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Mesenchymal stem cell cryopreservation agent

A cryopreservation agent and stem cell technology, applied in the field of bone marrow mesenchymal stem cell cryopreservation agent and its preparation, can solve the problems of cytotoxicity, stem cell damage, etc., and achieve simple and feasible operation, reasonable price, and improved survival rate of resuscitation Effect

Inactive Publication Date: 2017-10-10
陈印平
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, DMSO can be toxic to cells and cause irreversible damage to cryopreserved stem cells

Method used

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  • Mesenchymal stem cell cryopreservation agent

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Add 0.2% w / v sucrose, 0.5% w / v LDL, 3% w / v trehalose, 2% w / v lecithin, peptide 100ppm w / v, bFGF 1.5% w / v, vitamin E 1% w / v, glycerol 5% v / v Finally, use DMEM medium and fetal bovine serum to make up to 100ml according to the ratio of 1:1. The sequence of the polypeptide is shown as SEKTWFVWYSWRKQGCIEDSRPTYHYC.

Embodiment 2

[0023] Add 0.2% w / v sucrose, 0.5% w / v LDL, 3% w / v trehalose, 2% w / v lecithin, peptide 100ppm w / v, bFGF 1.5% w / v, vitamin E 1% w / v, glycerol 5% v / v Finally, use DMEM medium and fetal bovine serum to make up to 100ml according to the ratio of 1:1. The sequence of the polypeptide is shown as KAREWEPDRHMTHWIPGFHHMGCSYH.

Embodiment 3

[0030] The effect verification of embodiment 3 cryopreservation solution

[0031] The cell cryopreservation solutions prepared in the above Examples 1-2 and Comparative Examples I-III were respectively subjected to cell cryopreservation and recovery experiments according to the following methods.

[0032] Cell cryopreservation process:

[0033] Cultivate human leukemia stem cells growing into a monolayer, the cell density is about 6*10 9 cells / ml, add pH 7.0 PBS to wash the cell surface once.

[0034] Digest the cells with 0.25% trypsin solution for 4 minutes, discard the trypsin solution, add 15 ml of DMEM culture solution, blow gently with a pipette to make the cells uniform, centrifuge at 4000 rpm, discard the supernatant, add Examples 1-3 and compare Mix 2ml of the cryopreservation solution prepared in Example 1, and put it into a sterile cryopreservation tube;

[0035] Freezing: first freeze the cryopreservation tubes at 4°C for 30 minutes, then store them at -30°C for...

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PUM

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Abstract

The invention discloses a mesenchymal stem cell cryopreservation agent which comprises 0.2%w / v cane sugar, 0.5%w / v low-density lipoprotein, 3%w / v trehalose, 2%w / v lecithin, 100ppm w / v polypeptide, 1.5%w / v bFGF, 1%w / v vitamin E, 5% v / v glycerin and the balance of a DMEM (Dulbecco's Modified Eagle Medium) and fetal calf serum in a ratio of 1:1 till 100ml. In the mesenchymal stem cell cryopreservation agent disclosed by the invention, the glycerin, the trehalose, the vitamin E and the polypeptide have definite functions of preventing external damage to cells, the polypeptide also has a function of preventing cells from damage of ice crystal generated in cryopreservation, and the other components are essential for maintaining survival of specific MSCs (Mesenchymal Stem Cells). The cell cryopreservation agent disclosed by the invention is applied to cryopreservation and culture of cells, is particularly capable of increasing the survival rate of revived cryopreservation cells, and has relatively good application prospects.

Description

technical field [0001] The invention relates to the technical field of tissue cell culture, in particular to a cryopreservation agent for bone marrow mesenchymal stem cells and its preparation method and application. Background technique [0002] In addition to hematopoietic stem cells that can differentiate and develop into various blood cells, bone marrow also contains mesenchymal stem cells (MSCs) that produce non-hematopoietic tissues. Because it is easier to adhere to the wall and form fibroblast-like clones, some literatures MSCs are also called adherent cells (fibroblast colony forming units (CFU-F). Because they come from the support structure of bone marrow and can serve as a trophoblast to support the growth of hematopoietic stem cells, some people also call them bone marrow stromal cells. [0003] The multilineage differentiation potential of MSCs has been verified by in vivo and in vitro experiments. Friedenstem first confirmed that MSG has the ability to differ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
CPCA01N1/0221
Inventor 陈印平
Owner 陈印平
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