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A method for producing porcine circovirus type 2 antigen by whole suspension cell culture

A porcine circovirus and cell culture technology, applied in the directions of virus antigen components, biochemical equipment and methods, antiviral agents, etc., to achieve the effects of increasing harvest times, reducing production costs, and easy quality control

Active Publication Date: 2017-04-26
JIANGSU NANNONG HI TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to overcome the deficiencies of the existing rotary bottle culture method and microcarrier suspension culture method, and provide a method for large-scale high-density production of porcine circovirus type 2 vaccine antigen using reactor full suspension culture technology

Method used

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  • A method for producing porcine circovirus type 2 antigen by whole suspension cell culture
  • A method for producing porcine circovirus type 2 antigen by whole suspension cell culture

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] ——Production of porcine circovirus type 2 antigen by whole suspension cell culture

[0040] 1. Preparation of porcine circovirus type 2 antigen (1) Cultivation and passage of seed cells in shake flasks

[0041] Seed cells PK-15B1 (CCTCC No.C200936, provided by Professor Jiang Ping of Nanjing Agricultural University) were cultured and grown in shake flasks, samples were taken for cell counting, and the cell density reached 2×10 6 When cells / ml, let the shake flask stand still, and after the suspended cells naturally settle to the bottom of the bottle, use a pipette to suck off 2 / 3 of the supernatant, gently blow and beat the cells repeatedly, and add the seed cell growth solution (the composition of the cell growth solution (V / V) is: low sugar DMEM88%, NBCS10%, dextran sulfate 1%, double antibody 1%), according to 2×10 5 The density of each / ml is equally divided into several shake flasks, and the culture is continued at 37°C and 100r / min;

[0042] (2) Inoculation of s...

Embodiment 2

[0057] ——Production of porcine circovirus type 2 antigen by whole suspension cell culture

[0058] 1. Preparation of porcine circovirus type 2 antigen

[0059] (1) Cultivation and passage of seed cells in shake flasks

[0060] Samples were taken for cell counting, and the cell density reached 2.5×10 6 cells / ml, let the shake flask stand still, and after the suspended cells naturally settle to the bottom of the bottle, use a pipette to suck off 2 / 3 of the supernatant, gently blow and beat the cells repeatedly, add the seed cell growth solution, and mix according to 4×10 5 The density of each / ml is equally divided into several shake flasks, and the culture is continued at 37°C and 100r / min;

[0061] (2) Inoculation of seed cells

[0062] Centrifuge the healthy cell suspension obtained from the aforementioned culture at a speed of 3000r / min for 5min, discard the supernatant, resuspend the cells with PCV2CGMCC No.2389 venom (3750ml) with 5% of the final volume of culture, and i...

Embodiment 3

[0078] ——The comparative results of porcine circovirus type 2 inactivated vaccine (SH strain) produced by three different cell culture processes, see Table 1

[0079] Table 1 Comparison table of different cell culture processes

[0080]

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Abstract

The invention relates to a method for culturing and producing a PVC 2 antigen through reactor whole suspension cells. The PVC 2 antigen is cultured and produced in a whole suspended and serum-free mode through a bioreactor, and an inactivated vaccine is prepared. The method has the advantages that production cost can be greatly reduced; compared with a spinner bottle process, the cost of the unit antigen is reduced by more than 90%, and the input-output ratio is increased by 4 times to 10 times; compared with a traditional reactor cultivation process, the cost of the unit antigen is reduced by 50% to 70%, and the yield is increased by 3 times to 5 times; no carriers are needed, serum residuals are not generated, the produced vaccine is higher in safety, the difference between batches of the produced antigen is small, quality is stable and easy to control, the yield of the produced antigen can be obviously increased, and the quality of the produced antigen can be obviously improved.

Description

technical field [0001] The invention relates to a method for producing porcine circovirus type 2 vaccine antigen by using a bioreactor full-suspension cell culture technology, belonging to the field of veterinary biological products. Background technique [0002] At present, domestic porcine circovirus type 2 (Porcine circovirus 2, PCV2) vaccine antigen production mainly adopts the spinner bottle culture method, and a few adopts the microcarrier suspension culture method. The semi-finished product antigen titer produced by the spin bottle process is low, only 10 4.5 ~ 6.0 TCID 50 / ml, unable to provide qualified antigen stably (the antigen content of qualified antigen per milliliter should be ≥10 5.5 TCID 50 ), requiring high concentration. Moreover, the spinner process is labor-intensive, and 100 to 200 large spinner bottles are required to produce a batch, requiring multiple people to carry out long-term digestion and passage operations at the same time, which increas...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/12A61P31/20C12N7/00
Inventor 何海蓉孙石静王正春董彦鹏何叶峰胡芳张志华缪芬芳刘怡姜冲
Owner JIANGSU NANNONG HI TECH