Serum-free culture medium for mesenchymal stem cells and application thereof

A serum-free medium and mesenchymal stem cell technology, applied in the field of mesenchymal stem cell serum-free medium, can solve the limitations of mesenchymal stem cell research and application, cell productivity, limited number of passages, and limit mesenchymal stem cell application, etc. problems, to achieve the effect of eliminating instability, maintaining differentiation potential, and high-efficiency biological characteristics

Active Publication Date: 2019-10-15
苏州依科赛生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although the content of mesenchymal stem cells in bone marrow is the most abundant, its content in bone marrow tissue is extremely low, which makes collection difficult and causes great damage to the body, which greatly limits the research and application of mesenchymal stem cells. How to obtain high activity and high consistency Safer bone marrow mesenchymal stem cells are an important topic in the research and application of mesenchymal stem cells
Traditional mesenchymal stem cell culture still needs to add fetal bovine serum. The cultured mesenchymal stem cells carry bovine serum albumin through endocytosis and uptake of bovine serum albumin, which can cause immune response of the recipient. At the same time, bacteria, bacteria, The risk of viruses, and the limited yield and number of passages of cells cultured in serum greatly limit the application of mesenchymal stem cells

Method used

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  • Serum-free culture medium for mesenchymal stem cells and application thereof
  • Serum-free culture medium for mesenchymal stem cells and application thereof
  • Serum-free culture medium for mesenchymal stem cells and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] The serum-free medium for mesenchymal stem cells provided in this example consists of the following components: basal medium and additional components, the basal medium is DMEM / F12, and the additional components are recorded as MAX1 components, MAX1 components and DMEM / F12 The specific material composition is as follows:

[0033] (1) The components contained in the MAX1 component and the content of each component are (based on the volume of the MAX1 component):

[0034]

[0035] (2) The components contained in the DMEM / F12 component and the content of each component are (based on the volume of the DMEM / F12 component):

[0036]

[0037]

[0038] In this example, the basal medium DMEM / F12 can also be purchased directly from the reagent company.

[0039] In this example, the serum-free medium for mesenchymal stem cells was prepared with DMEM / F12 and MAX1 at a volume ratio of 1000mL: 80mL, and the preparation method was prepared by conventional methods.

Embodiment 2

[0041] This example provides verification of the effect of the serum-free medium for mesenchymal stem cells in Example 1.

[0042] In the experiments of this example, human bone marrow mesenchymal stem cells were used. The cells were derived from the ATCC standard cell bank (PCS-500-012, Lot 70011720, ATCC). These cells were used in all the experiments below.

[0043] 1. Experimental method

[0044] 1. Cell culture

[0045] Human bone marrow mesenchymal stem cells, at 5000 / cm 2 The density was seeded on 100mm cell culture dishes. The cell culture dishes used in the experiment were all pre-coated and contained the following human adhesion molecules: 10 μg / cm 2 Human collagen IV (collagen IV), 5μg / cm 2 Human fibronectin.

[0046] Add mesenchymal stem cell serum-free medium and control medium containing bovine serum (DMEM / F12 medium (Sigma, M2906) with 10% fetal bovine serum (ExCellBio, FND500)) for continuous culture, the amount added is about 15ml , the medium was replaced...

Embodiment 3

[0061] This example provides verification of the effects of various culture media.

[0062] In the experiments of this example, human bone marrow mesenchymal stem cells were used. The cells were derived from the ATCC standard cell bank (PCS-500-012, Lot 70011720, ATCC). These cells were used in all the experiments below.

[0063] 1. Experimental method

[0064] 1. Preparation of culture medium:

[0065] serial number Medium solution volume ratio (DMEM / F12: MAX1) M1 10:0.4 M2 10:0.6 M3 10:0.8 M4 10:1.0 M5 10:1.6 M6 commercial serum-free medium I M7 commercial serum-free medium II M8 serum containing medium

[0066] Wherein: the culture medium in numbering M1~M5 is the culture medium of basic medium and MAX1 under different volume proportions, and the component and content thereof of basic medium and MAX1 are the same as embodiment 1;

[0067] Commercial serum-free medium I: imported brand MSC serum-free medium (Milten...

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Abstract

The invention relates to a serum-free culture medium for mesenchymal stem cells and application thereof. The serum-free culture medium comprises a basal culture medium and additive components. The additive components comprise the following components: 2'-Deoxyadenosine, 2'-Deoxycytidine-HCl, 2'-Deoxyguanosine, L-glutamine, human serum albumin, recombinant human transferrin, recombinant human insulin, rhPDGF-BB, rhFGF-b, rhTGF-beta1, rhEGF, sodium selenate and hydrocortisone. The serum-free culture medium disclosed by the invention does not contain animal-derived heterologous component such asbovine serum, and can effectively replace serum to culture bone marrow mesenchymal stem cells; and due to the synergistic effect of the additive components, the culture medium has the advantages of shortening cell cycle, rapid proliferation and good cell consistency while culturing bone marrow mesenchymal stem cells, and can effectively maintain the molecular characteristics and differentiation potential of the bone marrow mesenchymal stem cells. The bone marrow mesenchymal stem cells obtained by the culture medium are suitable for further scientific research and clinical application research.

Description

technical field [0001] The invention belongs to the field of cell culture, and in particular relates to a serum-free medium for mesenchymal stem cells and an application thereof. Background technique [0002] Mesenchymal stem cells (MSCs) belong to the pluripotent stem cells of the mesoderm, mainly exist in connective tissue and organ mesenchyme, have the ability of self-renewal and multi-directional differentiation, and are widely found in umbilical cord tissue, fat, bone marrow, bone, etc. Muscle, periosteum, lung, liver, pancreas and other tissues as well as amniotic fluid and umbilical cord blood. Mesenchymal stem cells have a very strong ability to differentiate. Under specific conditions, they can proliferate and differentiate into various types of cells such as fat cells, bone cells, chondrocytes, nerve cells, and muscle cells. [0003] Among mesenchymal stem cells, bone marrow-derived mesenchymal stem cells (Bone Marrow-Derived Mesenchymal Stem Cells, BM-MSCs) are t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0662C12N5/0665C12N5/0667C12N2500/12C12N2500/24C12N2500/30C12N2500/32C12N2500/40C12N2501/11C12N2501/115C12N2501/135C12N2501/33
Inventor 于宝利陈旭陈刚
Owner 苏州依科赛生物科技股份有限公司
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