Novel method for preparing porcine circus-virus 2 type

A porcine circovirus, a new method of technology, applied in microorganism-based methods, biochemical equipment and methods, viruses/phages, etc., can solve high-tech, complex and other problems, reduce operating procedures, simplify operations, and shorten time. Effect

Inactive Publication Date: 2014-04-23
TECON BIOLOGY CO LTD
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Problems solved by technology

After the cells were inoculated with PCV2, they were cultured at 37°C for 3-4 days, and the virus titer reached 107.0 TCID50 / ml; the above methods all produced relatively high-titer viruses, but it still required complicated operations and a high technical level to complete

Method used

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  • Novel method for preparing porcine circus-virus 2 type
  • Novel method for preparing porcine circus-virus 2 type
  • Novel method for preparing porcine circus-virus 2 type

Examples

Experimental program
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Embodiment

[0027] 1) Preparation of PK-15 cells

[0028] will contain a recovery density of 1.0 x 10 6 Place a container bottle of 1.8ml of PK-15 cryopreserved cells in a 37°C water bath, shake and dissolve for 2min, centrifuge at 1000rmp for 10min, discard the supernatant, add 15ml of cell culture medium, and transfer to a 75cm 2 culture flask at 37°C, 5% CO 2 In the incubator, cultivate for 72 hours, digest the cells with 0.25% EDTA-trypsin 5ml, and then dilute to 1.0×10 with 5ml DMEM cell culture medium (containing 10% fetal bovine serum). 6 Centrifuge the PK-15 cell suspension at 1000rmp for 10min, discard the supernatant, add 13ml cell culture medium, and transfer to 75cm 2 culture flask at 37°C, 5% CO 2 In an incubator, cultivate for 72 hours to obtain the PK-15 cells to be prepared;

[0029] 2) Preparation of virus culture medium

[0030] Add 50 mL of fetal bovine serum to 950 mL of DMEM culture solution, mix well, and then add 20 mL of D-glucosamine solution with a final con...

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Abstract

The invention provides a novel method for preparing porcine circus-virus 2 type, wherein inoculated PK-15 cells are treated by D-glucosamine with different concentration gradients, and is screened by IFA; the method not only makes the OK-15 cells normal grow, but also substantially raises PVC2 virus titer; the PVC2 virus titer acquired by the method is not less than 105TCID50, and a method for propagating and operating PVC 2 by screened proper D-glucosamine is used for researching a PVC2 vaccine and a relative diagnostic reagent, and preparing the products.

Description

technical field [0001] The invention relates to the preparation of porcine circovirus. The new method for preparing porcine circovirus type II not only saves time, but also reduces pollution, improves virus titer, and is beneficial to the improvement of yield. Background technique [0002] Apply conventional synchronous inoculation or asynchronous inoculation for virus culture, then add D-glucosamine solution for treatment, after D-Hanks cleaning, add virus maintenance solution to continue culturing, not only the process is complicated, but also high concentration of D- Glucosamine causes lethal damage to cells, thereby affecting the proliferation of viruses on cells. At present, the vaccine made of PCV2 antigen has achieved initial success in the prevention of PMWS, but the difficulty in the production of PCV2 whole-virus inactivated vaccine is that the virus titer is low, which cannot meet the requirements for the production of whole-virus inactivated vaccine. The culture...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12R1/93
Inventor 张伟潘晓梅师小潇李延涛贺笋刘小兰
Owner TECON BIOLOGY CO LTD
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