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Preparation method of fresh mesenchymal stem cells

A mesenchymal stem cell, fresh technology, applied in the field of preparation of fresh mesenchymal stem cells, can solve problems such as cryopreservation, achieve the effect of increasing positive effects, avoiding thawing links, and reducing the reduction of cell activity

Pending Publication Date: 2018-07-17
刘金宏
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem in the prior art that mesenchymal stem cells need to be frozen at -196°C before use, the purpose of the present invention is to provide a method for preparing fresh mesenchymal stem cells

Method used

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  • Preparation method of fresh mesenchymal stem cells
  • Preparation method of fresh mesenchymal stem cells
  • Preparation method of fresh mesenchymal stem cells

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Experimental program
Comparison scheme
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Embodiment Construction

[0036] 1. The composition of the first culture medium

[0037] 1. Amino acids

[0038] 30mg / L glycine, 85mg / L L-arginine hydrochloride, 60mg / L L-cystine dihydrochloride, 585 mg / L L-glutamine, 45 mg / L L-histidine hydrochloride monohydrate, 105mg / L L-Isoleucine, 105 mg / L L-Leucine, 150 mg / L L-Lysine Hydrochloride, 30 mg / L L-Methionine, 65 mg / L L-Benzene Alanine, 40 mg / L L-Serine, 95 mg / L L-Threonine, 18 mg / L L-Tryptophan, 105 mg / L L-Tyrosine Disodium Salt, 95 mg / L L - Valine.

[0039] 2. Vitamins

[0040] 4mg / L choline chloride, 4mg / LD-calcium pantothenate, 5mg / L folic acid, 4mg / L nicotinamide, 4mg / L pyridoxal hydrochloride, 0.5mg / L riboflavin, 4mg / L thiamine hydrochloride, 8mg / L inositol.

[0041] 3. Inorganic salts

[0042] 200mg / L anhydrous calcium chloride, 0.1mg / L ferric nitrate nonahydrate, 98mg / L anhydrous magnesium sulfate, 400mg / L potassium chloride, 3700mg / L sodium bicarbonate, 6400mg / L sodium chloride, 125mg / L Sodium dihydrogen phosphate hydrate.

[0043] 4. O...

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Abstract

The invention discloses a preparation method of fresh mesenchymal stem cells. The preparation method comprises the following steps: S1, separating primary cells from umbilical cord Wharton jelly; S2,carrying out differential velocity adherence and separating and purifying umbilical cord mesenchymal stem cells; S3, discarding a first culture medium in a culture dish and changing into a fresh firstculture medium; S4, adding pancreatin for passaging adherent growth cells; S5, repeating step S3; S6, adding cells treated in S5 into a cryoprotectant, and putting at the temperature of -80 DEG C forpreserving; S7, thawing the cells frozen in S6; S8, adding a second culture medium into the cells thawed in S7, eluting the cryoprotectant, and extracting cell suspension for culturing; S9, centrifuging the cells cultured in S8, discarding supernatant and adding into a preservative solution. According to the preparation method of the fresh mesenchymal stem cells, disclosed by the invention, a cryopreservation link is eliminated, and the fresh mesenchymal stem cells are prepared; activity and quantity of the cells are remarkably improved, adverse reactions are fewer, and transportation and useare more convenient.

Description

technical field [0001] The invention relates to the technical field of stem cell preparation, in particular to a method for preparing fresh mesenchymal stem cells. Background technique [0002] Contemporary cell biology studies have shown that mesenchymal stem cells (mesenchymal stem cells, MSCs) with self-renewal and multi-directional differentiation potential can be applied to the treatment and rehabilitation of various diseases. For example, the application of embryonic stem cells has problems such as ethics, law, immune rejection and tumorigenicity; autologous bone marrow mononuclear cells have complex components, low content of stem cells with multidirectional differentiation potential, and limited differentiation potential; autologous bone marrow The source of mesenchymal stem cells is limited, and the proliferation and differentiation potential of cells significantly decreases with age, and long-term expansion in vitro reduces the differentiation and homing potential ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775A01N1/02
CPCA01N1/0221C12N5/0668C12N2509/00
Inventor 刘金宏
Owner 刘金宏
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