Method for culturing porcine circovirus type 2 by microcarrier bioreactor

Abstract

The invention discloses a method for culturing porcine circovirus type 2 by a microcarrier bioreactor. The method comprises the following steps: (1) cell dissociation in a spinner bottle; (2) cell filtration; (3) cell virus receiving and tank charging culture; (4) 24 h liquid exchange; (5) gaining of a primarily received antigen; (6) second-time cell dissociation and tank charging; (7) gaining ofa secondarily received antigen and liquid exchange culture; (8) gaining of a thirdly received antigen; and (9) melting of the primarily received antigen, the secondarily received antigen and the thirdly received antigen, filtration to remove a microcarrier to obtain an antigen liquid for titer detection. By adoption of the method, the cell wall-adhering efficiency can be improved, cell aggregatesare reduced, the cell density is improved, the yield of single-batch antigens can be increased, the titer of the antigens is improved, and the production cost is lowered.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for cultivating porcine circovirus type 2 in a microcarrier bioreactor. Background technique [0002] The culture method of porcine circovirus type 2 is generally a spinner bottle culture and a microcarrier bioreactor culture. [0003] The spinner bottle culture process is generally as follows: use spinner bottle culture cells, after a period of culture in spinner bottle cells, use enzyme digestion solution to digest, subculture and amplify the culture, and at the same time carry out synchronous inoculation. Connect it to a spinner bottle for cultivation, change the medium after cultivating for a period of time, and continue to cultivate until 72-96h to harvest the virus liquid. [0004] This method has the following disadvantages: due to the limited surface area of ​​the spinner bottle cultured cells, the cell density is low when the cells are cultured, and the...

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Problems solved by technology

[0004] This method has the following disadvantages: due to the limited surface area of ​​the cells cultured in the spinner bottle, the cell density is low when the cells are cultured, and the titer of the virus cultured after inoculation is also relatively low

[0006] There are following deficiencies in this method: 1, the cells are digested from the spinner bottle, and there are many cell clumps, and the cells are unevenly adhered to the wall in the microcarrier, and many clumps are attached to the microcarriers, and the clumps in the process of cultivating Cells are easy to die or fall off early, resulting in insufficient cell density and low titer of antigen harvested

2. The culture of circovirus is generally inoculated synchronously. After 48 hours of inoculation, the cells begin to shed a large number of cells. After 48 hours, due to the large number of shedding cells and insufficient virus reproduction, the titer of the antigen finally harvested is not high.

3. When harvesting the antigen, only the supernatant is harvested. Since there are still some infected cells on the carrier that cannot be shed during harvesting, a part of the virus is lost, resulting in a low titer of the harvested antigen.

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