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.

Inactive Publication Date: 2019-09-20
GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The first generation is a serum-free medium in a general sense, using various biological materials that can replace the function of serum, containing a large amount of animal-derived proteins and unknown components, such as animal or human-derived platelet lysates, etc.; its advantage is compared with containing Serum medium has higher experimental accuracy, repeatability, and stability, but because the chemical composition of the added substances is not clear, most of them contain a large amount of animal-derived protein, which is not conducive to the separation and purification of the target protein, and the cost is high
The second generation is a serum-free and animal-derived protein-free medium. Its additives do not use animal-derived protein at all, but replace it with various recombinant proteins or animal and plant protein hydrolyzates; its advantage is that its stability is improved compared with the first generation. The cost is reduced, and the effect is also improved accordingly. However, due to the high cost, it is only suitable for scientific research users with small demand, and it is difficult to be used by lar...

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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

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Experimental program
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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

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IPC IPC(8): C12N5/0775
CPCC12N5/0662C12N2500/12C12N2500/20C12N2500/22C12N2500/24C12N2500/32C12N2500/36C12N2500/38C12N2500/44C12N2500/46C12N2500/90C12N2501/11C12N2501/115C12N2501/734C12N2501/855C12N2501/999
Inventor 陈东煌陈海佳葛啸虎戚康艺姜交华岳坤
Owner GUANGZHOU SALIAI STEMCELL SCI & TECH CO LTD
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