., Tverskoy V.A., Vannikov A.V. Matrix synthesis of polyaniline in the presence of poly- (2-acrylamido-2-methyl-1-propane) sulfonic acid, Electrochemistry, 2004, v.40, No. 3, pp. 339-345.

PAN-O is a water insoluble powder.

The salt form of PAN-S is soluble in water and in aqueous-organic mixtures under certain conditions. Solutions are homogeneous highly dispersed systems, up to molecularly dispersed ones, stable for a long time. Atomic force microscopy data indicate the formation of interpolymer complexes of poly- (2-acrylamido-2-methyl-1-propane-sulfonic acid (PAMPSK) with a length of up to 50-300 nm and a diameter of 20-40 nm. In addition, linear associates of these complexes were found up to 1500 nm long (OLGribkova, GBMeshkov, VFIvanov, AAIsakova, AANekrasov, AVVannikov, IV Yaminsky, Nanoobjects of interpolymer complexes of polyaniline and PAMPSA in aqueous solutions, J. Phys .: Conf. Ser. 61 (2007 ) 359-363). PAN-O and PAN-S can be used in the form of powder or in the form of films, which expands the possibilities of using these sorbents. They differ in method and time In addition to synthesis, the degree of ordering of polymer units, both compounds were used later for binding viral particles and other proteins from various liquids.

This paper presents data on the study of the conditions of sorption and desorption of influenza A and B viruses with the proposed sorbents, sorption of non-viral proteins, as well as anti-influenza antibodies from immune sera using an immunosorbent, the matrix of which are the proposed sorbents.

Viruses

It is known that animal and bird viruses have many differences among themselves. This applies both to the size of virions, their genomes, the composition and properties of virion proteins, including surface ones (Gaidamovich S.Ya. Classification of viruses. In the book: “General and private virology.” M., Medicine, 1982, p. 26 -60). The viruses of the family of orthomyxoviruses (influenza viruses), paramyxoviruses, togoviruses (for example, flaviviruses) have an outer shell. The presence of glycoproteins in the outer shell determines their ability to be sorbed on animal erythrocytes.

To identify the sorption properties of the proposed polyanilines and for the preparation of immunosorbents, reference and epidemic strains of the influenza A virus (H1N1, H3N2) and B, isolated in the period 1977-2007, with different surface protein structures, were taken as representatives of viruses with an outer shell. and accordingly properties. Viruses were cultured in various systems (chicken embryos, MDCK tissue culture cells). We also used 10-50% sucrose concentrations purified in a gradient and concentrated allantoic viruses.

Methods

Definitions: 1) viruses in solution were carried out using the reaction of hemagglutinating activity (RGA); 2) antibodies in serum - the inhibition of hemagglutinating activity (RTGA) using 0.75% suspension of human red blood cells 0 (1) blood group; 3) the presence of protein in the preparations was determined by electrophoresis of samples in a polyacrylamide gel according to the method of Laemmli U.K. (Cleavage of structural proteins of the head ofbacteriophage T4, "Nature" 1970, v.227, p.680-685).

In studies with a sorbent at different stages, a shaker (Shaker PSU 2T plus) and a desktop low-speed centrifuge (Comb spin) were used.

Example 1. Obtaining a sorbent PAN-O.

The chemical synthesis of polyaniline was carried out at room temperature by adding a 0.11 M solution of ammonium persulfate in a 0.22 M solution of aniline hydrochloride in 2 M hydrochloric acid with stirring. The molar ratio of aniline and oxidizing agent was 2: 1 (Cao Y., Andreatta A., Heeger AJ, Smith P. Influence of chemical polymerization conditios on the properties ofpolyaniline, "Polymer", 1989, V.30, N.12, p. 2305-2311). The precipitate was filtered off, washed with distilled water, acetonitrile and ethanol, and then dried under vacuum. The resulting emeraldine salt was transferred to the base by treatment in a 15% aqueous ammonia solution for one hour. The precipitate was filtered off and dried in vacuo.

Example 2. Obtaining a sorbent PAN-S.

