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Method for extensive amplification of cell line and mesenchymal stem cell in vitro

A technology of mesenchymal stem cells and cell lines, applied in the field of expanding cell lines and mesenchymal stem cells, can solve the problems of limited number of umbilical cord mesenchymal stem cells and failure to achieve clinical application, and achieve easy measurement and monitoring, cost saving, Simple operation effect

Active Publication Date: 2012-01-18
UNION STEMCELL & GENE ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] However, how to obtain a large number of cells in the clinical application of stem cells has become a key issue, and it has also become a bottleneck problem, because both hematopoietic stem cells and umbilical cord mesenchymal stem cells are limited in cell number
Research on how to expand stem cells in large quantities in vitro has attracted more and more attention from researchers. Although some progress has been made, it is far from meeting the needs of clinical applications.

Method used

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  • Method for extensive amplification of cell line and mesenchymal stem cell in vitro
  • Method for extensive amplification of cell line and mesenchymal stem cell in vitro
  • Method for extensive amplification of cell line and mesenchymal stem cell in vitro

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1: (the following percentages are mass percentages)

[0042] Taking the African green monkey cells (COS1) as the object cells, the microcarrier culture was tried and finally compared with the total amount of cells that can be obtained by normal cell culture flask culture.

[0043] 1) Microcarrier treatment: Weigh 0.1g of microcarrier magnetic beads into a 50ml centrifuge tube, add 40ml of PBS to wash once, let it stand until all the magnetic beads have settled to the bottom, then gently suck out the PBS and then add 40ml of PBS to hydrate at 4°C for 3 hours . The hydrated microcarriers were sterilized by high-temperature steam at 121° C. for 30 minutes. The finally obtained microcarriers were soaked in 40ml of PBS for use.

[0044] 2) COS1 cell inoculation: After fully mixing the microcarrier mother solution, pipette 0.1 ml into 5 ml of DMEM medium and mix for use. The recovered COS1 cells were divided into 1*10 5Inoculate in 5ml DMEM medium (10% FBS / 1% d...

Embodiment 2

[0048] Rat adipose-derived mesenchymal stem cells (rADSC) were used as the target cells to try microcarrier culture and finally compared with the total amount of cells that can be obtained from normal cell culture flask culture, and the state of the cells was further detected by inducing differentiation of the obtained cells .

[0049] 1) Microcarrier treatment: Weigh 0.1g of microcarrier magnetic beads into a 50ml centrifuge tube, add 40ml of PBS to wash once, let it stand until all the magnetic beads have settled to the bottom, then gently suck out the PBS and then add 40ml of PBS to hydrate at 4°C for 3 hours . The hydrated microcarriers were sterilized by high-temperature steam at 121° C. for 30 minutes. The finally obtained microcarriers were soaked in 40ml of PBS for use.

[0050] 2) Isolation of rADSC: Obtain 1.5g of fat from the groin of 6-week-old Wistar rats under clean conditions; cut the adipose tissue into about 1cm with dissecting scissors 3 Large and small fr...

Embodiment 3

[0060] Taking African green monkey cells (COS1) as the target cells to try to use microcarriers for large-scale culture:

[0061] 1) Microcarrier treatment: Weigh 0.1g of microcarrier magnetic beads into a 50ml centrifuge tube, add 40ml of PBS to wash once, let it stand until all the magnetic beads have settled to the bottom, then gently suck out the PBS and then add 40ml of PBS to hydrate at 4°C for 3 hours . The hydrated microcarriers were sterilized by high-temperature steam at 121° C. for 30 minutes. The finally obtained microcarriers were soaked in 40ml of PBS for use.

[0062] 2) COS1 cell inoculation: After fully mixing the microcarrier mother solution, pipette 0.1ml and add it to 4ml DMEM medium, mix and set aside. The recovered COS1 cells were divided into 1*10 5 Inoculate in 5ml DMEM medium (10% FBS / 1% double antibody) at a density of / ml, aspirate and discard the liquid in the microcarrier tube, add the cell suspension to the microcarrier, mix well and put it in ...

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Abstract

The invention discloses a method for extensive amplification of cell line and mesenchymal stem cells in vitro. Microcarrier and Hyperflask cell culture container are combined for extensive amplification of cell line and mesenchymal stem cells in vitro. The invention combines commercialized microcarrier and Hyperflask for extensive amplification of cells in vitro in a small culture space, so as to obtain a large amount of target cells on the premise of saving costs.

Description

technical field [0001] The invention relates to a method for expanding cell lines and mesenchymal stem cells, in particular to a method for amplifying cell lines and mesenchymal stem cells in large quantities in vitro by using microcarriers combined with Hyperflask. Background technique [0002] In recent years, the research on stem cells has been deepened day by day. With the continuous accumulation of relevant knowledge, stem cells have become a new weapon for human beings to fight diseases. At present, studies on the multi-directional differentiation ability of stem cells have found that stem cells from different tissues can be applied to the treatment of organ and tissue diseases. The research on the treatment of various diseases has achieved relatively exciting results in the treatment of cardiovascular diseases, nerve damage diseases and metabolic diseases. [0003] However, how to obtain a large number of cells in the clinical application of stem cells has become a k...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/00C12N5/0775
Inventor 黄家学陈晓波韩洪起侯士芳张宇光韩俊领陈勇
Owner UNION STEMCELL & GENE ENG
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