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Method for preparing pseudorabies vaccines

A technology of pseudorabies virus and pseudorabies virus, which is applied in the field of pseudorabies vaccine production, can solve the problems of low virus yield, low virus titer, and large batch-to-batch difference, and achieve increased vaccine yield, stable genetics, and improved vaccine production. safety effect

Inactive Publication Date: 2011-02-23
PU LIKE BIO ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The invention provides a large-scale production method of pseudorabies vaccine, which can not only solve the shortcomings of low virus yield, low virus titer and large batch-to-batch difference in the traditional rotary bottle culture mode in vaccine production, but also eliminate The virus produced in the existing tidal production method has potential safety hazards in the subsequent production of live vaccines, and by optimizing parameters, high-titer virus liquids can be harvested multiple times, greatly improving the production of vaccines

Method used

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  • Method for preparing pseudorabies vaccines
  • Method for preparing pseudorabies vaccines
  • Method for preparing pseudorabies vaccines

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Embodiment 1 Utilizes Vero cells to continuously produce pseudorabies virus in a tidal bioreactor

[0060] Bioreactor: TideCell bioreactor, purchased from Saiyu Cell Technology Co., Ltd.

[0061] Carrier: BioNOC II carrier, purchased from Saiyu Cell Technology Co., Ltd.

[0062]CVD Nucleus Counting Kit: purchased from Saiyu Cell Technology Co., Ltd., Cat. No. AB0100, Lot No. 3007.

[0063] Glucose analyzer: purchased from Saiyu Cell Technology Co., Ltd.

[0064] Pseudorabies virus: Bartha-K61 strain (purchased from China Veterinary Drug Control Institute).

[0065] Medium: DMEM medium was purchased from GIBCO Company (product number: 12800-082, batch number: 1309922).

[0066] Trypsin: purchased from BD-Difco (product number: 215250, batch number: 5213753).

[0067] Fetal bovine serum: purchased from PAA company (article number: A15-151, batch number: A64905-0972)

[0068] Glucose: purchased from SIGMA Company (article number: 7021, batch number: MFCD00063774).

...

Embodiment 2

[0076] Example 2 BHK-21 cells continuously produce pseudorabies virus in a tidal bioreactor

[0077] Cell culture medium: MEM medium was purchased from GIBCO Company (product number: 61100-087, batch number: 678277), and prepared into cell culture medium as described in the product manual.

[0078] Bovine serum: purchased from Hyclone Company (article number: SV30087.02, batch number: NSK0069)

[0079] Pseudorabies virus: Fa gene deletion strain (TK - / gE - / gI - ) (see Chinese patent CN101186902A for this strain, which is deposited in the China Center for Type Culture Collection (CCTCC) with the accession number V200002).

[0080] Other materials and reagents are the same as in Example 1.

[0081] The BHK-21 cells used in this example are deposited in the American Type Culture Collection (ATCC) with the accession number CCL-10. For details, see the ATCC web page http: / / www.atcc.org / ATCCAdvancedCatalogSearch / ProductDetails / tabid / 452 / Default. aspx? ATCCNum=CCL-10&Template...

Embodiment 3

[0083] Example 3 MDBK cells continuously produce pseudorabies virus in a tidal bioreactor

[0084] Cell culture medium: MEM medium was purchased from GIBCO Company (product number: 61100-087, batch number: 678277), and prepared into cell culture medium as described in the product manual.

[0085] Newborn bovine serum (manufactured by GIBCO, product number 16010-14, batch number 693745)

[0086] Other materials and reagents are the same as in Example 1.

[0087] In this example, the MDBK cell (NBL-1) was deposited in the American Type Culture Collection (ATCC) with the accession number CCL-22. For details, please refer to the ATCC webpage. http: / / www.atcc.org / ATCCAdvancedCatalogSearch / ProductDetails / tabid / 452 / Default.aspx? ATCCNum=CCL-22&Template=cellBiology. In this embodiment, a tidal bioreactor with a carrier tank of 10L is adopted. The steps, methods and parameters adopted are basically the same as those in Example 1, and the cell growth solution and cell maintenance sol...

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Abstract

The invention relates to a method for producing pseudorabies vaccines by culturing passage cells by using a tidal bioreactor, which comprises the following steps of: (1) subculturing recovered cells; (2) performing high-density culture on the cells by using the specific bioreactor and a vector system to attach the cells onto vectors, and performing circulation, oxygenation and pH value and glucose regulation on a culture medium; (3) performing virus infection by using the bioreactor, replacing the culture medium with a virus culture medium, inoculating viruses and completely absorbing the viruses onto the cells; (4) culturing and amplifying the viruses under specific condition; (5) harvesting the viruses; and (6) adding a lyophilization protecting agent, and performing uniform mixing, quantitative subpackaging and lyophilization to obtain the pseudorabies vaccines.

Description

technical field [0001] The present invention relates to a kind of production method of pseudorabies vaccine, more specifically, the present invention relates to a kind of method that utilizes mammalian cell to produce pseudorabies vaccine on a large scale in the carrier tank of tidal type bioreactor. Background technique [0002] Pseudorabies (PR) is an infectious disease common to a variety of animals, causing acute and fatal infectious diseases in livestock and wild animals, with fever, itching and encephalomyelitis as the main symptoms, and pigs are its natural susceptible hosts. It mainly causes abortion, stillbirth and mummified fetuses in pregnant sows. Piglet infection is mainly neurological symptoms, while adult pigs are mainly recessive infection. This disease has become the main cause of reproductive failure syndrome in breeding pigs. Vaccine immunization is the fundamental measure to prevent and control pseudorabies, and inactivated vaccines, attenuated vaccines a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/02C12N7/00A61K39/245A61P31/20
Inventor 张许科孙进忠乔荣岑
Owner PU LIKE BIO ENG
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