The chemical synthesis of PAN-S was carried out at room temperature with successive mixing of aqueous solutions of PAMPSK, aniline and ammonium persulfate. The ratio of PAMPSK and aniline was 1: 1 mol-unit / mol, and the ratio of aniline and oxidizing agent was 2: 1 mol / mol. The concentrations of aniline and PAMPSC in the reaction mixture are 0.04 M (Ivanov V.F., Gribkova O.L., Cheberyako K.V., Nekrasov A.A., Tverskoy V.A., Vannikov A.V. "Matrix synthesis of polyaniline in the presence of poly- (2-acrylamido-2-methyl-1-propane-sulfonic acid) ", Electrochemistry, 2004, v.40, No. 3 p.339-345). The resulting PAN-C was precipitated with dioxane and dried under vacuum.

3. Sorption of influenza viruses on sorbents.

Sorbents in the experiments used in the form of powder or in the form of films. To determine the optimal ratio of the mass of sorbents in the form of a powder to a constant volume of the virus solution V = 200 μl, studies were conducted in which the mass of the sorbent ranged from 5 to 50 mg. As studies have shown, sorption of viruses occurred intensively with weighed portions of the sorbent at selected sorbent masses, therefore, in the future, all studies were carried out on a 5 mg sample.

The virus sorption procedure was as follows. The prepared sorbent was added to the virus-containing solution. For active contact of the virus with the sorbent, the reaction mixture was placed on a shaker. After the virus contacted the sorbent, the mixture was centrifuged at 2000 thousand rpm for 4 min, the supernatant was examined for the presence of the virus in the RGA. The precipitate containing the sorbent with the immobilized virus was investigated in experiments on desorption of the virus, as well as on the sorption of antibodies to the immunosorbent. Sorption of viruses was studied depending on the sorbents, antigenic viruses of influenza A and B, as well as on different experimental conditions.

A study of the sorption of different strains of influenza A and B viruses on any of the sorbents (PAN-O or PAN-S) is presented in Table 1. It was shown that all viruses successfully interacted with the sorbent under different conditions, regardless of the antigenic properties of surface proteins. Influenza viruses A and B were adsorbed onto the sorbent from any virus-containing liquid: from buffer solutions (Tris-HCl buffer with pH 7.2-STE, saline solution - FR), allantoic fluid of chicken embryos, both undiluted and diluted in FR, the culture fluid ( tissue culture cells MDCK). In the experiments, the weights of the sorbent ranged from 5 to 50 mg with a constant volume of virus-containing liquid V = 200 μl. The initial titers of the viruses studied in this experiment, in terms of 1 mg of sorbent, were in the range from 10 to 400 hemagglutinating units (GE). The final titers in terms of 1 mg of sorbent ranged from less than 2 to 10 GE. The drop in the hemagglutinating titer of influenza viruses contained in the solution after sorption occurred 8 - 256 times, depending on the initial concentration of viruses. Similar results were obtained using the proposed sorbents in the form of films.

The effect of temperatures on the sorption of viruses was analyzed. The data obtained showed that the adsorption of viruses with any of the sorbents occurred equally intensively in the temperature range from 4 to 37 ° C. The results of experiments with PAN-S are presented in table 2.

To determine the optimal contact time of the virus-sorbent, studies were conducted on the effect of the duration of the interaction of the virus with the sorbent on the sorption of viruses. The results of the study showed that the sorption of influenza viruses on any of the sorbents occurred almost identically in the time range from 15 to 120 minutes. Table 2 presents the results of experiments with PAN-S at different sorption times.

Thus, the sorption properties of the proposed sorbents were the same for different strains of the influenza virus and had a common pattern in the chosen experimental conditions.

