Serum-free culture medium for in-vitro culture and amplification of bone marrow mesenchymal stem cells

A technology of serum-free medium and bone marrow mesenchyme, which is applied in the field of cell biology, can solve the problems of residual animal serum, failure to achieve serum-free culture, and inability to promote bone marrow mesenchymal stem cell adhesion, etc., to achieve no environmental pollution, Maintain biological effects and multi-lineage differentiation potential, maintain the effect of multi-lineage differentiation potential

Pending Publication Date: 2021-04-16
CHENGDU ZHIA TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, none of these serum-free media can promote bone marrow mesenchymal stem cell attachment
When cultured in vitro, bone marrow mesenchymal stem cells need to be completely adhered to the wall in a serum-containing medium, and then replaced with a serum-free medium for culture, which will cause a certain amount of animal serum to remain in the medium. It is said that the true serum-free culture has not been achieved, and it may still have a great risk in clinical application

Method used

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  • Serum-free culture medium for in-vitro culture and amplification of bone marrow mesenchymal stem cells

Examples

Experimental program
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Effect test

Embodiment 1

[0022] A serum-free medium for culturing and expanding bone marrow mesenchymal stem cells in vitro, characterized in that it is an aqueous solution, and its components and contents are as follows: basal medium 20g / L, growth factor 5ng / mL, Kombucha fermentation broth 8mg / mL, serum protein 3mg / mL, selenomethionine 1ng / mL, sodium ferulate 0.1mg / mL, metformin 3ng / mL, 3'-deoxynicotinamide adenine dinucleotide 5ng / mL , L-glutamine 150mg / L, multivitamin 30μM, insulin 12mU / L.

[0023] The multivitamins are formed by mixing vitamin A, vitamin B2, vitamin C, vitamin D and vitamin E in a mass ratio of 1:1:0.8:2:1.

[0024] The basal medium is DMEM-F12; the DMEM-F12 culture solution is a mixture of F12 culture solution and DMEM culture solution at a volume ratio of 1:2.

[0025] The growth factors are transforming growth factor β1 (TGF-β1), heparin-binding epidermal growth factor (HB-β), insulin growth factor-1 (IGF-1), basic fibroblast growth factor (bFGF), epidermal growth factor The ...

Embodiment 2

[0029] A serum-free medium for culturing and expanding bone marrow mesenchymal stem cells in vitro, characterized in that it is an aqueous solution, and its components and contents are as follows: basal medium 25g / L, growth factor 8ng / mL, Kombucha fermentation broth 12mg / mL, serum protein 8mg / mL, selenomethionine 3ng / mL, sodium ferulate 0.2mg / mL, metformin 4ng / mL, 3'-deoxynicotinamide adenine dinucleotide 7ng / mL , L-glutamine 250mg / L, multivitamin 50μM, insulin 17mU / L.

[0030] The multivitamins are formed by mixing vitamin A, vitamin B2, vitamin C, vitamin D and vitamin E in a mass ratio of 1.3:1:0.9:2:1.3.

[0031] The basal medium is DMEM-F12; the DMEM-F12 culture solution is a mixture of F12 culture solution and DMEM culture solution at a volume ratio of 1:2.5.

[0032] The growth factors are transforming growth factor β1 (TGF-β1), heparin-binding epidermal growth factor (HB-β), insulin growth factor-1 (IGF-1), basic fibroblast growth factor (bFGF), epidermal growth factor ...

Embodiment 3

[0036] A serum-free medium for culturing and expanding bone marrow mesenchymal stem cells in vitro, characterized in that it is an aqueous solution, and its components and contents are as follows: basal medium 30g / L, growth factor 10ng / mL, Kombucha fermentation broth 16mg / mL, serum protein 13mg / mL, selenomethionine 6ng / mL, sodium ferulate 0.25mg / mL, metformin 6ng / mL, 3'-deoxynicotinamide adenine dinucleotide 8ng / mL , L-glutamine 350mg / L, multivitamin 70μM, insulin 19mU / L.

[0037] The multivitamins are formed by mixing vitamin A, vitamin B2, vitamin C, vitamin D and vitamin E in a mass ratio of 1.5:1:1:2:1.5.

[0038] The basal medium is DMEM-F12; the DMEM-F12 culture solution is a mixture of F12 culture solution and DMEM culture solution at a volume ratio of 1:3.

[0039] The growth factors are transforming growth factor β1 (TGF-β1), heparin-binding epidermal growth factor (HB-β), insulin growth factor-1 (IGF-1), basic fibroblast growth factor (bFGF), epidermal growth factor...

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Abstract

The serum-free culture medium is characterized in that the serum-free culture medium is an aqueous solution and comprises the following components: 20-40g / L of a basal culture medium, 5-15ng / mL of a growth factor, 8-22mg / mL of kombucha fermentation liquor, 3-20mg / mL of serum protein, 1-10ng / mL of selenomethionine, 0.1-0.4 mg / mL of sodium ferulate, 3-8ng / mL of metformin, 5-12 ng / mL of 3'-deoxidized nicotinamide adenine dinucleotide, 150 mg / L-550 mg / L of Lglutamine, 30-110 [mu] M of composite vitamin and 12 mU / L-24 mU / L of insulin. The invention further provides a preparation method and application of the serum-free culture medium for in-vitro culture and amplification of the bone marrow mesenchymal stem cells. The serum-free culture medium for in-vitro culture and amplification of bone marrow mesenchymal stem cells disclosed by the invention is good in biological safety, can effectively improve the amplification efficiency of human bone marrow mesenchymal stem cells, and maintains the biological effect and the multidirectional differentiation potential at the same time; the bone marrow mesenchymal stem cells with related requirements can be timely and accurately provided for medical scientific research teaching and clinical application.

Description

technical field [0001] The invention relates to the technical field of cell biology, in particular to a serum-free medium for culturing and expanding bone marrow mesenchymal stem cells in vitro. Background technique [0002] Bone marrow mesenchymal stem cells (MSCs) are the main component of the bone marrow stroma, providing structural and functional support for the hematopoietic function of the bone marrow. This stem cell is an adult stem cell with self-replication and multilineage differentiation potential. Under certain induction conditions, MSCs have the ability to differentiate into mesoderm cells such as osteoblasts, chondrocytes, myoblasts, tenocytes, adipocytes and stromal cells; at the same time, MSCs can also differentiate into ectoderm neurons and endoderm hepatic oval cell differentiation. Bone marrow mesenchymal stem cells can promote the reconstruction of the hematopoietic system, have effective immune regulation function, can inhibit the proliferation of T-l...

Claims

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Application Information

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IPC IPC(8): C12N5/0775
Inventor 向兴力
Owner CHENGDU ZHIA TECH CO LTD
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