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A kind of low-serum medium suitable for bhk-21 cell spinner bottle culture and preparation method thereof

A technology of BHK-21 and preparation method, applied in the field of low serum medium and its preparation, can solve the problems of bacterial, fungal, virus and mycoplasma contamination, restriction of vaccine production enterprises, uncertainty of chemical composition, etc., to shorten the doubling time, Conducive to separation and purification, the effect of reduced content

Active Publication Date: 2019-03-12
内蒙古金源康生物工程股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the production process in China mostly adopts MEM medium supplemented with 10% calf serum for spinner bottle adherent culture. Due to the increasingly tight supply of serum in China and the rising price, there are obvious restrictions on Chinese vaccine manufacturers.
In addition, from the perspective of production, serum contains a large number of complex proteins, which is extremely unfavorable for downstream purification and separation of biological products such as vaccines, cytokines, and monoclonal antibodies. Batch stability issues and also prone to bacterial, fungal, viral and mycoplasma contamination

Method used

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  • A kind of low-serum medium suitable for bhk-21 cell spinner bottle culture and preparation method thereof
  • A kind of low-serum medium suitable for bhk-21 cell spinner bottle culture and preparation method thereof
  • A kind of low-serum medium suitable for bhk-21 cell spinner bottle culture and preparation method thereof

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Experimental program
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Effect test

Embodiment 1

[0041] The invention relates to a low serum medium (JYK-LSM) suitable for BHK-21 cell spinner bottle adherent culture and its preparation method. The medium is P0240 medium (DMEM) produced by Shanghai Yuanju Biotechnology Co., Ltd. Inorganic salts, hormones, lipids, trace elements, hydrolyzates and other components were added as the basic medium. The materials used were purchased from Sigma Company unless otherwise specified. The specific additions and contents are as follows:

[0042] Sodium selenite 0.005mg / L,

[0043] Hydrocortisone 0.01mg / L,

[0044] Prostaglandin E10.1mg / L,

[0045] Triiodothyronine (T3) 0.01mg / L,

[0046] Linoleic acid 0.1mg / L,

[0047] Soy lecithin 15mg / L,

[0048] Cholesterol 3mg / L,

[0049] Putrescine 0.8mg / L,

[0050] Ethanolamine 20mg / L,

[0051] Cottonseed hydrolyzate 800mg / L,

[0052] Rice hydrolyzate 3000mg / L,

[0053] Wheat bran hydrolyzate 300mg / L,

[0054] Sodium pyruvate 50mg / L,

[0055] Recombinant insulin 25mg / L,

[0056] Ferric...

Embodiment 2

[0061] The invention relates to a low serum medium (JYK-LSM) suitable for BHK-21 cell spinner bottle adherent culture and a preparation method thereof. The low serum medium is a P0240 medium produced by Shanghai Yuanju Biotechnology Co., Ltd. ( DMEM) was used as the basic medium to add inorganic salts, hormones, lipids, hydrolyzates and other components. Unless otherwise specified, the materials used were purchased from sigma company. The specific additions and contents were as follows:

[0062] Sodium selenite 0.015mg / L,

[0063] Hydrocortisone 0.04mg / L,

[0064] Prostaglandin E1--0.025mg / L,

[0065] Triiodothyronine (T3) 0.004mg / L,

[0066] Linoleic acid 0.2mg / L,

[0067] Soy lecithin 18mg / L,

[0068] Cholesterol 5mg / L,

[0069] Putrescine 0.4mg / L,

[0070] Ethanolamine 5mg / L,

[0071] Cottonseed hydrolyzate 3000mg / L,

[0072] Rice hydrolyzate 300mg / L,

[0073] Wheat bran hydrolyzate 800mg / L,

[0074] Sodium pyruvate 150mg / L,

[0075] Recombinant insulin 5mg / L,

[...

Embodiment 3

[0081] The invention relates to a low serum medium (JYK-LSM) suitable for BHK-21 cell spinner bottle adherent culture and a preparation method thereof. The low serum medium is a P0240 medium produced by Shanghai Yuanju Biotechnology Co., Ltd. ( DMEM) was used as the basic medium to add inorganic salts, hormones, lipids, hydrolyzates and other components. Unless otherwise specified, the materials used were purchased from sigma company. The specific additions and contents were as follows:

[0082] Sodium selenite 0.03mg / L,

[0083] Hydrocortisone 0.1mg / L,

[0084] Prostaglandin E1--0.01mg / L,

[0085] Triiodothyronine (T3) 0.001mg / L,

[0086] Linoleic acid 0.3mg / L,

[0087] Soy lecithin 20mg / L,

[0088] Cholesterol 10mg / L,

[0089] Putrescine 0.04mg / L,

[0090] Ethanolamine 1mg / L,

[0091] Cottonseed hydrolyzate 300mg / L,

[0092] Rice hydrolyzate 800mg / L,

[0093] Wheat bran hydrolyzate 3000mg / L,

[0094] Sodium pyruvate 300mg / L,

[0095] Recombinant insulin 2mg / L,

...

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Abstract

The present invention discloses a low-serum culture medium suitable for BHK-21 cell flask rotation culture, and a preparation method thereof. According to the present invention, in the flask rotation adherence culture, the serum consumption is substantially to 1% from the conventional 10%, the cell growth rate is rapid, the achieved cell density is high, and the high vitality is maintained; and during the culture medium development process, a plurality of compounds having special functions, lipid components, yeast, plant hydrolyzates and recombinant proteins are used to replace the conventionally-added animal-derived components in the culture medium, hormones, lipids, trace elements, hydrolyzates and other components are added to replace the partial functions of the serum so as to substantially reduce the serum addition amount, and reduce the serum cost while reduce the probability of the pollution caused by exogenous pathogenic bacteria carried by the serum and the animal-derived proteins in the culture medium, and the protein content in the culture medium is substantially reduced, such that the purification process at the late stage is easily achieved, and the stability of the cell product is improved.

Description

technical field [0001] The invention relates to the technical field of medicine and biological product manufacturing, in particular to a low-serum medium suitable for BHK-21 cell spinner bottle culture and a preparation method thereof. Background technique [0002] BHK-21 cell is a cell line approved by the World Health Organization and China Biological Products Regulations for the production of vaccines. It has the advantages of wide source, high biological safety, sensitivity to infection by various viruses, and high virus proliferation titer. The continuously passaged BHK-21 cell line can be used for the propagation and purification of various viruses, such as foot-and-mouth disease virus, encephalomyocarditis virus, reovirus, vesicular stomatitis virus, etc. All or part of the rabies vaccines, foot-and-mouth disease vaccines and pseudorabies vaccines produced in China are produced using BHK-21 cells. At present, the production process in China mostly uses MEM medium sup...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
Inventor 赵峻岭
Owner 内蒙古金源康生物工程股份有限公司
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