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Human umbilical cord mesenchymal stem cell serum-free culture medium and preparation method and application thereof

A technology of serum-free medium and stem cells, applied in the field of stem cells, can solve the problems of high price, short storage time, unsatisfactory cell proliferation effect, etc., and achieve the effect of low cost, good growth state, and good stability between batches

Inactive Publication Date: 2020-01-17
湖南丰晖生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At the same time, the current commercial serum-free medium is expensive, the storage time is short, and the cell proliferation effect is not ideal

Method used

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  • Human umbilical cord mesenchymal stem cell serum-free culture medium and preparation method and application thereof
  • Human umbilical cord mesenchymal stem cell serum-free culture medium and preparation method and application thereof
  • Human umbilical cord mesenchymal stem cell serum-free culture medium and preparation method and application thereof

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Experimental program
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Effect test

Embodiment 1

[0025] Embodiment 1 of the present invention is: preparation and use of human umbilical cord mesenchymal stem cell serum-free medium:

[0026] 1. Preparation of serum-free medium:

[0027] In order to study the influence of the concentrations of human serum albumin, transferrin and recombinant human insulin on the medium of the present invention, medium 1-9 was prepared for comparative experiments. In culture medium 1-9, all use DMEM / F12 type culture medium as basal culture medium, other same components also include: Glycine (7.5mg / L), alanine (8.9mg / L), asparagine (13.2 mg / L), aspartic acid (13.3mg / L), glutamic acid (14.7mg / L), proline (11.5mg / L), serine (10.5mg / L), glutamine (5mM) , sodium selenite (5ng / mL), recombinant human epidermal growth factor (10ng / mL), basic fibroblast growth factor (10ng / mL), HEPES (10-25mM all can, what used in the present embodiment is 10mM); except above-mentioned identical components, also comprise recombinant human serum albumin, transferrin ...

Embodiment 2

[0033]Embodiment two of the present invention is: the cell culture control experiment of different component concentration culture medium formula:

[0034] Umbilical cord mesenchymal stem cells were recovered, and the cells were divided into 9 groups for MTT experiment. Cells were inoculated into 96-well plate, 3000 per well, 5 wells in each group, cultured in medium 1-9 in 9 groups, and cell proliferation was detected at 0, 24, 48, 72, and 96 hours. The cells adhered to the wall 4 hours after inoculation, took a 96-well plate, added 10 μL of MTT reagent, incubated at 37°C for 4 hours, and detected the OD value of the cells at 490 nm with a microplate reader, which was recorded as 0 hour. Afterwards, a 96-well plate was taken every 24 hours for MTT detection until the measurement was completed in 96 hours, the data was sorted out, and the average value was calculated. The analysis results are shown in Table 2 and figure 2 (with time as the horizontal and vertical scale, and ...

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Abstract

The invention discloses a human umbilical cord mesenchymal stem cell serum-free culture medium and a preparation method and application thereof. The human umbilical cord mesenchymal stem cell serum-free culture medium comprises a basic culture medium, amino acid substances, trace elements, recombinant human serum albumin, transferrin, recombinant human insulin, recombinant human epidermal growth factors, alkaline fibroblast growth factors and buffer salt, wherein the amino acid substances comprise glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, serine and glutamine. The serum-free culture medium is clear in components, stable in batch, free of serum, antibiotics and other serum analogues, and free of heterogeneous pollution and immunological rejection risks and safe touse; by the serum-free culture medium, normal cell adherence and good cell growth state can be kept; the serum-free culture medium has stemness and is suitable for the growth and stemness maintenanceof human umbilical cord mesenchymal stem cells.

Description

technical field [0001] The invention belongs to the technical field of stem cells, and in particular relates to a serum-free medium for human umbilical cord mesenchymal stem cells and a preparation method and application thereof. Background technique [0002] Mesenchymal stem cells (MesenchymaL Stem Cell, MSC) is a kind of adult stem cells with multi-directional differentiation potential, which can differentiate into osteoblasts, adipocytes, fibroblasts, stromal cells and other cells, and has immune regulation, tissue Repair, inflammatory chemotaxis and other functions can be isolated from bone marrow, fat, umbilical cord, placenta and other tissues. Due to their low immunogenicity and strong proliferative ability, MSCs have been widely used in the treatment of clinical diseases and medical cosmetology in recent years. Bone marrow-derived mesenchymal stem cells are the earliest method of obtaining them, but their application is limited due to the strict requirements for the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0668C12N2500/25C12N2500/32C12N2500/60C12N2500/90C12N2501/11C12N2501/115C12N2501/998
Inventor 许澎黄巧胡清云
Owner 湖南丰晖生物科技有限公司
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