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Common epitope, antibody, identification method and application of multiple subtype influenza virus HA2 proteins

An influenza virus, common antigen technology, applied in the direction of virus antigen components, antiviral immunoglobulins, viruses/phages, etc., can solve the problem of not having a broad spectrum, and achieve the effect of improving sensitivity and reducing steps.

Active Publication Date: 2020-07-24
YANGZHOU UNIV
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Epitopes identified on the HA1 protein of influenza A viruses tend to be subtype-specific and not broad-spectrum

Method used

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  • Common epitope, antibody, identification method and application of multiple subtype influenza virus HA2 proteins
  • Common epitope, antibody, identification method and application of multiple subtype influenza virus HA2 proteins
  • Common epitope, antibody, identification method and application of multiple subtype influenza virus HA2 proteins

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Embodiment

[0033] 1. Serum Preparation

[0034] Table 1 Background information of 13 strains of influenza virus and the hemagglutination inhibitory titer of their immune sera

[0035]

[0036] Preparation of whole virus immunized chicken serum: IAV was inactivated with formaldehyde, mixed with Tween 80 and white oil to make inactivated oil emulsion vaccine, and immunized 3-week-old SPF chickens with a dose of 0.3 mL / bird. A second immunization was performed 2 weeks later with the same dose. Serum was collected in the second week after the second immunization, and the hemagglutination inhibition (HI) titer was greater than or equal to 6log2 for future use (see Table 1 for details).

[0037] Preparation of peptide-immunized chicken serum: 100 μg of BSA-coupled peptide (synthesized by Shanghai GL Biochem Company) was dissolved in 0.1 mL of normal saline and mixed with an equal amount of Freund’s complete adjuvant (Sigma, USA), and subcutaneously immunized at 21 days old SPF chicken. T...

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Abstract

The invention relates to a multi-subtype influenza virus HA2 protein common epitope, an antibody, an identification method and application. The antigen epitope can be combined with immune serum of H1,H3, H4, H5, H6, H7, H8, H9 and H10 subtype influenza viruses. The method comprises the following steps: selecting an H5N1 subtype influenza virus strain S, designing 18 polypeptides with the length of 20 amino acids and overlapped 10 amino acids according to the sequence of the HA2 protein of the H5N1 subtype influenza virus strain S, artificially synthesizing, spotting to a modified silica gel film to prepare a polypeptide chip, and screening universal epitopes by using antiserum aiming at different subtype IAVs. The recombinant virus with the mutated epitope is constructed, and the bindingcapacity of the epitope and immune serum is analyzed through western blot. Results show that 485-FYHKCDNECME-495 on the HA2 protein has broad-spectrum binding activity with H1, H3, H4, H5, H6, H7, H8,H9 and H10 subtype IAV antiserum. K / D mutation in the recombinant virus leads to reduction of the serum binding activity, which indicates that the K / D mutation is a key amino acid of the serum binding activity. The peptide fragment also comprises an HLA-DR1 (Human Leukocyte Antigen-DR1) restrictive CD4 T cell epitope.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a common antigenic epitope, antibody, identification method and application of multiple subtype influenza virus HA2 proteins. Background technique [0002] Type A influenza virus (Influenza Avirus, IAV) consists of 8 segments of single-stranded negative-sense RNA, prone to antigenic drift and antigenic shift. Currently, 18 HA subtypes and 11 NA subtypes have been found, among which avian influenza virus (AIV) includes 16 HA subtypes and 9 NA subtypes. According to genetic evolution analysis, these 18 HA subtypes can be divided into two groups, Group 1 is H1, H2, H5, H6, H8, H9, H11, H12, H13, H16 and H17; Group 2 is H3, H4, H7, H10, H14 and H15. Due to the persistent endemic and seasonal epidemics, the antigenicity of IAV of the same subtype has also changed greatly, and different lineages have gradually been derived. The monitoring and control of IAV is also becoming more and mor...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/11G01N33/569G01N33/68C07K16/10A61K39/145A61P31/16
CPCA61K39/12A61K2039/552A61P31/16C07K14/005C07K16/1018C12N2760/16122C12N2760/16134G01N33/56983G01N33/6854G01N2333/11G01N2469/10
Inventor 彭大新王秋霞陈素娟秦涛孙志豪
Owner YANGZHOU UNIV
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