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Serum-free culture medium for mesenchymal stem cells

A serum-free medium and mesenchymal stem cell technology, applied in the field of mesenchymal stem cell serum-free medium, can solve the problems of affecting the growth of keratinocytes, accelerating keratinocyte differentiation, and fibroblast pollution, so as to prevent the occurrence and development of cancer , reduce blood viscosity, inhibit vasodilation

Active Publication Date: 2012-05-02
CHENGDU QINGKE BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The medium used in the existing mesenchymal stem cell culture methods mostly contains animal serum, which on the one hand causes contamination of fibroblasts, affects the growth of keratinocytes, and accelerates the differentiation of keratinocytes; on the other hand, due to the existence of serum and proteins with unknown components, Interfering with basic research in cell biology, toxicology, pathology, etc.; at the same time, the use of serum-supplemented media to construct artificial tissues for in vivo transplantation may cause immune reactions or introduce unsafe factors (such as viruses or other pathogens, etc.), Hence the need to develop chemically defined serum-free media
[0004] Due to the lack of composition of serum-free medium, the growth rate is slow and the characteristics of aging are prone to appear in the process of culturing cells in these medium, which affects the mass expansion of cells

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1: Adipose-derived mesenchymal stem cells are subcultured in culture medium;

[0039] Medium 1: serum-free medium for mesenchymal stem cells provided by the present invention;

[0040] Medium 2: L-DMEM+10%FBS (Fetal Bovine Serum);

[0041] Medium 3: STEMPRO MSC SFM serum-free medium.

[0042] The above-mentioned L-DMEM is provided by Gibco Company, the article number is 11885-084.

[0043] The above-mentioned STEMPRO MSC SFM serum-free medium is provided by Gibco Company, the product number is A10332-01.

[0044] Steps:

[0045] 1.1 Dilute adipose-derived mesenchymal stem cells at 10,000 cells / cm 2 The density was planted into three T75 cell culture flasks, and medium 1, medium 2, and medium 3 were added respectively. After the medium and the adipose-derived mesenchymal stem cell suspension were fully mixed, the three cell culture flasks were put into to 37°C, CO 2 The concentration was 5% in the cell culture incubator, and thereafter every 3 days accor...

Embodiment 2

[0052] Embodiment 2: Subculture of umbilical cord mesenchymal stem cells in culture medium;

[0053] Medium 1: serum-free medium for mesenchymal stem cells provided by the present invention;

[0054] Medium 2: L-DMEM+10%FBS (Fetal Bovine Serum);

[0055] Medium 3: STEMPRO MSC SFM serum-free medium.

[0056] The above-mentioned L-DMEM is provided by Gibco Company, the article number is 11885-084.

[0057]The above-mentioned STEMPRO MSC SFM serum-free medium is provided by Gibco Company, the product number is A10332-01.

[0058] Steps:

[0059] 2.1. The umbilical cord mesenchymal stem cells were divided into 10000 cells / cm 2 The density was planted into three T75 cell culture bottles, and medium 1, medium 2, and medium 3 were added respectively. After the medium and the umbilical cord mesenchymal stem cell suspension were fully mixed, the three cell culture bottles were put into to 37°C, CO 2 Cell culture was carried out in a cell incubator with a concentration of 5%, and ...

Embodiment 3

[0066] Example 3: Biological characteristics of umbilical cord mesenchymal stem cells and adipose mesenchymal stem cells subcultured in culture medium;

[0067] Medium: serum-free medium for mesenchymal stem cells provided by the present invention.

[0068] Steps:

[0069] 3.1 The umbilical cord mesenchymal stem cells and adipose-derived mesenchymal stem cells were divided into 10000 cells / cm 2 The densities were respectively planted in 2 T175 cell culture flasks, added to the serum-free medium of mesenchymal stem cells provided by the present invention for subculture, and each type of stem cells were subcultured 5 times and 10 times respectively;

[0070] 3.2 Harvest the cells in the cell culture flask, digest with 0.25% trypsin for 2 min, then blow off the mesenchymal stem cells and terminate the digestion with medium;

[0071] 3.3 Centrifuge the umbilical cord-derived mesenchymal stem cells and adipose-derived mesenchymal stem cells obtained in step 3.2 at a speed of 200g...

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Abstract

The invention discloses a serum-free culture medium for mesenchymal stem cells. The serum-free culture medium for the mesenchymal stem cells comprises the following ingredients: fibronectin at the final concentration of 25mu g / ml, basic fibroblast growth factors at the final concentration of 10ng / ml, human epidermal growth factors at the final concentration of 15ng / ml, recombinant human insulin at the final concentration of 1mg / ml, human transferrin at the final concentration of 0.55mg / ml, human blood albumin in a volume ratio of 5 percent, sodium selenite at the final concentration of 0.67mug / ml, L-carnitine at the final concentration of 5mM and resveratrol at the final concentration of 30mu M. When the mesenchymal stem cells are cultured by the serum-free culture medium for the mesenchymal stem cells, animal-derived serum is not contained, so infection risks can be controlled; the L-carnitine and the resveratrol which are added into the serum-free culture medium for the mesenchymalstem cells can effectively improve the growth state of the mesenchymal stem cells; and the growth speed of the mesenchymal stem cells is remarkably improved, and the biological characteristics of themesenchymal stem cells are kept unchanged.

Description

technical field [0001] The invention relates to the technical field of stem cell culture medium, in particular to a serum-free culture medium for mesenchymal stem cells. Background technique [0002] Mesenchymal Stem Cells (MSCs) are an important member of the stem cell family, derived from the mesoderm and ectoderm in the early stages of development, and have the characteristics of multi-directional differentiation potential, hematopoietic support and promotion of stem cell implantation, immune regulation and self-replication. And increasingly people's attention. Mesenchymal stem cells are widely present in various tissues throughout the body, cultured and expanded in vitro, and can differentiate into cardiomyocytes, nerve cells, and fat cells under specific conditions. With the in-depth research on the biological characteristics and functions of mesenchymal stem cells by scholars at home and abroad, mesenchymal stem cells can be isolated from various tissues. Mesenchymal...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
Inventor 申重阳郑榆坤海泉
Owner CHENGDU QINGKE BIOTECH
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