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Method for producing porcine reproductive and respiratory syndrome JXA1-R strain virus by using low serum cultured Marc-145 cells

A JXA1-R, low-serum technology, applied in the field of veterinary biology, can solve problems such as no, and achieve the effect of increasing production costs

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

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Problems solved by technology

After searching, there is no relevant literature report on the production of porcine reproductive and respiratory syndrome JXA1-R strain virus by culturing Marc-145 cells in low serum

Method used

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  • Method for producing porcine reproductive and respiratory syndrome JXA1-R strain virus by using low serum cultured Marc-145 cells
  • Method for producing porcine reproductive and respiratory syndrome JXA1-R strain virus by using low serum cultured Marc-145 cells
  • Method for producing porcine reproductive and respiratory syndrome JXA1-R strain virus by using low serum cultured Marc-145 cells

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Experimental program
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Embodiment 1

[0033] Embodiment 1: the method that low serum culture Marc-145 cell produces porcine reproductive and respiratory syndrome JXA1-R strain virus, it comprises the following steps:

[0034] S1. Subculture and culture of cells for preparation:

[0035] Take a T75 flask and culture a monolayer of Marc-145 cells, digest the cells with EDTA-trypsin cell dispersion solution, the EDTA-trypsin cell dispersion solution is a mixture of 0.15% trypsin and 0.02% EDTA in Hank's solution solution; blow and disperse the cells with cell growth solution, add 20ml of cell growth solution, and place Marc-145 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. The above-mentioned cell growth medium is DMEM culture medium with 10% serum content;

[0036] S2. Adapting cells to low serum medium: including the following sub-steps:

[0037] S21. Domestication of first generation cells:

[0038]Inoculate the Ma...

Embodiment 2

[0049] Embodiment 2: the method that low serum culture Marc-145 cell produces porcine reproductive and respiratory syndrome JXA1-R strain virus, it comprises the following steps:

[0050] S1. Subculture and culture of cells for preparation:

[0051] Take a T75 flask and culture a monolayer of Marc-145 cells, digest the cells with EDTA-trypsin cell dispersion solution, the EDTA-trypsin cell dispersion solution is a mixture of 0.15% trypsin and 0.02% EDTA in Hank's solution Blow and disperse the cells with cell growth medium, add 20ml of cell growth medium, place Marc-145 cells in a carbon dioxide incubator at a temperature of 37±2°C for 72 hours, and perform amplified culture when a good cell monolayer is formed. The above-mentioned cell growth medium is MEM culture medium with 10% serum content;

[0052] S2. Adapting cells to low serum medium: including the following sub-steps:

[0053] S21. Domestication of first generation cells:

[0054] The Marc-145 cells expanded and c...

Embodiment 3

[0065] Example 3: Effect of low serum medium on acclimating Marc-145 cells

[0066] Use T75 cell flasks, 37 ℃ static culture adapted to the fourth generation of Marc-145 cells in low serum medium, the ratio of serum addition is 1%, 2%, 3%, 4%, 5% five gradients, and the serum content used is set to The Marc-145 cells cultured in 10% DMEM was used as the control group, and the results were as follows: figure 1 , figure 2 , image 3 , Figure 4 , Figure 5 , Figure 6 See Table 1 for the indicators during the cell culture process.

[0067] Table 1: Effects of MD series low-serum medium on acclimatization of Marc-145 cells

[0068]

[0069] It can be seen from Table 1 that the average cell yield, cell activity and 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 porcine reproductive and respiratory syndrome JXA1-R strain virus by using low serum cultured Marc-145 cells, and belongs to the technical field of veterinary biology. The method comprises the following steps: S1, carrying out passage and culture on cells for preparation; S2, domesticating the cells to adapt to a low serum culture medium; S3, domesticating the cells to adapt to a low serum culture environment; and S4, reproducing a cytotoxic strain. By adopting the method for producing the porcine reproductive and respiratory syndrome JXA1-R strain virus by using the low serum cultured Marc-145 cells, disclosed by the invention, the production cost can be significantly reduced, and the downstream purification efficiency can also be improved; and moreover, the production scale can be expanded rapidly and stably, and balanced and stable quality can be easily achieved.

Description

technical field [0001] The invention belongs to the technical field of veterinary biology, and in particular relates to a method for producing porcine reproductive and respiratory syndrome JXA1-R strain virus by culturing Marc-145 cells with low serum. Background technique [0002] At present, the production technology of animal vaccines in my country is relatively backward. Most of the cells used in the production of viruses come from spinner bottle culture, and the most basic synthetic cell culture medium is mainly used, and even natural medium (such as hydrolyzed milk protein) is added with about 10% newborn bovine serum , The problems of vaccine safety, quality and cost brought about by these animal-derived ingredients have been highlighted. The main manifestations are: ①The large amount of bovine serum makes the growth of cells highly dependent on serum, but serum is a mixture of uncertain components, and there are differences in components between batches, which directl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12N5/073C12R1/93
Inventor 徐宏军胡来根牟和平岳丰雄田伟王洁清
Owner 成都史纪生物制药有限公司
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