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Low serum culture medium for BHK-21 cell culture and corresponding virus production

A technology of BHK-21 and cell culture, applied in the direction of cell culture active agent, microorganism-based method, virus, etc., can solve the problems of difficult to obtain semi-finished products, large amount of serum, short storage time, etc., and achieve a small reduction in poison price , Improve virus titer, pH stability effect

Inactive Publication Date: 2020-01-03
山东巨山能源科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The traditional BHK-21 cell culture method is to add 8% calf serum to DMEM high-glucose medium to cultivate cells, which requires a large amount of serum and high cost; the titer of the semi-finished product of pseudorabies vaccine produced by the traditional method is between 7.0-8.0, and it is under freezing conditions The price of poison drops rapidly, the storage time is short, and it is difficult to obtain semi-finished products that meet the standards for seedlings
[0004] There are some low-serum cell culture products on the market. Although the problem of serum dosage has been solved, the problem of rapid decline in the poison price of semi-finished products of pseudorabies vaccine has not been solved. Based on this, we propose a method for BHK-21 cell culture and corresponding virus production. low serum medium

Method used

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  • Low serum culture medium for BHK-21 cell culture and corresponding virus production
  • Low serum culture medium for BHK-21 cell culture and corresponding virus production

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Experimental program
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Effect test

Embodiment 1

[0018] A low-serum medium for BHK-21 cell culture and corresponding virus production, including the following components: glycine 37.5mg / L, L-alanine 8.9mg / L, L-arginine hydrochloride 84mg / L L, L-asparagine 256.8mg / L, L-aspartic acid 13.3mg / L, L-cystine dihydrochloride 63mg / L, cysteine ​​99mg / L, L-glutamic acid 195.3mg / L L, L-glutamine 398.8mg / L, L-histidine hydrochloride monohydrate 42mg / L, L-isoleucine 105mg / L, L-leucine 105mg / L, L-lysine Hydrochloride 146mg / L, L-methionine 30mg / L, L-phenylalanine 66mg / L, L-proline 11.5mg / L, L-serine 167.5mg / L, L-threonine 95mg / L, L-Tryptophan 16mg / L, L-Tyrosine Disodium Dihydrate 104mg / L, L-Valine 94mg / L, 4-Hydroxyethylpiperazineethanesulfonic acid (HEPES) 1287mg / L, sodium dihydrogen phosphate monohydrate 125mg / L, milk protein hydrolyzate 400mg / L, bovine serum albumin 200mg / L, bovine transferrin 7.5mg / L, recombinant human insulin 12.5mg / L, ammonium metavanadate 3.00E-04mg / L, copper sulfate pentahydrate 0.00125mg / L, manganese dichloride 5...

Embodiment 2

[0021]A low-serum medium for BHK-21 cell culture and corresponding virus production, including the following components: glycine 10mg / L, L-alanine 5mg / L, L-arginine hydrochloride 50mg / L, L-Asparagine 250mg / L, L-Aspartic Acid 10mg / L, L-Cystine Dihydrochloride 50mg / L, Cysteine ​​45mg / L, L-Glutamic Acid 100mg / L, L-Glutamine Aminoamide 200mg / L, L-histidine hydrochloride monohydrate 20mg / L, L-isoleucine 50mg / L, L-leucine 50mg / L, L-lysine hydrochloride 100mg / L , L-methionine 20mg / L, L-phenylalanine 50mg / L, L-proline 5mg / L, L-serine 100mg / L, L-threonine 50mg / L, L-tryptophan Acid 10mg / L, L-tyrosine disodium dihydrate 50mg / L, L-valine 50mg / L, 4-hydroxyethylpiperazineethanesulfonic acid (HEPES) 1000mg / L, dihydrogen phosphate monohydrate Sodium 50mg / L, milk protein hydrolyzate 50mg / L, bovine serum albumin 100mg / L, bovine transferrin 5mg / L, recombinant human insulin 5mg / L, ammonium metavanadate 0.0001mg / L, copper sulfate pentahydrate 0.0005 mg / L, manganese dichloride 0.00001mg / L, sodium...

Embodiment 3

[0024] A low-serum medium for BHK-21 cell culture and corresponding virus production, including the following components: glycine 50mg / L, L-alanine 15mg / L, L-arginine hydrochloride 150mg / L, L-Asparagine 350mg / L, L-Aspartic Acid 15mg / L, L-Cystine Dihydrochloride 120mg / L, Cysteine ​​110mg / L, L-Glutamic Acid 200mg / L, L-Glutamine Aminoamide 400mg / L, L-histidine hydrochloride monohydrate 50mg / L, L-isoleucine 120mg / L, L-leucine 120mg / L, L-lysine hydrochloride 200mg / L , L-methionine 50mg / L, L-phenylalanine 100mg / L, L-proline 15mg / L, L-serine 200mg / L, L-threonine 120mg / L, L-tryptophan Acid 20mg / L, L-tyrosine disodium dihydrate 120mg / L, L-valine 120mg / L, 4-hydroxyethylpiperazineethanesulfonic acid (HEPES) 1500mg / L, dihydrogen phosphate monohydrate Sodium 150mg / L, milk protein hydrolyzate 600mg / L, bovine serum albumin 600mg / L, bovine transferrin 15mg / L, recombinant human insulin 15mg / L, ammonium metavanadate 0.0005mg / L, copper sulfate pentahydrate 0.0015 mg / L, manganese dichloride 0.0...

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Abstract

The invention discloses a low serum culture medium for BHK-21 cell culture and corresponding virus production. The low serum culture medium is prepared from the following components: amino acid, a vitamin, inorganic salts, a protein, microelements and auxiliary components; the amino acid comprises glycine, L-arginine monohydrochloride, L-asparaginate, L-glutamic acid, L-glutamine, L-lysine hydrochloride and L-serine; and the protein comprises 4-HEPES, sodium dihydrogen phosphate monohydrate, milk protein hydrolysate, bovine serum albumin and recombinant human insulin. According to the low serum culture medium for the BHK-21 cell culture and the corresponding virus production, serum is no longer needed to be added in the maintenance stage of a pseudorabies vaccine produced from a BHK-21 cell, dosage of the serum is further reduced, the good growth state of cells can still be maintained, virus titer is improved, reduction of poison value of a semi-finished product is small, and preservation time of the semi-finished product is prolonged.

Description

technical field [0001] The invention relates to the field of biopharmaceutical vaccine production, in particular to a low-serum medium used for BHK-21 cell culture and corresponding virus production. Background technique [0002] BHK cells refer to BabyHamster Syrian Kidney cells, established in 1961. The original cell line is fibroblast, anchorage dependent. In 1963, a single cell clone was obtained. After countless passages, the cells can grow in suspension. It is widely used to multiply various viruses and produce veterinary vaccines. [0003] The traditional BHK-21 cell culture method is to add 8% calf serum to DMEM high-glucose medium to cultivate cells, which requires a large amount of serum and high cost; the titer of the semi-finished product of pseudorabies vaccine produced by the traditional method is between 7.0-8.0, and it is under freezing conditions The poison price drops rapidly, the storage time is short, and it is difficult to obtain semi-finished product...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12N7/00C12R1/93
CPCC12N5/0686C12N7/00C12N2500/90C12N2500/32C12N2500/38C12N2500/25C12N2500/12C12N2500/60C12N2500/34C12N2500/46C12N2500/30C12N2500/40C12N2500/36C12N2500/20C12N2500/14C12N2500/24C12N2500/16C12N2501/998C12N2501/999C12N2710/16751
Inventor 房圆瑗齐智刘海英巩俊廷
Owner 山东巨山能源科技有限公司
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