Method for preparing inactivated Japanese encephalitis vaccine by bioreactor

A bioreactor, porcine encephalitis technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganisms, etc., can solve the problems of difficulty in improving the vaccine quality of antigen content, high production costs, and obvious side effects, etc. To achieve the effect of improving oxygen transfer mode, low cost and controllable quality

Inactive Publication Date: 2013-03-06
TANGSHAN YIAN BIOLOGICAL ENG CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the inactivated JE vaccine is safe and reliable, it has high production costs, a large number of irrelevant antigenic components, and obvious side effects
Although the attenuated vaccine has good protection, there may be unsafe factors such as potential virulence reversion, poisoning, detoxification, and verti...

Method used

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  • Method for preparing inactivated Japanese encephalitis vaccine by bioreactor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Example 1 Vero cell subculture expansion culture adapted to the Japanese encephalitis virus P3 strain

[0015] Dilute the JE virus M53 generation P3 strain JE virus seeds with standard MEM medium, inoculate suckling mice in the brain, collect dead mouse brain tissue, grind to make brain tissue suspension, centrifuge, and take the supernatant to make Japanese encephalitis mouse brain virus.

[0016] Properly dilute the prepared Japanese encephalitis virus mouse brain suspension with cell maintenance solution, inoculate Vero cells that grow into a single layer, and culture the virus at 34.0±2.0°C, and harvest when the cytopathy reaches about 75% For the virus supernatant, according to the above method, the P3 strain Japanese encephalitis virus was continuously passaged in Vero cells, and the virus content was measured at 10 7.0 More than PFU / ml. It shows that the Japanese encephalitis virus P3 strain is sensitive to Vero cells and can be used for vaccine production.

Embodiment 2

[0017] Embodiment 2 utilizes bioreactor and Cytodex-1 microcarrier system to carry out Vero cell culture

[0018] The working volume of the bioreactor is 10L-500L, the Cytodex-1 microcarrier system uses 10g / L, and the cell density ranges from 5×10 5 Increase to 5~7×10 6 .

Embodiment 3

[0019] Example 3 Inoculation of Japanese encephalitis virus P3 strain to carry out viral infection of Vero cells

[0020] When the cells grow into a monolayer, carry out the P3 strain Japanese encephalitis virus infection according to inoculation 0.005~0.5M.O.I, according to the concentration of glucose, lactate concentration, valley Aminoamide, oxygen consumption rate and pH value indicators are used to control; virus culture temperature is controlled at 34.0±2.0°C, pH value is 7-8, 20-60 rpm, and dissolved oxygen is 10-100%.

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Abstract

The invention provides a method for preparing inactivated Japanese encephalitis vaccine by a bioreactor. The method includes by the aid of the bioreactor and a Cytodex-1 microcarrier system, subjecting high-density Vero cells adaptive to Japanese encephalitis virus P3-strain to virus infection by utilizing the P3-strain Japanese encephalitis virus with good antigenicity; according to multiplication conditions of the Japanese encephalitis virus P3-strain on the Vero cells, harvesting virus liquid after the cytopathic effect reaches a certain degree so as to increase virus titer from 105-6PFU/ml to 107-8PUF/ml at least and increase virus content by 10 times at least; and inactivating harvest virus liquid, adding additives, and then packing to obtain high-quality vaccine products. The prepared vaccine is safe and effective, production process is stable, and production quality is controllable.

Description

technical field [0001] The invention relates to a vaccine preparation method, in particular to a method for preparing porcine Japanese encephalitis inactivated vaccine using a bioreactor. Background technique [0002] Porcine Japanese encephalitis, also known as Japanese Japanese encephalitis, is a mosquito-borne zoonotic infectious disease caused by the Japanese Japanese encephalitis virus. Mainly through the bite of blood-sucking insects (mainly mosquitoes), the spread of the virus is expanded through the pig-mosquito-pig cycle, making it the main host and source of infection for Japanese encephalitis virus. Pigs of various ages, breeds, and sexes are susceptible to the disease, and diseased breeding pigs can be transmitted vertically to piglets. The disease is prevalent in summer and autumn, reaching its peak in June to July in the south and August to September in the northeast. It exists widely in large-scale pig farms in my country and is endemic. Beijing, Yunnan, Hun...

Claims

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Application Information

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IPC IPC(8): A61K39/12C12N7/00A61P31/14C12R1/93
CPCY02A50/30
Inventor 岳雷郭鑫陈静李淑芬韩中山张振山
Owner TANGSHAN YIAN BIOLOGICAL ENG CO LTD
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