Culture method of mesenchymal stem cells contained in umbilical cord blood

A technology of mesenchymal stem cells and culture methods, which is applied in the field of culture of umbilical cord blood containing mesenchymal stem cells, can solve the problem of low success rate of umbilical cord blood mesenchymal stem cell isolation, and achieve risk reduction, immunogenicity, good safety, No effects of exogenous contamination

Inactive Publication Date: 2021-08-13
上海南滨江细胞生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the success rate of umbilical cord blood mesenchymal stem cells is low. Therefore, it is a technical problem to be solved to provide a convenient, simple and high success rate in vitro culture method of umbilical cord blood mesenchymal stem cells.

Method used

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  • Culture method of mesenchymal stem cells contained in umbilical cord blood
  • Culture method of mesenchymal stem cells contained in umbilical cord blood

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Experimental program
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Effect test

Embodiment 1

[0028] A method for culturing umbilical cord blood containing mesenchymal stem cells, specifically comprising the following steps:

[0029] Step S1, collection and transportation of umbilical cord blood; umbilical cord blood comes from fetuses with normal full-term delivery in October 2020. The sample is approved by the hospital ethics committee and consented by the mother and family members before taking the sample. The amount of umbilical cord blood is 90ml; Serum volume fraction is 10%;

[0030] Step S2, extraction of mesenchymal stem cells: put umbilical cord blood and 6% hydroxyethyl starch aqueous solution in a volume ratio of 4:1 into a beaker, stir at room temperature at a speed of 25 r / min for 5 minutes, and then let stand for 30 minutes. Then take the supernatant, centrifuge at 1000r / min for 10min, discard the supernatant, resuspend the cells in the culture medium and add it to Ficoll separation medium containing an equal volume, centrifuge at 1800r / min for 20min, ta...

Embodiment 2

[0040] A method for culturing umbilical cord blood containing mesenchymal stem cells, specifically comprising the following steps:

[0041] Step S1, collection and transportation of umbilical cord blood; umbilical cord blood comes from fetuses with normal full-term delivery in October 2020. The sample is approved by the hospital ethics committee and consented by the mother and family members before taking the sample. The amount of umbilical cord blood is 90ml; Serum volume fraction is 15%;

[0042] Step S2, extraction of mesenchymal stem cells: put umbilical cord blood and 6% hydroxyethyl starch aqueous solution in a volume ratio of 4:1 into a beaker, stir at room temperature at a speed of 28 r / min for 5 minutes, and then let stand for 38 minutes. Then take the supernatant, centrifuge at 1000r / min for 10min, discard the supernatant, resuspend the cells in the culture medium and add it to Ficoll separation medium containing an equal volume, centrifuge at 1800r / min for 20min, ta...

Embodiment 3

[0052] A method for culturing umbilical cord blood containing mesenchymal stem cells, specifically comprising the following steps:

[0053] Step S1, collection and transportation of umbilical cord blood; umbilical cord blood comes from fetuses with normal full-term delivery in October 2020. The sample is approved by the hospital ethics committee and consented by the mother and family members before taking the sample. The amount of umbilical cord blood is 90ml; Serum volume fraction is 20%;

[0054] Step S2, extraction of mesenchymal stem cells: put umbilical cord blood and 6% hydroxyethyl starch aqueous solution in a volume ratio of 4:1 into a beaker, stir at room temperature at a speed of 30 r / min for 5 minutes, and then let stand for 45 minutes. Then take the supernatant, centrifuge at 1000r / min for 10min, discard the supernatant, resuspend the cells in the culture medium and add it to Ficoll separation medium containing an equal volume, centrifuge at 1800r / min for 20min, ta...

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Abstract

The invention discloses a culture method of mesenchymal stem cells contained in umbilical cord blood, belongs to the technical field of stem cell culture, and solves the problems that the success rate of separation of umbilical cord blood mesenchymal stem cells is low and the cell proliferation rate is low in the culture process in the prior art. The culture method specifically includes the following steps: step S1, collecting and transporting umbilical cord blood; step S2, extracting the mesenchymal stem cells; step S3, performing primary culture; step S4, performing subculture; and step S5, performing cryopreservation. According to the culture method, firstly, effective umbilical cord blood is acquired, sedimentation of red blood cells is accelerated by adding macromolecular hydroxyethyl starch with the combination of a density gradient centrifugation method, the success rate of separation of a single mesenchymal stem cell is increased; a specially-made culture medium used for the primary culture contains astragalus polysaccharide, and the astragalus polysaccharide can also maintain the phenotype and characteristics of the umbilical cord blood mesenchymal stem cells while promoting cell proliferation; and the culture method provided by the invention is simple to operate, free of exogenous pollution, safe and reliable and high in cell proliferation rate.

Description

technical field [0001] The invention belongs to the technical field of stem cell culture, and in particular relates to a culture method for umbilical cord blood containing mesenchymal stem cells. Background technique [0002] Umbilical cord blood mesenchymal stem cells are a type of adult stem cells isolated and cultured from umbilical cord blood, with high potential for self-renewal and multilineage differentiation. These characteristics of umbilical cord blood mesenchymal stem cells have attracted the attention of many scholars at home and abroad. At present, the clinical treatment of umbilical cord blood mesenchymal stem cell transplantation has made some progress. [0003] Umbilical cord blood refers to the blood that remains in the umbilical cord and placenta near the fetus after the fetus is delivered and the umbilical cord is ligated. There are two types of stem cells in cord blood: hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs). The possibility o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775A01N1/02
CPCC12N5/0665A01N1/0221C12N2509/00C12N2500/02C12N2500/90C12N2501/90C12N2533/52C12N2501/599C12N2500/84
Inventor 占震锋李庆静
Owner 上海南滨江细胞生物科技有限公司
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