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Method for producing hog cholera C-strain virus by culturing ST Cells in low serum

A low-serum, cell-based technology, applied in the field of veterinary biology, can solve unforeseen problems and achieve the effects of reducing production costs, quickly stabilizing production scale, and expanding production scale

Inactive Publication Date: 2015-05-20
成都史纪生物制药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After searching, there is no relevant literature report on the production of CSF strain C virus by culturing ST cells with low serum

Method used

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  • Method for producing hog cholera C-strain virus by culturing ST Cells in low serum
  • Method for producing hog cholera C-strain virus by culturing ST Cells in low serum
  • Method for producing hog cholera C-strain virus by culturing ST Cells in low serum

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1: the method that low serum culture ST cell produces classical swine fever C strain virus, it may further comprise the steps:

[0037] S1. Subculture and culture of cells for preparation:

[0038] Take a T75 flask and culture it to cover a monolayer of ST cells, digest the cells with EDTA-trypsin cell dispersion, the EDTA-trypsin cell dispersion is a mixture of 0.15% trypsin and 0.02% EDTA in Hank's solution; Blow and blow dispersed cells with cell growth medium, add 20ml of cell growth medium, and culture ST cells in a carbon dioxide incubator at 37±2°C for 72 hours. When a good cell monolayer is formed, carry out amplified culture. DMEM culture fluid with 10% serum content;

[0039]S2. Adapting cells to low serum medium: including the following sub-steps:

[0040] S21. Domestication of first generation cells:

[0041] Inoculate the ST cells expanded and cultivated in step S1 into the first-generation low-serum medium for domestication, which is DMEM cul...

Embodiment 2

[0055] Embodiment 2: the method that low serum culture ST cell produces classical swine fever C strain virus, it may further comprise the steps:

[0056] S1. Subculture and culture of cells for preparation:

[0057] Take a T75 flask and culture a monolayer of ST cells, and digest the cells with EDTA-trypsin cell dispersion, which is a mixture of 0.15% trypsin and 0.02% EDTA in Hank's solution. Blow and blow dispersed cells with cell growth medium, add 20ml of cell growth medium, and culture ST cells in a carbon dioxide incubator at a temperature of 37±2°C for 72 hours. When a good cell monolayer is formed, carry out amplified culture. MEM medium with 10% serum content;

[0058] S2. Adapting cells to low serum medium: including the following sub-steps:

[0059] S21. Domestication of first generation cells:

[0060] Inoculate the ST cells expanded in step S1 into the first-generation low-serum medium for acclimatization, the first-generation low-serum medium is MEM culture fl...

Embodiment 3

[0074] Embodiment 3: Low serum medium acclimates ST cell effect

[0075] Use T75 cell flasks, 37°C statically culture the fourth-generation ST cells adapted to low serum medium, add serum in five gradients of 1%, 2%, 3%, 4%, and 5%, and set up a serum content of 10% The ST cells cultured in DMEM were used as the control group, and the results Such as figure 1 , figure 2 , image 3 , Figure 4 , Figure 5 , Image 6 As shown, the indicators in the process of cell culture are shown in Table 1 . (Merck MD series dry powder low serum medium including DMEM and MEM)

[0076] Table 1 : Effect of MD series low serum medium on acclimating ST cells

[0077]

[0078]

[0079] Depend on Table 1 It can be seen that the average harvested amount of cells, the average cell activity and the average specific growth rate of the experimental group were higher than those of the control group.

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Abstract

The invention discloses a method for producing a hog cholera C-strain virus by culturing ST Cells in low serum, and belongs to the technical field of veterinary medicine and biology. The method comprises the following steps: S1, preparing a cell subculture and culture solution; S2, domesticating cells to adapt to a low-serum culture medium; S3, domesticating cells to adapt to a low-serum culture environment; S4, breeding a cell virus strain; S5, breeding prepared venom. The method for producing the hog cholera C-strain virus by culturing ST Cells in low serum provided by the invention can remarkably reduce the production cost and can also improve the downstream purifying efficiency, and can quickly and stably expand production scale, so that the quality is easy to balance and stabilize.

Description

technical field [0001] The invention belongs to the technical field of veterinary biology, and in particular relates to a method for producing hog C strain virus by culturing ST cells with low serum. Background technique [0002] ST cells are pig testis (swine testis) cells, which grow adherently in vitro, proliferate slowly during culture, and tend to form clumps during digestion and passage. ST cells are sensitive to various viruses, such as classical swine fever virus (CSFV), porcine parvovirus (PPV), pseudorabies virus, etc. At present, the method of cultivating ST cells to produce vaccines mostly adopts the spinner bottle process, and uses DMEM and MEM medium to add 10% serum for culture. [0003] At present, whether it is the traditional spinner bottle culture technology or the advanced bioreactor suspension culture technology, the domestic production technology of swine fever vaccine adopts the high serum content cell culture technology. The traditional process has ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12N5/071C12R1/93
Inventor 徐宏军胡来根牟和平岳丰雄王洁清
Owner 成都史纪生物制药有限公司
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