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Antibody against hemagglutinin of influenza A H1N1 virus

A technology of hemagglutinin protein and influenza virus, which is applied in the field of antibodies against H1N1 influenza virus hemagglutinin protein, and can solve the problem of no specific monoclonal antibody for H1N1 influenza virus hemagglutinin protein

Active Publication Date: 2011-11-16
CENT FOR EXCELLENCE IN MOLECULAR CELL SCI CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, there is currently no monoclonal antibody specifically directed against the hemagglutinin protein (HA) of influenza A (H1N1) virus in the art

Method used

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  • Antibody against hemagglutinin of influenza A H1N1 virus
  • Antibody against hemagglutinin of influenza A H1N1 virus
  • Antibody against hemagglutinin of influenza A H1N1 virus

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preparation example Construction

[0050] Preparation of monoclonal antibodies

[0051] The monoclonal antibody of the present invention is prepared using hybridoma technology (see Kohler et al., Nature 256; 495, 1975; Kohler et al., Eur.J.Immunol.6:511, 1976; Kohler et al., Eur.J.Immunol.6 : 292, 1976; Hammerling et al., In Monoclonal Antibodies and T Cell Hybridomas, Elsevier, N.Y., 1981). The preparation method of a kind of hybridoma cell of monoclonal antibody of the present invention is: (1) utilize type A H1N1 influenza virus vaccine to immunize mice; (2) separate the splenocytes of immunized mice, and SP2 / 0 myeloma Cell line fusion; (3) adding mouse peritoneal feeder cells to co-culture with fusion cells, and screening positive strains; (4) cloning and screening by limiting dilution method to obtain monoclonal antibody cell lines, and screening from the cell lines can A cell line producing the desired monoclonal antibody.

[0052] The monoclonal antibody of the present invention can also be obtained by...

Embodiment approach

[0053] After the hybridoma cells are obtained, monoclonal antibody production techniques well known to those skilled in the art can be used to mass-produce the monoclonal antibody of the present invention. As an embodiment of the present invention, the monoclonal antibody can be prepared by the following preparation method, which includes the steps of: (1) providing adjuvant pretreated mice; (2) intraperitoneally inoculating the mice The hybridoma cells secrete monoclonal antibodies; (3) ascitic fluid is extracted, and the monoclonal antibodies are obtained by separation. As a way, the method of isolating monoclonal antibody from ascites is: collect ascites, precipitate with ammonium sulfate, and then use Protein G prepacked chromatographic column to purify to obtain high-purity anti-H1N1 influenza virus HA monoclonal antibody .

[0054] In addition, the hybridoma cells can also be cultured and expanded in vitro according to conventional animal cell culture methods, so as to ...

Embodiment 1

[0088] Example 1. Preparation of anti-H1N1 influenza virus hemagglutinin protein (HA) monoclonal antibody

[0089] 1. Influenza A (H1N1) virus vaccine

[0090] In the present invention, influenza A H1N1 virus vaccine (obtained from Hualan Biotechnology Co., Ltd., concentration 100ug / ml) was used to immunize BALB / c mice to prepare monoclonal antibodies.

[0091] 2. Animal immunization

[0092] Take 100 μg of H1N1 influenza virus vaccine mixed with an equal volume of Freund's adjuvant, emulsify completely, and inject subcutaneously on the back and feet of BALB / c mice to immunize mice. Vaccine once every 3 weeks, and 3 times in a row. Complete Freund's adjuvant was used for the first immunization, and incomplete Freund's adjuvant was used for the second and third immunizations. One week after the third immunization, blood was collected from the orbit, the serum was separated, and the titer of anti-H1N1 influenza virus vaccine antibody was measured. A mouse with a high titer w...

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Abstract

The invention relates to monoclonal antibody against hemagglutinin of influenza A H1N1 virus. The invention further provides a hybridoma cell strain secreting the monoclonal antibody. The invention further provides a kit containing the monoclonal antibody. With the present invention, the monoclonal antibody combines with conformational epitope of the hemagglutinin of the influenza A H1N1 virus; the monoclonal antibody has high affinity with a HA1 fragment of the hemagglutinin of the influenza A H1N1 virus; the monoclonal antibody does not generate cross reactions with other proteins; the monoclonal antibody has very high specificity and sensitivity.

Description

technical field [0001] The invention belongs to the field of biotechnology; more specifically, the invention relates to an antibody against hemagglutinin protein of influenza A (H1N1) virus. Background technique [0002] Influenza virus is a virus that causes influenza in humans and animals. It belongs to the Orthomyxoviridae taxonomically and is a single-stranded negative-sense RNA virus. Human influenza viruses can be divided into three types according to the antigenicity of their nucleoprotein (NP), A, B, and C (A, B, C). For type A influenza virus, according to the antigenicity of its surface glycoproteins hemagglutinin (HA) and neuraminidase (NA), it can be further divided into different subtypes. Currently, Type A viruses have 16 HA subtypes (H1-H16) and 9 NA subtypes (N1-N9). Can cause acute respiratory infection and even severe systemic immune response. Influenza viruses spread rapidly through the air, and there are often periodic pandemics around the world, causi...

Claims

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Application Information

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IPC IPC(8): C07K16/10C12N5/20G01N33/577G01N33/543A61K39/42A61P31/16C12R1/91
Inventor 孙兵黄潇边超凌志洋崔骏吴洪强孙轶卓胡贵玉
Owner CENT FOR EXCELLENCE IN MOLECULAR CELL SCI CHINESE ACAD OF SCI
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