4. Desorption of the virus from the immunosorbent.

As you know, an immunosorbent is a substance with antigens or antibodies adsorbed on it, used to extract antibodies or antigens from complex mixtures, respectively. Of particular interest was to determine how stable the virus is associated with an immunosorbent prepared on the basis of PAN-O or PAN-S. For this, we investigated the desorption of the virus from immunosorbents in two solutions of STE buffer and FR. Immunosorbents were mixed with solutions and incubated at 4, 22 and 37 ° C for 1, 24, 48 hours. Then, immunosorbents were precipitated and centrifuged for 5 minutes. The supernatant was examined for the presence of virus in the RGA. Virus desorption from immunosorbents was studied on allantoic viruses and after their purification by ultracentrifugation (Table 3). Thus, it was found that desorption of viruses from the surface of both sorbents does not occur in the selected temperature and time regimes.

5. The interaction of the immunosorbent with antibodies (immunosorption).

To determine whether the antigenic activity of virionic surface proteins, in particular the most important one, hemagglutinin, is preserved in the virus immobilized on a sorbent, we studied the interaction of sorbed viruses A / Sichuan / 2/87 A (H3N2) and B / Malaysia / 2506/04 s antibodies from immune sera obtained against these viruses. The prepared immunosorbents were mixed with a solution of immune serum containing homologous antibodies to this virus. The mixture was contacted under various temperature and time conditions, then centrifuged. The supernatant was examined for the presence of antibodies in solution in rtga. Analysis of the serum solution before and after interaction with the immunosorbent showed that serum titers were significantly reduced when using any of the proposed immunosorbents. Antibody binding occurred using any of the immunosorbents under various temperature and time conditions. Table 4 shows the results obtained at a temperature of 4 ° C.

6. Drying of viruses on a sorbent.

Of particular interest was to establish the ability to maintain immunosorbent for a long time. For this purpose, the method of lyophilization of viral preparations, widely used in virology and allowing the preservation of influenza viruses for many years, was applied.

To determine the effect of lyophilization on the properties of viruses combined with polyaniline powder, this complex was lyophilized in ampoules under standard lyophilization conditions for influenza viruses. After various storage periods (several months), the resulting preparation was examined for the activity of viruses in rtga. For this, the lyophilized immunosorbent was incubated with immune serum at a temperature of 4 ° C for 18 h and then the resulting complex was centrifuged. The serum titer in the supernatant decreased 4 times, which indicates the interaction of the virus, which is part of the immunosorbent with antibodies. These data also indicate the preservation of the antigenic properties of viruses in combination with polyaniline powder of both PAN-O and PAN-S sorbents after lyophilization.

7. Interaction of sorbents with proteins of non-viral nature.

It was important to study the adsorption of non-viral proteins on the PAN-O and PAN-S sorbents: 1% bovine serum albumin, which is widely used in virology in immunological reactions, immune serum proteins, and allantoic fluid proteins. Therefore, we sorbed these proteins onto sorbents at T = 22 ° C for 30 min. After centrifugation of the complex (sorbent + proteins), the amount of unsorbed protein in the supernatant was evaluated qualitatively by the presence of protein bands in PAG and their comparison with protein bands in the preparations before sorption on PAN-O and PAN-S. The Lowry method, widely used to estimate the concentration of protein in solutions, could not be used in the case of the use of PAN-O or PAN-S sorbents (Lowry OK, Roenbrough NJ Faff NJ, Randal RJ Protein measurement with folin reagent. "J. Biol. Chem." , 1951, Volume 193, pp. 265-267).

The fact is that part of the sorbent particles was so small that they did not precipitate during centrifugation up to 10,000 rpm and were present in test solutions, creating non-specific staining. Analysis of biological materials during polyacryamide gel electrophoresis before and after sorption on PAN-O or PAN-S sorbents showed that 1% bovine serum albumin sits by about 70%, allantoic proteins are completely immobilized, and immune serum proteins are sorbed by about 90%. Figures 1 and 2 show the electrophoresis of albumin and serum proteins.

