Serum free culture medium for growing various cells derived from kidney tissue

A serum-free culture medium and culture medium technology, applied in the field of cell engineering, can solve the problems of high price, legacy problems, and unsuitability for other cell lines, and achieve the effects of low price, simple preparation method, and improved biological safety

Inactive Publication Date: 2011-02-16
国家兽用生物制品工程技术研究中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] 1) The serum is susceptible to contamination by viruses, mycoplasma or other pathogens;
[0004] 2) Quality differences between different batches of serum cause quality differences between final product batches;
[0006] 4) It is difficult to completely remove some serum components, which seriously affects the quality of the final product and poses a safety hazard
[0009] 1) The serum-free medium still contains animal protein sourc

Method used

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  • Serum free culture medium for growing various cells derived from kidney tissue
  • Serum free culture medium for growing various cells derived from kidney tissue
  • Serum free culture medium for growing various cells derived from kidney tissue

Examples

Experimental program
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Effect test

Embodiment 1

[0089] The preparation of embodiment 1 serum-free culture medium of the present invention

[0090] The raw materials in Table 1 are all products of sigma company. According to the operation steps provided by the product manual of sigma company, the raw materials in table 1 are classified and dissolved according to their respective solubility characteristics, and then mixed at 25 °C to adjust the pH value of the resulting mixture to 7.2. Set the volume to 1000ml to obtain the culture medium. The content of each component in the culture medium is shown in the table below.

[0091] When mixing the ingredients, it is best to add an appropriate amount of non-pyrogenic ultra-clean pure water to 800ml, and stir for 30 minutes in the dark to fully dissolve, adjust the pH value to 7.2, and then adjust the volume to 1000ml. The prepared target culture medium was sterilized by filtration with a 0.22 μm filter membrane, and stored in a refrigerator at 4°C in the dark until use.

[0092] ...

Embodiment 2

[0096] The acclimation and culture steps of HEK293 cells, Marc145 cells, Vero cells and BHK21 cells in serum-free medium are as follows:

[0097] 1) The normally cultured HEK293 cells, Marc145 cells, Vero cells and BHK21 cells were adapted to reduce the amount of serum:

[0098] Step 1: Culture HEK293 cells, Marc145 cells, Vero cells and BHK21 cells using conventional methods (the amount of serum in DMEM medium is 10%), and then inoculate the above cells into DMEM supplemented with 5% (volume concentration) serum Cultured in culture medium, after 1-2 generations of adaptation, it can fully adapt to the culture conditions of DMEM supplemented with 5% serum;

[0099] The second step: the cells adapted to the DMEM medium supplemented with 5% serum were respectively inoculated into the DMEM medium with 2% serum concentration for culture, HEK293 cells and Marc145 cells were adapted for 2 passages, and Vero cells and BHK21 cells were adapted for 3 passages. Can be fully adapted to ...

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Abstract

The invention relates to a serum free culture medium for growing various cells derived from the kidney tissue. In the serum free culture medium, dulbecco's modified eagle medium (DMEM)/ nutrient mixture F-12(F12)(1:1) is used as a basic culture medium and comprises the components including yeast autolysate, soy protein hydrolysate, amino acids, a fatty acid combining agent, an antioxidant reagent, reduced glutathione, a polyamine mixture, ethanolamine, beta-mercaptoethanol, glycerol, minerals, Pluronic F-68 and the like, and the pH value is 7.2. The culture medium of the invention can be used for culturing various cell lines derived from the kidney tissue, such as HEK293 cells, Marc145 cells, Vero cells and BHK21 cells and for propagating viruses.

Description

technical field [0001] The invention relates to a serum-free medium used for the growth of various kidney tissue-derived cells, especially capable of supporting the cultivation of HEK293 cells, Vero cells, Marc145 cells or BHK21 cells, and belongs to the field of cell engineering. Background technique [0002] The host cells currently used for the amplification of recombinant viral vectors, recombinant protein expression or large-scale preparation of vaccines are basically mammalian kidney tissue-derived cells, such as human embryonic kidney cells (HEK293), African green ape kidney cells (Vero), Rhesus monkey kidney cells (Marc145) and young hamster kidney cells (BHK21), etc. These cells usually need to add a certain amount (5-10%) of bovine serum in the traditional culture process. The serum contains growth factors, hormones, carrier proteins, adhesion factors, trace elements and other nutrients required for cell growth, which can Promotes cell growth. However, in large-s...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 冯磊侯继波
Owner 国家兽用生物制品工程技术研究中心
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