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Method of preparing hepatitis A inactivated vaccine

A technology for inactivated vaccines and hepatitis A, applied in biochemical equipment and methods, pharmaceutical formulas, and resistance to vector-borne diseases, etc., can solve the problem that hepatitis A vaccines cannot meet the domestic market demand, and improve vaccine production capacity and increase Virus yield, the effect of achieving large-scale production

Active Publication Date: 2011-05-18
SHENZHEN KANGTAI BIOLOGICAL PROD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

my country has a large population, and the hepatitis A vaccine is far from meeting the domestic market demand

Method used

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  • Method of preparing hepatitis A inactivated vaccine
  • Method of preparing hepatitis A inactivated vaccine
  • Method of preparing hepatitis A inactivated vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Human embryonic lung diploid cells (MRC-5) cultured in a monolayer cell factory were taken, the cells were digested with 0.125w / v% trypsin-EDTA, and MEM cell culture medium containing 10% calf serum was added. Wherein, the MEM is 0.03% Glu, 0.08% NaHCO 3 , 0.01% penicillin and 0.01% streptomycin. MEM was purchased from Beijing Qingda Tianyi Biotechnology Co., Ltd., production batch number: 080302.

[0030] Make the cell concentration 1-1.5 million cells per milliliter, add hepatitis A virus to the cell solution at 0.05-0.1 MOI, and carry out mixed adsorption at 20-30°C for 30-90 minutes. Dilute the mixed and adsorbed cell-virus mixture 10 times with MEM cell culture medium containing 10% calf serum, then inoculate it at 0.5 MOI in cell factories on layers 2-40 to proliferate hepatitis A virus, and culture it statically at 35°C±0.5°C for 21 -24 days, during which the cell maintenance solution was changed every 7 days (the cell maintenance solution was MEM cell culture ...

Embodiment 2

[0036]Human embryonic lung diploid cells (MRC-5) cultured in a monolayer cell factory were taken, the cells were digested with 0.125w / v% trypsin-EDTA, and MEM cell culture medium containing 10% calf serum was added. Wherein, the MEM is 0.03% Glu, 0.08% NaHCO 3 , 0.01% penicillin and 0.01% streptomycin. MEM was purchased from Beijing Qingda Tianyi Biotechnology Co., Ltd., production batch number: 080302.

[0037] Make the cell concentration 1-1.5 million cells per milliliter, add hepatitis A virus to the cell solution at 0.05-0.1 MOI, and carry out mixed adsorption at 20-30°C for 30-90 minutes. Dilute the mixed and adsorbed cell-virus mixture 10 times with MEM cell culture medium containing 10% calf serum, then inoculate it at 0.5 MOI in cell factories on layers 2-40 to proliferate hepatitis A virus, and culture it statically at 35°C±0.5°C for 21 -24 days, during which the cell maintenance solution was changed every 7 days (the cell maintenance solution was MEM cell culture s...

experiment example 3

[0042] The Hepatitis A vaccine prepared in Example 1 and Example 2 of the present invention was subjected to an in vivo efficacy test in mice, the endpoint dilution was calculated, and the immunogenicity was observed.

[0043] km mice (purchased from the Institute of Pathogenic Microbiology, Chinese Academy of Military Medical Sciences), male and female, weighing 16-18 g, were randomly divided into groups. Test is divided into 12 groups, every group is 10, and test vaccine 8 groups: inject respectively the vaccine that embodiment 1 and embodiment 2 prepare; , Ai Basu: Product batch number: 3001424.02). The test vaccine and the reference vaccine were serially diluted 4 times with the vaccine diluent, totaling 4 dilutions: original times, 1:4, 1:16, 1:64. Each mouse was intraperitoneally injected with 1.0ml, and after 4 weeks of immunization, blood was collected from the inner canthus of the eye, the serum was separated by centrifugation, and stored at -20°C for future use. Th...

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Abstract

The invention provides a method of preparing a hepatitis A inactivated vaccine, which comprises the following steps of inoculating mixed and absorbed hepatitis A virus strain SH and a human embryo diploid cell MRC-5 to hepatitis A virus propagated in a cell factory, digesting the cell in the virus propagation peak to obtain cell virus liquid, removing impurity proteins of the cell by ultrasonication, chloroform extraction and ultrafiltration through ultrafiltration membranes, degerming and filtering by gel filtration chromatography and purification, and absorbing by an aluminium hydroxide adjuvant so that the hepatitis A inactivated vaccine is prepared. The result of in vivo effectiveness experiments performed on a mouse shows that the hepatitis A inactivated vaccine prepared by the method of the invention has higher ED50 and better immunogenicity than the contract strain. The method of the invention can improve the safety of the vaccine, simplify the technique, shorten the productionperiod, reduce the production cost, and realize the scale production of the hepatitis A inactivated vaccine.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a new method for preparing an inactivated hepatitis A vaccine. Background technique [0002] Hepatitis A, referred to as hepatitis A, is an acute infectious disease caused by hepatitis A virus that seriously endangers human health. Hepatitis A is mainly transmitted through the "fecal-oral" route, or transmission among individuals, or outbreaks of hepatitis A are caused by water or food contaminated with hepatitis A virus. After older children and adults are infected with hepatitis A, more than 70% are clinical infections, and the case fatality rate is 0.3% to 0.6%; the case fatality rate of patients over 50 years old is 1.8%. After chronic liver disease is infected with hepatitis A, the risk of acute liver failure raised. With the improvement of living conditions, the number of adults infected with hepatitis A tends to increase, and the proportion of clinical hepatitis A increases,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/29C12N7/00A61P31/14
CPCY02A50/30
Inventor 张现臣魏文进黄秋香钟汉斌孟红彦王春雨
Owner SHENZHEN KANGTAI BIOLOGICAL PROD
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