Serum-free culture medium for mesenchymal stem cells

A serum-free medium and mesenchymal stem cell technology, applied in the field of serum-free medium for mesenchymal stem cells, can solve the problems of difficult product development, unclear chemical composition of added substances, unfavorable separation and purification of target proteins, etc.

A serum-free medium and mesenchymal stem cell technology, applied in the field of serum-free medium for mesenchymal stem cells, can solve the problems of difficult product development, unclear chemical composition of added substances, unfavorable separation and purification of target proteins, etc.

CN110257328AInactive Publication Date: 2019-09-20GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD +1
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  • Serum-free culture medium for mesenchymal stem cells
  • Serum-free culture medium for mesenchymal stem cells
  • Serum-free culture medium for mesenchymal stem cells

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0121] The preparation of the serum-free medium described in this application can be prepared according to the methods well known to those skilled in the art. After mixing the components at a specific time, they are filtered and sterilized in a filter membrane to obtain the mesenchymal stem cells (UC-MSCs). ) serum-free medium.

[0122] The mesenchymal stem cell-promoting polypeptide composition provided by the present invention has clear chemical composition, no animal source, no serum, extremely low protein content, stable performance, can realize rapid proliferation of UC-MSCs, solve the problem of insufficient cell number, and maintain interstitial Biological properties and immunophenotypic stability of mesenchymal stem cells.

Embodiment 1

[0124] Example 1 Preparation of serum-free medium

[0125] 1) Based on α-MEM basal medium, prepare serum-free medium according to the following raw material ratio:

[0126] Lipid mixture: 1vol%;

[0127] Glutamine: 2mM;

[0128] Hydrocortisone: 50µg / L;

[0129] Zinc sulfate: 0.5mg / L;

[0130] Sodium butyrate: 1mM;

[0131] Ferric citrate: 50µM;

[0132] Vitamin C: 100µM;

[0133] Recombinant human epidermal growth factor: 20µg / L;

[0134] Recombinant human basic fibroblast growth factor: 20µg / L;

[0135] RGD short peptide: 10µg / L;

[0136] Palmitoyl tripeptide-5: 25µg / L;

[0137] Palmitoyl Tetrapeptide-7: 25µg / L;

[0138] Lithium chloride: 5mM;

[0139] L-Glutathione: 4mg / L;

[0140] Soybean trypsin inhibitor: 100mg / L;

[0141] β-mercaptoethanol: 2.5mg / L;

[0142] Ethanolamine: 0.2g / L;

[0143] Sodium selenite: 0.00067mg / L;

[0144] Sodium pyruvate: 0.1mM;

[0145] Hepes: 0.01M;

[0146] NaHCO 3 : 3mM;

[0147] Dissolve the above-mentioned components in wat...

Embodiment 2

[0235] Example 2 Morphological observation and activity detection of UC-MSCs

[0236] Select the P3 generation UC-MSCs to carry out the experiment, and the UC-MSCs are divided into 1×10 4 / cm 2 Density inoculated in T25 culture flasks, with 3 replicates in each group, placed in 5% CO 2 Cultivate in an incubator at 37°C, collect images of UC-MSCs after 48 hours of cultivation, and the results are as follows: figure 1 as shown, figure 1 Middle panel A is the morphology of UC-MSCs in control group 1, panel B is the morphology of UC-MSCs in control group 2, panel C is the morphology of UC-MSCs in control group 3, and panel D is the morphology of UC-MSCs in control group 4 Figure E is the morphology of UC-MSCs in experimental group 1, Figure F is the morphology of UC-MSCs in experimental group 2, and Figure G is the morphology of UC-MSCs in experimental group 3.

[0237] Depend on figure 1 It can be seen that the UC-MSCs in each group grew in a single layer of adherent, most o...

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Abstract

The invention provides a serum-free culture medium for mesenchymal stem cells. The serum-free culture medium specifically comprises a lipid mixture, glutamine, hydrocortisone, zinc sulfate, sodium butyrate, ferric citrate, vitamin C, recombinant epidermal growth factors, recombinant alkaline fibroblast growth factors, RGD oligopeptides, palmitoyl tripeptide-5, palmitoyl tetrapeptide-7, lithium chloride, L-glutathione, a soybean pancreatin inhibitor, beta-mercaptoethanol, ethanolamine, sodium selenite, Hepes, sodium bicarbonate and a basic culture medium. The serum-free culture medium is free of human and animal resources, clear in chemical component, rich in nutrition, capable of achieving UC-MSCs rapid proliferation, and capable of solving the problem of cells number insufficiency and maintaining the biological features and immunophenotyping stability of the mesenchymal stem cells.

Description

technical field [0001] The invention relates to the technical field of medium, in particular to a serum-free medium for mesenchymal stem cells. Background technique [0002] Mesenchymal stem cells (MSCs) are derived from the mesoderm in the early stage of development and are a type of non-hematopoietic stem cells that are widely distributed in bone marrow, subcutaneous fat, periosteum, muscle, synovium, synovial fluid, liver, peripheral tissue, umbilical cord, cord blood and placenta. MSCs have high self-renewal ability and multi-directional differentiation potential, and can be cultured and expanded in vitro. They can not only support the growth of hematopoietic stem cells, but also have the function of immune regulation; under different induction conditions, they can differentiate into bone, cartilage, and muscle in vitro. , nerve, myocardium, endothelium and fat, etc., still have multi-directional differentiation potential after continuous subculture and cryopreservation...

Claims

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Application Information

Patent Timeline
20 Sep 2019
Publication
CN110257328A
IPC
C12N5/0775
CPC
C12N5/0662; C12N2500/12; C12N2500/20; C12N2500/22; C12N2500/24; C12N2500/32; C12N2500/36; C12N2500/38
Inventors
陈东煌; 陈海佳