Monoclone antibody of swine fever virus resistant wild strain E2 protein, preparation method and application thereof

A monoclonal antibody and swine fever virus technology, applied in hybridoma cell lines, the application of monoclonal antibodies in clinical diagnosis of swine fever, the field of monoclonal antibodies, can solve the problems of differentiation, limited application, difficult natural infection, etc.

Inactive Publication Date: 2008-10-29
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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Problems solved by technology

The biggest disadvantage of conventional attenuated vaccine immunization is that it is difficult to distinguish it from natural infection by serological methods
Scholars at home and abroad have prepared a variety of CSFV monoclonal antibodies (Wensvoort G, Terpstra C, Boonstra J, et al. Production of monoclonal antibodies against swine fever virus and their use in laboratory diagnosis. Vet Microbiol, 1986, 12 (2): 101 -108; Qiu Huishen, Wang Zaishi, Fang Guoan. Research on Diagnostic Reagents of Monoclonal Antibody of Swine Fever. China Livestock and Poultry Infectious Diseases, 1991, 6: 20-26; 1991, 6: 20-26; Ma Gang, Li Zuosheng, Yu Xing Long et al. Preparation of monoclonal antibody against classical swine fever virus E2 protein and preliminary analysis of its epitope. Chinese Journal of Veterinary Medicine, 2002, 22(2):

Method used

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  • Monoclone antibody of swine fever virus resistant wild strain E2 protein, preparation method and application thereof
  • Monoclone antibody of swine fever virus resistant wild strain E2 protein, preparation method and application thereof
  • Monoclone antibody of swine fever virus resistant wild strain E2 protein, preparation method and application thereof

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Embodiment 1

[0020] The preparation of embodiment 1 monoclonal antibody

[0021] 1. Purification and detection of recombinant protein

[0022] Sf9 cells in the logarithmic growth phase were infected with the 4th generation recombinant baculovirus rAcV-Shimen-E2 and rAcV-HCLV-E2 respectively, cultured at 27°C for 96h, and the supernatant of the cell culture medium was collected, centrifuged at 3000r / min, 4°C for 10min , to remove cell debris; then put the supernatant in a dialysis bag and concentrate it by embedding with PEG 8000; the concentrated supernatant was placed in Ni 2+ Ion affinity chromatography equilibrium solution (50mmol / L Na 3 PO 4 , 500mmol / LNaCl, pH7.0) and dialyzed at 4°C overnight. Then mix the dialyzed supernatant with the balanced resin, gently shake at 4°C for 1 hour, elute with 250mmol / L imidazole, collect the eluate, conduct SDS-PAGE electrophoresis analysis, and then transfer the target protein On the nitrocellulose membrane, with the anti-E2 protein monoclonal ...

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Abstract

The invention discloses a monoclonal antibody against virulent strain E2 protein of classical swine fever virus and a hybridoma cell strain secreting the monoclonal antibody. The hybridoma cell strain is obtained by using hog cholera lapinized virus vaccine strain E2 protein expressed by Baculovirus as tolerogen, selecting Shimen strain E2 protein as immunogen, immunizing mouse by cyclophosphamide immunosuppression method, carrying out cell fusion, and sieving hybridoma cell strain capable of stably secreting monoclonal antibody against E2 protein. The monoclonal antibody can react with Shimen strain and can produce specific reaction with virulent strain of classical swine fever viruses of 1.1, 2.1, 2.2 and 2.3 gene sub-groups. The monoclonal antibody has neutralization activity and does not react with hog cholera lapinized virus vaccine strain, so that the monoclonal antibody can be used for differentiating virulent strain of classical swine fever virus and hog cholera lapinized virus vaccine strain, which establishes the foundation for establishing a method for differentiating wild virus infection of classical swine fever and vaccine immunity and for researching the molecular difference between CSFV virulent strain and mild strain.

Description

technical field [0001] The present invention relates to a monoclonal antibody, in particular to a monoclonal antibody against the E2 protein of the wild strain of classical swine fever virus, and the present invention also relates to a hybridoma cell strain secreting the anti-E2 protein monoclonal antibody and the anti-E2 protein monoclonal antibody and the monoclonal antibody in classical swine fever The application in clinical diagnosis belongs to the field of animal disease inspection and quarantine. Background technique [0002] Classical swine fever (CSF) is a highly contagious infectious disease characterized by high fever and hemorrhage caused by classical swine fever virus (CSFV), and is one of the major infectious diseases that endanger the pig industry , the World Organization for Animal Health (OIE) included it in the OIE List of Diseases (OIE Listeddiseases), and my country also listed it as a first-class infectious disease. CSFV belongs to the genus Pestivirus o...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/10G01N33/577
Inventor 仇华吉夏照和彭伍平成丹侯强王万利孙元涂长春
Owner HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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