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Application of human embryonic lung fibroblast strains in preparation of rabies inactivated vaccine

A fibroblast and inactivated vaccine technology, applied in the application field of human embryonic lung fibroblast cell line in the preparation of rabies inactivated vaccine, can solve the problem of inability to guarantee cell matrix tumorigenesis and cell DNA, difficult to cultivate cells, and difficult to spread Universal use and other issues, to achieve clear property rights of genetic resource stakeholders, clear genetic background information, and facilitate the production scale

Inactive Publication Date: 2013-12-25
云南沃森生物技术股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there are four kinds of rabies vaccines mass-produced in the world: the first is the diploid cell rabies vaccine represented by the United States, but the cells are difficult to cultivate, and the generation is high, and more importantly, the cost of the vaccine is high, and it is difficult to be widely used; A freeze-dried and inactivated vaccine represented by French Vero cells as a culture substrate, but the tumorigenicity and cellular DNA of the cell substrate cannot be guaranteed
The third and fourth types of hamster kidney cells represented by China and chicken embryo and duck embryo vaccines from Japan cannot guarantee that the cells do not carry exogenous factors, nor can they guarantee the tumorigenicity of the cell matrix because the cells are derived from common animals. sex and cellular DNA, and the availability of animal organs are limiting factors for industrial production
Chinese patent 201210371784.X discloses a human embryonic lung diploid fibroblast cell line Walvax-2, but it is not clear whether this cell is sensitive to rabies virus, and it is not known whether this cell can be applied to the production of rabies vaccine
But the disadvantage is low yield and high cell generation, which limits the improvement of virus yield and further use
More importantly, the virus titer of rabies virus cultured on existing diploid cells is relatively low, and it can only be cultured in a limited number of cell flasks, making it difficult to implement large-scale industrial production

Method used

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  • Application of human embryonic lung fibroblast strains in preparation of rabies inactivated vaccine
  • Application of human embryonic lung fibroblast strains in preparation of rabies inactivated vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Preparation of embodiment 1 Walvax-2 cell strain

[0036]1.1. Acquisition of experimental materials: After obtaining the approval of the national health management department and the ethics committee, investigate the age, occupation and health status of the couples who donate pregnant women, as well as the three generations of parents of the fetus. Fetuses with a family history of genetic defects. After obtaining the consent of the parents of the fetus and their relatives, sign the "Donor Consent".

[0037] 1.2. Primary culture: embryos sent to the laboratory in time after induction of water sac, the lung tissue was taken out under sterile conditions, the membrane on the surface of the lung tissue was torn off, and cut into 1-2 mm 3 The tissue pieces were washed with PBS for 3 to 4 times; the tissue pieces were transferred into the Erlenmeyer flask; digested with trypsin for 30 minutes, neutralized with serum, centrifuged at 1000rpm for 5 minutes to discard the trypsin...

Embodiment 2

[0043] Embodiment 2 Adaptation of rabies virus CTN-1V5 virus and aG virus on cell Walvax-2 respectively

[0044] 2.1 Resuscitate Walvax-2 cells by conventional methods, use MEM containing 10% calf serum to subculture to 25-30 passages, grow into a monolayer, digest, disperse into single cells, add complete culture medium, centrifuge and remove the supernatant, the cells Suspending the sediment with serum-free medium to obtain a suspension of human embryonic lung diploid cells dispersed as single cells;

[0045] 2.2 Inoculate the fresh rabies virus CTN-1V5 seed into the human embryonic lung diploid cell suspension dispersed into single cells at 0.0001-1 MOI, absorb at 36°C for 90 minutes, and add cell culture medium to a suitable volume of 8 ml , cultivate at 30-40°C, form a dense monolayer in 2-5 days, discard the cell culture medium, replace with MEM maintenance solution containing 5% calf serum and 1% sucrose solution, maintain for 5 days, harvest the supernatant, and add 5-...

Embodiment 3

[0055] The rabies inactivated vaccine effect comparison prepared by embodiment 3 Walvax-2 cells and MRC-5 cell lines

[0056] The rabies inactivated vaccine prepared by Walvax-2 cell line is compared with the vaccine effect prepared by MRC-5 cells, and the method of Example 1 is used to prepare rabies inactivated vaccine on MRC-5 cells, and there are the following differences.

[0057] Table 2: Comparison of the effects of inactivated rabies vaccines prepared by Walvax-2 cells and MRC-5 cell lines

[0058] difference

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Abstract

The invention discloses an application of human embryonic lung fibroblast strains Walvax-2 in the preparation of a rabies inactivated vaccine. The human embryonic lung fibroblast strains Walvax-2 are susceptible to rabies viruses, and when the human embryonic lung fibroblast strains are used for producing the rabies virus vaccine, the diploid cell rabies virus inactivated purified vaccine for people can be obtained, wherein the vaccine is high in antigen purity, good in immune effect, high in safety and low in price.

Description

[0001] Field of the invention: [0002] The invention belongs to the technical field of vaccines, in particular, the invention relates to the application of human embryonic lung fibroblast strains in the preparation of rabies inactivated vaccines, and the application of human embryonic lung fibroblast strains in the preparation of inactivated vaccines for treating rabies Background technique: [0003] Rabies is a zoonotic natural foci disease caused by rabies virus infection. Rabies is the infectious disease with the highest death toll and case fatality rate in China's legal reports. According to the 2003 World Rabies Survey Report, the number of cases in China is second only to India, ranking second in the world. The Chinese Ministry of Health has listed rabies as a second-class infectious disease for management, but in recent years, the rabies epidemic in China has been on the rise. The epidemic situation is very serious. There is no effective treatment for rabies, and safe...

Claims

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Application Information

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IPC IPC(8): C12N7/00A61K39/205A61P31/14C12R1/93
Inventor 杨喆马波李伟何丽芳王丽丽陈敏王馨尹孝兵任正勇高显专胡超
Owner 云南沃森生物技术股份有限公司
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