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A method for constructing a human umbilical cord mesenchymal stem cell bank

A technology of quality stem cells and construction methods, which is applied in the field of stem cell bank construction, can solve the problems of loss and discarding, and achieve the effects of less cell damage, simple operation, and fast construction methods

Active Publication Date: 2012-02-22
章毅 +7
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the umbilical cord tissue produced along with the umbilical cord blood is discarded, and the irrecoverable and important resource of umbilical cord mesenchymal stem cells contained in it is lost in vain

Method used

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  • A method for constructing a human umbilical cord mesenchymal stem cell bank
  • A method for constructing a human umbilical cord mesenchymal stem cell bank

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Construction of human umbilical cord mesenchymal stem cell bank:

[0025] (1) After confirming the safety of the human umbilical cord through strict testing procedures (ABO / Rh blood type testing, HLA typing testing, syphilis antibody testing, HIV immune testing, CMV antibody testing, hepatitis B antigen antibody testing, etc.), the buffer solution Rinse repeatedly to remove residual blood, and cut it to about 1mm with a sterile knife 3 -1.5mm 3 small pieces;

[0026] (2) Take 10ml of small fragments, put them in a 50ml centrifuge tube, add 10ml of 0.05% collagenase, and digest at 37°C for 1 hour to obtain the digested mixture;

[0027] (3) Add phosphate buffer (PH=7.2-7.4) to the digested mixture until it fills the 50ml centrifuge tube, mix well, and centrifuge at 600g for 15min;

[0028] (4) Discard the supernatant, add 20ml of phosphate buffer (PH=7.2-7.4), resuspend the precipitate, pass through a 200-mesh sieve, collect the filtrate, centrifuge at 450g for 10min,...

Embodiment 2

[0031] Construction of human umbilical cord mesenchymal stem cell bank:

[0032] (1) After confirming the safety of the human umbilical cord through strict testing procedures (ABO / Rh blood type testing, HLA typing testing, syphilis antibody testing, HIV immune testing, CMV antibody testing, hepatitis B antigen antibody testing, etc.), the buffer solution Rinse repeatedly to remove residual blood, and cut it to about 1mm with a sterile knife 3 -1.5mm 3 small pieces;

[0033] (2) Take 10ml of small fragments, put them in a 50ml centrifuge tube, add 20ml of 0.1% collagenase, and digest at 37°C for 2 hours to obtain the digested mixture;

[0034] (3) Add phosphate buffer (PH=7.2-7.4) to the digested mixture until it fills the 50ml centrifuge tube, mix well, and centrifuge at 600g for 15min;

[0035] (4) Discard the supernatant, add 20ml of phosphate buffer (PH=7.2-7.4), resuspend the precipitate, pass through a 200-mesh sieve, collect the filtrate, centrifuge at 450g for 10min,...

Embodiment 3

[0038] Construction of human umbilical cord mesenchymal stem cell bank:

[0039] (1) After confirming the safety of the human umbilical cord through strict testing procedures (ABO / Rh blood type testing, HLA typing testing, syphilis antibody testing, HIV immune testing, CMV antibody testing, hepatitis B antigen antibody testing, etc.), the buffer solution Rinse repeatedly to remove residual blood, and cut it to about 1mm with a sterile knife 3 -1.5mm 3 small pieces;

[0040] (2) Take 10ml of small fragments, put them in a 50ml centrifuge tube, add 30ml of 0.2% collagenase, and digest at 37°C for 3h to obtain the digested mixture;

[0041] (3) Add phosphate buffer (PH=7.2-7.4) to the digested mixture until it fills the 50ml centrifuge tube, mix well, and centrifuge at 600g for 15min;

[0042] (4) Discard the supernatant, add 20ml of phosphate buffer (PH=7.2-7.4), resuspend the precipitate, pass through a 200-mesh sieve, collect the filtrate, centrifuge at 450g for 10min, and ...

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Abstract

The invention discloses a method for constructing an umbilical cord mesenchymal stem cell bank using the umbilical cord of a newborn as a resource, aiming at fewer steps, simpler and easier operations when constructing the umbilical cord mesenchymal stem cell bank. The present invention consists of the following steps: (1) taking a human umbilical cord for detection, washing with a buffer to remove residual blood, and cutting into pieces to obtain human umbilical cord tissue; (2) adding collagenase to digest; (3) adding buffer to dilute and digest (4) Discard the supernatant, add buffer to resuspend the pellet, sieve, collect the filtrate, centrifuge, and repeat twice to obtain human umbilical cord mesenchymal stem cells; (5) The obtained human umbilical cord The mesenchymal stem cells are stored in liquid nitrogen, and stored according to their ABO / Rh typing and HLA typing, and a cell information file for retrieval is established, that is, a human umbilical cord mesenchymal stem cell bank is constructed. Compared with the existing method, the method of the present invention is faster, more convenient, and less damage to cells.

Description

technical field [0001] The invention relates to a method for constructing a stem cell bank, in particular to a method for constructing a human umbilical cord mesenchymal stem cell bank. technical background [0002] In addition to human hematopoietic stem cells in human bone marrow, there is also a cell population with stem cell characteristics, called human mesenchymal stem cells (MSCs). Human mesenchymal stem cells have the potential of self-renewal and multidirectional differentiation, and can differentiate into three germ layer cells under different induction conditions, such as bone cells, chondrocytes, muscle cells, fat cells and nerve cells. In addition, MSCs also express IL6, G-CSF and SCF and other factors required for the growth of hematopoietic cells, and can maintain the proliferation of long-term cultured hematopoietic progenitor cells, showing that MSCs can support hematopoiesis and promote hematopoietic recovery. At the same time, studies have shown that MSCs...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02C12N5/08
Inventor 高峰钱燕翔陈亮
Owner 章毅
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