Japanese encephalitis vaccine prepared by human embryonic lung fibroblasts and preparation method thereof

A technology of fibroblasts and Japanese encephalitis, applied in the field of bioengineering, can solve the problems of inability to use human embryonic lung fibroblasts, and achieve the effects of quality control, safety assurance, and pollution avoidance

Active Publication Date: 2009-09-09
CHENGDU KANGHUA BIOLOGICAL PROD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0008] However, commonly used Japanese encephalitis strains P3, SA14-14-2 and Nakayama are not sensitive to human embryonic lung fibroblasts. Therefore, Japanese encephalitis vaccines have not been able to use human embryonic lung fibroblasts for a long time.
There is no report on the systematic use of human embryonic lung fibroblasts to produce Japanese encephalitis vaccine

Method used

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  • Japanese encephalitis vaccine prepared by human embryonic lung fibroblasts and preparation method thereof
  • Japanese encephalitis vaccine prepared by human embryonic lung fibroblasts and preparation method thereof
  • Japanese encephalitis vaccine prepared by human embryonic lung fibroblasts and preparation method thereof

Examples

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Embodiment 1

[0040] Use revived human embryonic lung fibroblast WI-38, the cell nutrient solution is MEM culture solution, which contains 8-10% calf serum and 24-32U / ml gentamicin, adjust the pH to 7.0-7.2, and then Cultured in a 15L spinner bottle at 37°C to form a dense monolayer and then amplified according to the seeding ratio of 1:8. When the expansion reached the production batch, after 6 days, the cells grew into a dense monolayer. Discard the cell nutrient solution and use pH7.0 Earle's solution of -7.4 washes the cell surface, inoculates the P3 virus seed of the virulent strain of Japanese encephalitis virus in human embryonic lung fibroblasts, and uses the third generation working virus seed (P3V) of the P3 strain on WI-38 cells. 3 ), virus seed concentration MOI 0.001, cell maintenance medium is MEM solution containing 0.2% (w / w) human albumin, its pH is 7.4-7.6, cultured at 33°C, after 24h, replace with fresh cell maintenance medium and continue to cultivate for 48- After 72 ho...

Embodiment 2

[0042] Use resuscitated human embryonic lung fibroblasts MRC-5, the cell concentration is 10 6 cells / ml, cultured in a 50L bioreactor with 5g / L Cytodex 2, working volume 40L, rotating speed 40-45 rpm, dissolved oxygen 25-50%. The cell nutrient solution uses MEM culture solution containing 8%-10% calf serum and 24-32U / ml gentamycin, pH 7.0-7.2, and the culture temperature is 37°C. Observe the growth of the cells at any time, adjust the pH of the culture solution according to the automatic replenishment of the reactor according to the growth of the cells, and continue culturing for 6 days until the cell density reaches 10 7 In the case of cells / ml, after washing twice with Earle's solution, add MEM solution without calf serum and containing 0.2% (w / w) human albumin, pH7.4-7.6. Then inoculate human embryonic lung fibroblast JE virus attenuated strain SA14-14-2 virus seed, use the third generation working virus seed of SA14-14-2 strain passed on MRC-5 cells, virus seed concentrat...

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Abstract

The invention discloses a Japanese encephalitis vaccine prepared by human embryonic lung fibroblasts and a preparation method thereof, comprising culture and expansion of the human embryonic lung fibroblasts. The method comprises the following steps: Japanese encephalitis virus strain P3, strain SA14-14-2 or strain Nakayama is naturalized and inoculated to fit the human embryonic lung fibroblasts, and the seeds of Japanese encephalitis viruses are prepared on the human embryonic lung fibroblasts; wherein, an inactivated Japanese encephalitis vaccine also comprises the steps of harvesting viruses, inactivating viruses, concentrating, purifying and the like; an attenuated live vaccine also comprises the steps of harvesting viruses, concentrating, purifying and the like. Due to being prepared by healthy human embryonic lung fibroblasts, the two kinds of Japanese encephalitis vaccines do not contain any adventitious pollution agent and tumorigenicity, has high purity after being purified and has the advantages of good immune effect and high security. The preparation method of the invention is suitable for large-scale industrial production and can meet the preparation processes of Japanese encephalitis vaccines required by domestic and abroad markets.

Description

technical field [0001] The invention relates to the field of bioengineering, in particular to an inactivated Japanese encephalitis vaccine and an attenuated live vaccine prepared from human embryonic lung fibroblasts, and a preparation process thereof. Background technique [0002] Japanese encephalitis is an infectious disease caused by acute viral infection of the central nervous system. The source of JE infection is infected people or animals, and then spread to other people or animals through mosquito bites. Pigs and mosquitoes are considered to be the main long-term storage hosts and diffusion hosts of the virus. The annual incidence of Japanese encephalitis in endemic areas is as high as 10-100 / 100,000. Nearly 3 billion people live in Japanese encephalitis endemic areas, mainly including Southeast Asia and the Western Pacific region, where more than 70 million newborns are born every year. Japanese encephalitis causes at least 50,000 clinical cases and 10,000 deaths i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/12A61K47/42A61K47/26A61P31/14
CPCY02A50/30
Inventor 蔡勇朱文漓
Owner CHENGDU KANGHUA BIOLOGICAL PROD
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