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Cell culture medium and its preparation method and application

A culture medium and cell technology, applied in the field of culture medium, can solve the problems of inducing immune response, animal-derived pollution, mycoplasma pollution, etc., and achieve the effect of easy-to-obtain raw materials, simple steps, and avoid inducing immune response

Active Publication Date: 2018-11-16
宁波医诺生物技术有限公司
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  • Summary
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  • Description
  • Claims
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Problems solved by technology

Although the above medium can guarantee the excellent self-renewal ability of stem cells and induced differentiation cells, due to the introduction of a large amount of animal-derived substances into the medium, it brings uncertainties to the self-renewal and differentiation process of stem cells and induced differentiation cells. factors, increasing the error; at the same time, it may introduce animal-derived contamination, such as mycoplasma contamination; in addition, it will induce immune response during the cell therapy

Method used

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  • Cell culture medium and its preparation method and application
  • Cell culture medium and its preparation method and application
  • Cell culture medium and its preparation method and application

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preparation example Construction

[0027] The present invention also provides a preparation method of a cell culture medium, the preparation method comprising:

[0028]a. Mix DMEM medium, F12 medium, sodium selenate, sodium bicarbonate, vitamin C, insulin, vascular endothelial growth factor and transforming growth factor β signaling pathway inhibitor to obtain mixture W1;

[0029] b. adjusting the pH of the mixture W1 to prepare the mixture W2;

[0030] c. Sterilizing the mixture W2 to prepare a cell culture medium.

[0031] In the above preparation method, the amount of each raw material can be selected in a wide range, but in order to provide more sufficient nutrients and a more stable living environment for the cultured cells in order to prepare the cell culture medium, preferably, the DMEM The volume ratio of the medium to the F12 medium is 1:1-1:4, the concentration of the vitamin C is 1-100 μg / ml, the concentration of the insulin is 1-100 μg / ml, the vascular endothelial growth factor The concentration i...

preparation example 1

[0038] 1) DMEM medium, F12 medium, sodium selenate, vitamin C, sodium bicarbonate, insulin, activin A, bone morphogenic protein 4, Rho protein kinase inhibitor (brand Y27632), glycogen synthase kinase- 3 inhibitor (brand CHIR99021) and glycogen synthase kinase-3 inhibitor (brand BIO) were mixed to prepare mixture W3; wherein, the volume ratio of DMEM medium to F12 medium was 1:2, and the concentration of vitamin C The concentration of insulin is 50 μg / ml, the concentration of insulin is 50 μg / ml, the concentration of activin A is 55 ng / ml, the concentration of bone morphogenic protein 4 is 60 ng / ml, and the concentration of glycogen synthase kinase-3 inhibitor (brand name is CHIR99021) The concentration of glycogen synthase kinase-3 inhibitor (brand is BIO) is 25 μM, the concentration of sodium selenate is 20 μg / L, and the concentration of sodium bicarbonate is 500 mg / L.

[0039] 2) adding sodium hydroxide to the above mixture W3 to adjust the pH to 7.8 to prepare mixture W4; ...

preparation example 2

[0042] Cell culture medium A2 was prepared according to the method of Preparation Example 1, except that no Rho protein kinase inhibitor was used in step 1), and the concentration of Rho protein kinase inhibitor was 10 μM.

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Abstract

The invention discloses a cell culture medium, and a preparation method and application thereof. The cell culture medium contains a DMEM, an F12 culture medium, sodium selenate, sodium bicarbonate, vitamin C, insulin, a vascular endothelial growth factor and a transforming growth factor beta signaling pathway inhibitor. The cell culture medium does not contain animal-derived ingredients, and can provide adequate nutrients and stable living environment for culture cells; in addition, the cell culture medium can effectively induce and differentiate mesendoderm progenitor cells into progenitor cells of vascular endothelial cells.

Description

technical field [0001] The invention relates to a culture medium, in particular to a cell culture medium and its preparation method and application. Background technique [0002] Stem cells refer to a type of cell that is capable of self-renewal, division, and differentiation into other cell types. Stem cells can be divided into embryonic stem cells (embryonic stem cells, ES cells) and adult stem cells (adult stem cells) according to the stage of stem cells in development. Embryonic stem cells are a kind of pluripotent stem cells (pluripotent stem cells), which can proliferate indefinitely and differentiate into more than 200 types of cells in the human body, forming tissues and organs of the body, that is, stem cells with the potential to differentiate into various cells and tissues. Since Evans and Kaufman isolated embryonic stem cells from the inner cell mass (ICM) of mice in 1981, ES cell research has become a hot research topic for scientists from all over the world. I...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071
Inventor 张竞方秦小平
Owner 宁波医诺生物技术有限公司
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