Method for culturing gosling plague virus by use of goose embryo continuous cell line and bioreactor

A bioreactor and gosling plague virus technology, which is applied in the field of veterinary biological products, can solve the hidden dangers of embryonic body-derived exogenous virus contamination of biological safety, the cumbersome process of primary fibroblasts, and the treatment of embryonic body waste to disperse poison, etc. It can improve the virus titer, improve the controllability, and eliminate potential safety hazards.

Active Publication Date: 2014-12-17
浙江美保龙生物技术有限公司
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  • Abstract
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Problems solved by technology

The direct use of embryoid bodies for virus propagation is easily limited by the source of non-immune embryoid bodies. At the same time, the use of embryoid bodies for virus propagation, because the cultured virus suspension contains a large amount of miscellaneous proteins will bring safety hazards to subsequent vaccine preparations. In addition, the embryo body waste disposal Improper use also presents the danger of poisoning
Although the spinner bottle primary cell culture technology is relatively mature, the process of preparing primary fibroblasts is cumbersome, and there are large differences between batches. The amount of cultured virus is small and the toxicity is low. Security risks

Method used

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  • Method for culturing gosling plague virus by use of goose embryo continuous cell line and bioreactor
  • Method for culturing gosling plague virus by use of goose embryo continuous cell line and bioreactor

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Experimental program
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Effect test

Embodiment 1

[0025] Embodiment 1 Cultivate and propagate gosling plague virus on the AP20C type rapid flow perfusion reactor with goose embryo passage cell line, comprising the following steps:

[0026] (1) Selection of bioreactor: Rapid flow reactor type AP-20C with a volume of 10L.

[0027] (2) Selection of cells: goose embryo passage cell line CGBQ, a passage cell line suitable for the growth of gosling plague virus, non-carcinogenic, purchased from the American Type Culture Collection.

[0028] (3) Selection of virus strains: gosling plague virus SHM 319 strain was selected, microbial deposit number: ATCC VR-696, preserved in the American Type Culture Collection, and can be purchased from the American Type Culture Collection.

[0029] (4) Cultivation of species of cells: Propagate CGBQ cells with the cell growth medium in T175 square flasks, generally passaged at a ratio of 1:3 to 1:5, digest and count the cells after the cells have grown into a monolayer, and inoculate the cell suspen...

Embodiment 2

[0039] Embodiment 2 and the comparison of spinner bottle culture

[0040] (1) Spinner flask cell culture and virus propagation

[0041] Spinner bottle cell culture: culture CGBQ cells on the spinner bottle, after the monolayer grows, the number of cells is 5×10 8 a / L. Rotary bottle virus propagation: 30 hours after CGBQ cells were inoculated with GPV, 80% of the cells showed pathological changes, and the virus titer was 10 -7.0 / 0.1ml.

[0042] (2) The comparison results of CGBQ breeding GPV in spinner bottle and reactor culture are shown in Table 1.

[0043] Table 1 Comparison of reactor and spinner bottle results

[0044]

[0045] Cultivate goose plague virus CVCC AV240 strain, CVCC AV239 strain, NEAU0671 strain or other goose plague virus strains by the method described in the present invention, and finally the same technical effect as the present invention can be achieved.

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Abstract

The invention discloses a method for culturing gosling plague virus by use of a goose embryo continuous cell line and a bioreactor, belonging to the technical field of veterinary biological products. The method comprises the following steps of: 1) culture of master cells; 2) suspension culture of cells for virus reproduction; and 3) culture and harvesting of virus. The method disclosed by the invention can shorten the production period and reduce personnel allocation, and realizes low pollution probability and small floor area; and the obtained virus has high titer and small inter-batch difference.

Description

technical field [0001] The invention belongs to the technical field of veterinary biological products, and in particular relates to a method for cultivating gosling plague virus using a goose embryo subculture cell line and a bioreactor. Background technique [0002] Gosling plague (Gosling Plague, GP) is an acute or subacute septic infectious disease of goslings caused by Gosling Plague Virus (GPV). The disease mainly affects goslings and muscovy ducks within 30 days after hatching. It has the characteristics of rapid transmission, high morbidity and mortality, and the mortality rate can reach 90% to 100%. Fang Dingyi first discovered the disease in Yangzhou, my country in 1956, and isolated the virus from goose embryos in 1961. After 1965, the existence of the disease was successively reported in Hungary, Germany, the Netherlands, the former Soviet Union, Italy, the United Kingdom, France, Yugoslavia, Vietnam, Israel and other countries. At present, there are outbreaks a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00C12R1/93
Inventor 李阳闫艳丽刘雪徐晓艳王二先
Owner 浙江美保龙生物技术有限公司
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