Method for preparing Seneca virus by suspension cell line

A suspension cell and cell line technology, applied in the field of veterinary biological products, can solve problems such as difficult control and lack of vaccines, and achieve the effect of reducing batch-to-batch variation and increasing production capacity

Active Publication Date: 2019-12-13
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because there is no commercial vaccine for the time being, once the SVA epidemic occurs, it is difficu

Method used

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  • Method for preparing Seneca virus by suspension cell line
  • Method for preparing Seneca virus by suspension cell line
  • Method for preparing Seneca virus by suspension cell line

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] The domestication of the ST cell line of embodiment 1 adherence

[0045] Materials used in this example: adherent ST cell line, ATCC number: CRL-1746; basic medium: GibcoEMEMbasic, serum-free medium: Gansu Jianshun Biotechnology Co., Ltd. CDPK15259; virus: Seneca virus FJ / 001 strain (SVV / FJ / 001 strain), which was isolated and preserved in our laboratory, and its microorganism preservation number is: CCTCCNO:V201802.

[0046] First, the adherent ST cell line was domesticated into a suspension ST-S cell line, and the specific steps were as follows:

[0047] 1. Take out one sticky ST cell line from the liquid nitrogen tank, and quickly put it in a 37°C water bath to melt it, then transfer it to a 15ml centrifuge tube and add 5-10ml 10% FBSEMEM medium, centrifuge at 1000rpm Discard the supernatant after 5 minutes, and gently blow and beat the cell pellet with 10ml of complete medium containing 10% FBSEMEM, then place it in a Corning T25 ventilated culture bottle and place ...

Embodiment 3

[0063] The determination of embodiment 3 suspension ST-S cell line serum-free medium

[0064] In order to eliminate the influence of serum in the low-serum medium on the subsequent virus culture and reduce the cost for industrial production, the present invention screened a variety of serum-free medium to cultivate the suspension ST-S cell line. The medium screening comparison is as follows.

[0065]

[0066] Finally, the culture medium of Gansu Jianshun Biotechnology Co., Ltd. is selected, and the initial density of cell culture is 0.8~1.5×10 6 cell / ml, the cell density reaches 6.0~8.0×10 within 72 hours 6 cell / ml, culture temperature is 35~37℃, culture speed is 100~110rpm, CO 2 The concentration is 5%, and the Adolf Konai ISF1-X box-type temperature-controlled shaker is preferred for the culture shaker.

Embodiment 4

[0067] Example 4 Preparation of Seneca virus using suspension ST-S cell line

[0068] Materials used in this example: serum-free medium: Gansu Jianshun Biotechnology Co., Ltd. CDPK15259; virus: Seneca virus is FJ / 001 strain (SVV / FJ / 001 strain), which was isolated and preserved in this laboratory, and its microorganism The preservation number is: CCTCCN0:V201802; TPCK trypsin from bovine pancreas, purchased from sigma company, article number: 9002-07-7; ventilated Erlenmeyer shaker flask: Corning 125ml.

[0069] 4.1 Determination of the optimum pH for inoculation

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Abstract

The invention provides a method for preparing Seneca virus by a suspension cell line. The method comprises the steps as follows: (1) an adherent ST cell line is domesticated into a suspension ST-S cell line; and (2) Seneca virus is cultured and prepared by the suspension ST-S cell line obtained in the step (1). The Seneca virus is cultured by the domesticated suspension ST-S cell line, the capacity of the obtained virus is effectively improved, and a foundation is laid for large-scale mass production of Seneca virus vaccines.

Description

technical field [0001] The invention relates to a method for preparing Seneca virus by using a suspension cell line, in particular to a method for preparing Seneca virus by using a suspension culture subculture cell line, and belongs to the technical field of veterinary biological products. Background technique [0002] Senecavirus A (SenecavirusA, SVA), also known as Senecavirus (Senecavalleyvirus, SVV), is a new virus in the world and newly introduced into my country. Members, infected pigs cause blisters and ulcers on the nose and hoof crowns of the pigs, resulting in lameness, and the clinical symptoms are indistinguishable from those of foot-and-mouth disease. Before 2014, SVA only occurred sporadically in the United States and Canada, but since 2015, SVA outbreaks have occurred in Brazil, the United States, Thailand, China, etc., and have continued to spread and become more harmful. Since SVA was introduced into my country in 2015, epidemics have occurred in many provi...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N5/071
CPCC12N5/0683C12N7/00C12N2770/32051
Inventor 郑海学曹伟军杨帆朱紫祥魏婷郑敏蒋保余田宏张克山刘永杰茹毅党文刘湘涛
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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