Enterovirus 71 (EV71) monoclonal antibody and application thereof

A monoclonal antibody and virus technology, applied in the field of virus immunity biomedicine, can solve the problems of high test cost, long detection time, difficult to popularize and use, etc., and achieve the effect of strong guiding significance, high neutralization activity and so on.

Active Publication Date: 2012-04-11
SINOVAC BIOTECH
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Problems solved by technology

At the same time, the test cost is high, many large-scale instruments and equipment and skilled technicians are requ

Method used

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  • Enterovirus 71 (EV71) monoclonal antibody and application thereof
  • Enterovirus 71 (EV71) monoclonal antibody and application thereof

Examples

Experimental program
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Example Embodiment

[0039] Example 1 Establishment of anti-EV71 monoclonal antibody cell line

[0040] 1. Immunogen preparation: The immunogen is the EV71 inactivated virus (the virus was deposited at the General Microbiology Center (CGMCC) of the China Microbial Culture Collection and Management Committee on May 8, 2009, and the deposit number is CGMCC No.3062) inoculated with Vero cells , Obtained by culturing, inactivating and purifying.

[0041] 2. Animal immunization: immunize BALB / c mice with EV71 inactivated virus. For the first time, the mixed emulsification of Freund's complete adjuvant and EV71 was used to immunize animals according to methods known in the art; 10 days later, the mixed emulsification of Freund's incomplete adjuvant and EV71 was used to boost the immunization of mice; one week later, the serum anti-EV71 activity was measured by indirect ELISA. The mice with the highest antibody titer were immunized with EV71 purified liquid intraperitoneally.

[0042] 3. Cell fusion to establ...

Example Embodiment

[0044] Example 2 Preparation of monoclonal antibodies

[0045] BALB / c mice were intraperitoneally immunized with paraffin. The hybridoma cells were cultured, collected and washed once by centrifugation, resuspended in serum-free culture medium, adjusted the cell density, and injected intraperitoneally into mice sensitized for 7 days. After 7-12 days of inoculation, the abdominal cavity of the mice was obviously swollen. The ascites was collected with a syringe, and the ascites was collected by centrifugation at 1500 rpm for 10 minutes. The monoclonal antibody cell line was established and stored in liquid nitrogen.

Example Embodiment

[0046] Example 3 Determination of the specificity of monoclonal antibodies

[0047] The A9 monoclonal antibody purified by ammonium sulfate was used to coat the ELISA plate, and the EV71 inactivation solution of three genotypes A, B, and C, Cox A16 virus, hepatitis A virus, Japanese encephalitis virus, varicella virus, etc. were added, and labeled with HRP The rabbit anti-EV71 polyclonal antibody is a labeled antibody for antigen detection. The results showed that only the EV71 virus reacted, and the monoclonal antibody did not cross-react with other enteroviruses and other hand, foot and mouth viruses such as Cox A16, and the specificity was good.

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Abstract

The invention provides an enterovirus 71 (EV71) monoclonal antibody and an application thereof. The monoclonal antibody is derived from an EV71 hybrid tumor cell strain, which has a preservation number of CGMCC (China General Microbiological Culture Collection) No.5330. The monoclonal antibody and related reagents thereof have important significance in promoting the treatment and research of severe cases and death cases caused by EV71 in China, and have extremely important value in further promoting clinical research, epidemic development and passive treatment of EV71.

Description

technical field [0001] The invention relates to the field of virus immune biomedicine, in particular to an enterovirus 71 monoclonal antibody and its application. Background technique [0002] Hand, foot and mouth disease (Hand, foot and mouth diseases, HFMD) is a fecal-oral disease that is widely prevalent in the world, and it has been reported in most parts of the world. For a long time, mild hand, foot and mouth disease broke out periodically all over the world. The disease was first reported in New Zealand in 1957, the Coxsackie virus was isolated for the first time in 1958, and the name of hand, foot and mouth disease was proposed in 1959. In 1969, enterovirus 71 (EV71 virus) was isolated from stool samples of infants with central nervous system diseases in the United States, and the virus was first confirmed. The interval between the prevalence of HFMD is 2-3 years. The early pathogens were mainly Cox A16, and later EV71 and Cox A16 appeared alternately, becoming the...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/10G01N33/577A61K39/42A61P31/14C12R1/91
Inventor 蔡芳高强王琳薛晨宝张立志尹卫东
Owner SINOVAC BIOTECH
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