Method for preparing pseudorabies live vaccines from DF1 continuous cells and product prepared by method

A technology for passaged cells and pseudorabies, which is applied in the field of biomedicine, can solve the problems of many cell fragments and impurities, easy to cause allergic reactions, large differences between cell batches, etc., and achieves high virus content, good immune protection, and production technology. stable effect

Inactive Publication Date: 2016-04-27
广东永顺生物制药股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Primary chick embryo fibroblast cells were used to culture pseudorabies virus, and the titer of toxin production was not high; the production of primary chicken embryo fibroblast cells had a large difference between batches of cells due to the difference in raw materials of chicken embryo fibroblasts; chicken embryo fibroblast There are many cell fragments and miscellaneous proteins in the first generation cells, which may easily cause allergic reactions in vaccinated pigs

Method used

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  • Method for preparing pseudorabies live vaccines from DF1 continuous cells and product prepared by method
  • Method for preparing pseudorabies live vaccines from DF1 continuous cells and product prepared by method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] The preparation of embodiment 1 pseudorabies live vaccine

[0031] Such as figure 1 Shown, a kind of method utilizing DF1 passage cell to produce pseudorabies live vaccine comprises the steps:

[0032] (1) Cell selection for seedling production: DF1 passage cells;

[0033] (2) Passage and culture of cells for seedling production: the above-mentioned DF1 passage cells were digested and passaged by EDTA-trypsin cell dispersion liquid, and continued to be cultured with cell growth liquid, and when a good monolayer was formed, they were used for continued passage or virus inoculation;

[0034] In the process of subculture and daily maintenance of cell culture, a lot of expense is required in terms of culture equipment, culture medium and various preparations, and once the cells leave the living body to start primary culture, its various biological characteristics will gradually change and become With the increase of the number of passages and the change of the environment...

Embodiment 2

[0048] Example 2 The safety test of pseudorabies DF1 passage cell live vaccine to pigs

[0049] 1 Materials and methods

[0050] 1.1 Experimental vaccines

[0051] Three batches of live cell vaccines for pseudorabies DF1 developed in the laboratory, the batch numbers are DF201301, DF201302, and DF201303.

[0052] 1.2 Experimental animals

[0053] Healthy pigs that were negative for neutralizing antibodies to pseudorabies virus were obtained from the experimental animal farm of Guangdong Yongshun Biopharmaceutical Co., Ltd.

[0054] 1.3 Safety test on pigs

[0055] 1.3.1 Safety trial of a single-dose immunization

[0056] Select healthy pigs aged 20-25 days and 50-55 days, and immunize pigs of different ages with 3 batches of laboratory products in a single dose. Each batch of vaccines is used to immunize 5 pigs, and each pig is intramuscularly injected with 1 head After inoculation, the temperature was measured and observed once a day in the morning and afternoon for 14 d...

Embodiment 2

[0076] Embodiment 2 pseudorabies DF1 subculture cell live vaccine toxin production test

[0077] 1 material

[0078] 1.1 Virus seeds were provided by Guangdong Yongshun Biopharmaceutical Co., Ltd.

[0079] 1.2 The cells were provided by Guangdong Yongshun Biopharmaceutical Co., Ltd.

[0080] 2 methods

[0081] Resuscitate the cells from the working cell bank and passage the cells, take the F10, F15, F20, F25, F30 generation cells that have formed a good monolayer, discard the nutrient solution, and inoculate with 0.1%~3% (ml / ml) production poison Keep the seed maintenance solution at 36-37°C to continue culturing, observe twice a day after inoculation, harvest the virus liquid when the CPE reaches 50%-100%, and store the harvested venom at -20°C or below. The cytotoxins harvested from the working cells of each passage were taken to measure the virus content respectively. And carry out two repeated experiments.

[0082] 3 results

[0083] The cells recovered from the work...

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Abstract

The invention provides a method for preparing pseudorabies live vaccines from DF1 continuous cells and a product prepared by the method. The method includes the steps of culturing a pseudorabies virus attenuated strain by the DF1 continuous cells; obtaining cell culture virus fluid; adding stabilizers and antibiotics and obtaining the pseudorabies DF1 live vaccines after vacuum freeze drying. The method has the advantages that the DF1 continuous cells which are heterogenous continuous cells are used for culturing pseudorabies attenuated viruses, so that the possibility that homologous cells have unknown swine-source microorganisms can be avoided and pure vaccines can be better guaranteed.

Description

technical field [0001] The invention relates to a method for producing pseudorabies live vaccine by using DF1 subcultured cells and its products, which belong to the technical field of biomedicine. Background technique [0002] The cells used in the production of live pseudorabies vaccines in my country are chicken embryo fibroblast primary cells. Primary chick embryo fibroblast cells were used to culture pseudorabies virus, and the titer of toxin production was not high; the production of primary chicken embryo fibroblast cells had a large difference between batches of cells due to the difference in raw materials of chicken embryo fibroblasts; chicken embryo fibroblast There are many cell fragments and miscellaneous proteins in the first generation cells, and the inoculated pigs are prone to cause allergic reactions. Contents of the invention [0003] In view of this, the purpose of the present invention is to provide a method for producing live pseudorabies vaccine by u...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/245A61P31/22
Inventor 吴文福林旭埜张毓金任向阳赖月辉黄秋雪牛晓芸
Owner 广东永顺生物制药股份有限公司
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