Thus, the proposed sorbents — polyaniline (base and salt) prepared in two ways — are capable of absorbing enveloped viruses for a short time over a wide range of temperatures, using influenza viruses. Influenza viruses A and B, regardless of their antigenic structure, degree of purification and method of cultivation, using the proposed sorbents can be removed from virus-containing liquids. At the same time, immobilized viruses retained the ability to bind to specific antibodies contained in immune sera, even several months after lyophilization. These sorbents can also be used for other viruses. The data obtained indicate a wide range of protein materials that can be removed from various biological fluids using the proposed polyaniline samples prepared by different methods. These are specific antiviral antibodies and non-viral proteins: allantoic fluid proteins, immune serum proteins, including albumin.

These sorbents can be used to disinfect liquids. In recent years, cases of transmission of influenza A virus from birds to people in areas of close contact between birds and humans have become more frequent, the present invention can be used to remove viruses from reservoirs infected with avian influenza viruses. The obtained immunosorbents based on the proposed polyanilines can be used to determine the spectrum of antibodies in immune sera during diagnostic studies. The combination of the revealed adsorption properties of polyaniline preparations with unique physicochemical properties (conductivity, electrochromism, inertness), economy (cheap raw materials), allows us to start developing virus detectors or virus complexes with antibodies for rapid diagnosis of viral infections. The claimed compound as a sorbent is suitable not only for the removal of enveloped viruses (for example, influenza A and B viruses), but also for other viruses, for example poliomyelitis virus, bacteriophage.

Table 1. Adsorption of influenza A and B viruses on polyaniline sorbents Viruses Type of Degree of purification Virus titer in solutions (RGA) sorbent Before sorption After sorption A / Sichuan / 2/87 A (H3N2) Purified 4000 <2 PAN-O A / New Caledonia / 20/99 A (H1N1) Allantoic. Diluted. 1: 1 128 <2 64 2 A / Texas / 77 A (H3N2) Allantoic. Diluted. 1: 1 256 <2 128 <2 A / Philippines / 2/82 A (H3N2) Allantoic 64 2 A / California / 7/04 A (H3N2) Allantoic 256 <2 B / Shanghai / 361/02 AT Allantoic. Diluted. 1: 1 256 2 32 <2 A / Moscow / 12/03 A / Moscow / 48/03 B / Moscow / 18/03 A (H1N1) MDCK 16 2 A (H3N2) 128 16 AT 256 64 A / Sichuan / 2/87 A (H3N2) Purified, concentric. Divorced at STE 2048 8 A / California / 7/04 A (H3N2) 512 16 B / Malaysia / 2506/04 AT 2048 8 A / New Caledonia / 20/99 A (H1N1) Allantoic 1024 four A / Sichuan / 2/87 A (H3N2) 2048 8 A / California / 7/04 A (H3N2) 1024 four B / Shanghai / 361/02 AT 512 four A / Vladimir / 44/07 A (H1N1) MDCK 64 <2 PAN-S A / Yaroslavl / 58/07 A (H1N1) 128 <2 B / Yaroslavl / 29/06 AT 256 <2 Note: experiments were carried out under the following conditions T = 22 ° C, contact 1 = 15 min

Table 2. Sorption of influenza A / Sichuan / 2/87 on polyaniline sorbents (PAN-S) depending on the time of sorption and temperature. The dependence of virus sorption on Virus T min T ° C Virus titer in solution (RGA) before sorption after sorption Contact time A / Sichuan / 2/87 5 twenty 2048 8 fifteen twenty 16 thirty twenty 8 Temperature A / Sichuan / 2/87 fifteen four 2048 8 fifteen twenty 8 fifteen 37 8

Table 3. Desorption of influenza A and B viruses from polyaniline sorbents. Virus Type of Degree of purification Sorption Desorption 1 Desorption 2 Sorbent Virus titer before sorption Virus titer after sorption T min T, ° С Virus titer T min T, ° С Virus titer PAN-S A / New Caledonia / 20/99 AH1N1 Allantois. 1024 four 60 min four ± 2 N.S. N.S. N.S. 22 ± 2 37 <2 A / California / 7/04 AH3N2 Allantois. 1024 <2 48 h 22 <2 48 h four <2 B / Shanghai / 361/02 AT Allantois. Intelligence. STE 64 2,4 48 h 22 <2 48 h four <2 <2 PAN-0 A / Sichuan / 2/87 AH3N2 Cleared. 4096 <2 60 min four <2 24 h four <2 22 <2 22 <2 37 <2 36 <2 B / Shanghai / 361/02 AT Allantois. Intelligence. STE 16 <2 60 min 22 <2 24 h <2

