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Monoclonal antibody ZJU61-01 for resisting H6N1 avian influenza virus hemagglutinin protein and application of monoclonal antibody ZJU61-01

A monoclonal antibody and avian influenza virus technology, which is applied in the field of biotechnology, can solve the problems of infecting pigs and human new influenza viruses and the like, and achieves the effect that the preparation method is simple and feasible

Active Publication Date: 2021-05-25
THE FIRST AFFILIATED HOSPITAL ZHEJIANG UNIV COLLEGE OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the H6 subtype avian influenza virus is currently a low-pathogenic avian influenza virus, it is widely circulated in poultry and has the potential to repeatedly infect pigs and humans and mutate to evolve into a new type of influenza virus that infects humans.

Method used

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  • Monoclonal antibody ZJU61-01 for resisting H6N1 avian influenza virus hemagglutinin protein and application of monoclonal antibody ZJU61-01
  • Monoclonal antibody ZJU61-01 for resisting H6N1 avian influenza virus hemagglutinin protein and application of monoclonal antibody ZJU61-01
  • Monoclonal antibody ZJU61-01 for resisting H6N1 avian influenza virus hemagglutinin protein and application of monoclonal antibody ZJU61-01

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Experimental program
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Effect test

Embodiment 1

[0043] Embodiment 1. Preparation method of monoclonal antibody against H6N1 avian influenza virus hemagglutinin protein

[0044] (1) Immunization of mice: for the first immunization, the H6N1 avian influenza virus hemagglutinin whole protein and the adjuvant were uniformly mixed in equal volumes, and the total volume was 600 microliters. Each BALB / C mouse 0.1 ml (containing H6N1 avian influenza virus hemagglutinin whole protein antigen 30 micrograms), intramuscularly injected into the inner thigh. On the 21st day, a booster shot was given in the same way. On the 35th day, a small amount of tail blood was collected for enzyme-linked immunosorbent assay, and the antibody titer reached 1:16000. Immediately, a booster immunization was injected into the tail vein, and cell fusion was carried out 3 days later.

[0045] (2) Cultivation of mouse myeloma cell SP2 / 0: the SP2 / 0 myeloma cell line from BALB / C mouse was cultured and passaged in DMEM medium containing 10% bovine serum, and ...

Embodiment 2

[0053] Embodiment 2. Carry out the qualitative detection of H6 subtype avian influenza virus with this monoclonal antibody

[0054] The anti-H6N1 avian influenza virus hemagglutinin protein monoclonal antibody prepared by the present invention can be used for qualitative detection of H6 subtype avian influenza virus, and the identification method can be realized by the following methods:

[0055] H6 immunochromatographic colloidal gold test strip:

[0056] (1) Preparation of colloidal gold solution: get 1 200 ml glass bottle, add 49.5 ml of ultrapure water, then add 0.5 ml of 1% chloroauric acid to prepare 50 ml of 0.01% chloroauric acid aqueous solution, heat and boil and continue stirring Add 1.8 milliliters of 1% trisodium citrate solution at one time under certain conditions, continue to stir and heat, when the color of the solution turns completely transparent purple red, stop heating after maintaining for 5 minutes, add water to the original volume, and cool to room temp...

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Abstract

The invention provides a monoclonal antibody ZJU61-01 for resisting H6N1 avian influenza virus hemagglutinin protein and application of the monoclonal antibody ZJU61-01. The heavy chain amino acid sequence of the monoclonal antibody ZJU61-01 is as shown in SEQ ID No.2, and the light chain amino acid sequence of the monoclonal antibody ZJU61-01 is as shown in SEQ ID No.4. The monoclonal antibody is further subjected to physical and chemical property analysis and identification, and a method for detecting the H6 subtype avian influenza virus through a colloidal gold immunochromatography test strip and an enzyme-linked immunosorbent assay by taking the monoclonal antibody as a probe is established. The invention provides an effective tool for auxiliary diagnosis of H6 subtype avian influenza virus infection in a clinical sample, and can be popularized and applied to various detection technologies and clinical and experimental researches.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to the preparation and application of anti-H6N1 avian influenza virus hemagglutinin protein monoclonal antibody ZJU61-01, which is the hybridization of monoclonal antibody secreting anti-hemagglutinin protein by using cell engineering and antibody engineering technology The tumor cell line was induced ascites from the same strain of mice to prepare the monoclonal antibody ZJU61-01 against hemagglutinin protein, which was identified as IgG1 and κ type, and then the antibody was detected by affinity purification, electrophoresis, immunization and other technologies. application. Background technique [0002] The long-term influenza virus monitoring found that among the avian influenza A viruses, the H6 subtype avian influenza virus is the most frequently isolated, with a wider host range and a wider epidemic range. The H6N1 strain has crossed the species barrier and directly infected human...

Claims

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Application Information

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IPC IPC(8): C07K16/10C12N5/20G01N33/577G01N33/569G01N33/543C12R1/91
CPCC07K16/1018G01N33/577G01N33/56983G01N33/54346G01N2333/11G01N2469/10
Inventor 吴海波杨帆吴南屏姚航平
Owner THE FIRST AFFILIATED HOSPITAL ZHEJIANG UNIV COLLEGE OF MEDICINE