Table 4. Study of the sorption of antibodies to influenza A and B viruses on polyaniline immunosorbents at 4 ° C. Immunosorbent Sorbed substance
Flu virus immune serum Serum titer (rtga) T h Before sorption After sorption PAN-O + virus A (H3N2) A (H3N2) 1: 1280 1:80 eighteen PAN-C + virus A (H3N2) A (H3N2) 1: 640 1:40 PAN-C + virus B AT 1: 1280 1:40 24

Claims (20)

1. Polyaniline in the form of an emeraldine base of the formula

n = 20-10000
or an interpolymer complex of polyaniline with poly- (2-acrylamido-2-methyl-1-propanesulfonic acid), i.e. salts of emeraldine with poly-2-acrylamido-2-methyl-1-propanesulfonic acid of the formula

n = 50-50000
as a sorbent for the removal of viruses, proteins of non-viral nature and to obtain an immunosorbent based on it for the isolation of antiviral antibodies. 2. The method of removing viruses by immobilizing them on a sorbent, including incubating it with a virus-containing liquid under certain temperature and time conditions, characterized in that the sorption is carried out using any of the sorbents according to claim 1. 3. The method according to claim 2, characterized in that the liquid contains viruses having an outer shell. 4. The method according to claim 2, characterized in that the liquid contains influenza viruses. 5. The method according to claim 2, characterized in that the liquid contains influenza A (H1N1) viruses. 6. The method according to claim 2, characterized in that the liquid contains influenza A viruses (H3N2). 7. The method according to claim 2, characterized in that the liquid contains influenza viruses B. 8. The method according to any one of claims 3 to 7, characterized in that the sorption is carried out at a temperature of 4-37 ° C for 15 or more minutes. 9. The method according to any one of claims 3 to 7, characterized in that the sorption is carried out with stirring. 10. The method of immunosorption, including mixing a solution containing specific antibodies and a sorbent with a fixed antigen on it, incubation at certain temperature and time conditions, centrifugation, characterized in that any of the antibodies obtained using the method according to claim 1 is used to isolate specific antibodies .2 sorbents with viruses fixed on it. 11. The method according to claim 10, characterized in that for the extraction of mixtures of specific antibodies to the virus, use any of the sorbents with viruses fixed on it, obtained by the method according to claim 3. 12. The method according to claim 10, characterized in that for the extraction of specific antibodies to the influenza virus from complex mixtures, use any of the sorbents with viruses fixed on it, obtained by the method according to claim 4. 13. The method according to claim 10, characterized in that for the extraction of specific antibodies to the influenza A (H1N1) virus from complex mixtures, use any of the sorbents with viruses fixed on it, obtained by the method according to claim 5. 14. The method according to claim 10, characterized in that for the extraction of specific antibodies to the influenza A (H3N2) virus from complex mixtures, use any of the sorbents with viruses fixed on it obtained by the method of claim 6. 15. The method according to claim 10, characterized in that for the extraction of specific antibodies to influenza B virus from complex mixtures, use any of the sorbents with viruses fixed on it, obtained by implementing the method according to claim 7. 16. The method according to any one of paragraphs.11-15, characterized in that to isolate specific antibodies, the incubation is carried out for 16 hours at 4 ° C. 17. The method of sorption of proteins of non-viral nature from complex mixtures using a sorbent, characterized in that the immobilization is carried out using any of the sorbents according to claim 1. 18. The method according to 17, characterized in that the proteins of the immune serum, including albumin, are removed from the mixtures. 19. The method according to 17, characterized in that albumin is removed from the mixtures. 20. The method according to 17, characterized in that the proteins of the allantoic fluid of the chicken embryos are removed from the mixtures.

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