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Isolation method of adipose-primitive mesenchymal stem cells

A technology of mesenchymal stem cells and separation method, which is applied in the field of separation of adipose primitive mesenchymal stem cells, can solve the problems of inability to remove miscellaneous cells, unfavorable clinical application of cells, etc., and achieve the effect of complete digestion and uniformity, good cell activity, and improved purity

Active Publication Date: 2020-02-21
钜威细胞(厦门)医学科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Chinese patent ZL201210336264.5 discloses a "method for the isolation and culture of human adipose stem cells and the construction of a stem cell bank". Improve, but can not remove a large number of miscellaneous cells, cells need to be purified by continuous passage, making the cultured cells unfavorable for clinical application

Method used

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  • Isolation method of adipose-primitive mesenchymal stem cells
  • Isolation method of adipose-primitive mesenchymal stem cells

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Experimental program
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Effect test

Embodiment 1

[0029] Prepare P-type collagenase mother solution: add 1 g P-type collagenase to 100 ml of normal saline, mix well and filter, then dilute P-type collagenase mother solution with normal saline to a final concentration of 0.1% (m / v).

[0030] Obtain adipose tissue: Obtain superficial abdominal adipose tissue through tumescent liposuction, transport the obtained adipose tissue in an environment of 2-8°, and separate within 12 hours.

[0031] Wash adipose tissue: put the adipose tissue in a centrifuge tube, 20ml / tube, add normal saline to 40ml, centrifuge at 200rpm for 3min, discard the lower layer of blood, repeat the above steps once.

[0032] Digestion of adipose tissue: take 10ml of adipose tissue, add an equal volume of 0.1% (m / v) type P collagenase, mix well, and digest with shaking at 150rpm at 37°C for 50min.

[0033] Centrifugal collection: the digested product is filtered through a 150-mesh sieve, and the primary cell suspension is collected. After the cell primary susp...

Embodiment 2

[0036] Prepare P-type collagenase mother solution: add 1 g P-type collagenase to 100 ml of normal saline, mix well and filter, then dilute P-type collagenase mother solution with normal saline to a final concentration of 0.4% (m / v).

[0037] Obtain adipose tissue: Obtain superficial abdominal adipose tissue through tumescent liposuction.

[0038] The collected adipose tissue was transported in an environment of 2-8°, and the separation operation was performed within 12 hours.

[0039] Wash adipose tissue: put the adipose tissue in a centrifuge tube, 20ml / tube, add normal saline to 40ml, centrifuge at 200rpm for 3min, discard the lower layer of blood, repeat the above steps once.

[0040] Digestion of adipose tissue: take 10ml of adipose tissue, add an equal volume of 0.4% (m / v) type P collagenase, mix thoroughly, and digest with shaking at 150rpm at 37°C for 16min.

[0041] Centrifugal collection: the digested product is filtered through a 150-mesh sieve, and the primary cell...

Embodiment 3

[0044] Preparation of P-type collagenase mother solution: add 1 g P-type collagenase to 100 ml of normal saline, mix well and filter, then dilute P-type collagenase mother solution with normal saline to a final concentration of 1% (m / v).

[0045] Obtain adipose tissue: Obtain superficial abdominal adipose tissue through tumescent liposuction, transport the obtained adipose tissue in an environment of 2-8°, and separate within 12 hours.

[0046] Wash adipose tissue: put the adipose tissue in a centrifuge tube, 20ml / tube, add normal saline to 40ml, centrifuge at 200rpm for 3min, discard the lower layer of blood, repeat the above steps once.

[0047] Digestion of adipose tissue: take 10ml of adipose tissue, add an equal volume of 1% (m / v) type P collagenase, mix thoroughly, and digest with shaking at 150rpm at 37°C for 8min.

[0048] Centrifugal collection: the digested product is filtered through a 150-mesh sieve, and the primary cell suspension is collected. After the cell prim...

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Abstract

The invention discloses a separation method of original adipose mesenchymal stem cells. The separation method comprises the following steps: adopting collagenase P to digest adipose tissues, and performing centrifugal treatment so as to obtain adipose mesenchymal stem cells. The collagenase P (collagenageP) enables the adipose tissues to be digested more thoroughly, and the tissues are digested uniformly without reminding obvious tissues, so that the obtained mesenchymal stem cells are good in activity, quick in adherence and good in uniformity.

Description

technical field [0001] The invention relates to a method for separating adipose primitive mesenchymal stem cells. Background technique [0002] Stem cells are a kind of primitive cells with self-renewal ability and multi-lineage differentiation potential under suitable microenvironment. Stem cells can be divided into embryonic stem cells, pluripotent stem cells, and committed stem cells in various tissues such as hematopoietic stem cells, neural stem cells, etc., according to the sequence in which they appear during individual development. At present, there have been experimental reports on the use of stem cells to treat myocardial infarction, lupus erythematosus, rheumatoid arthritis, nerve damage, and muscle atrophy. [0003] Adult stem cells are a kind of superpotent stem cell population with multilineage differentiation potential existing in human tissues, which can be induced to differentiate into various tissue cells under specific circumstances. Among adult stem cel...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0775
Inventor 吴炯
Owner 钜威细胞(厦门)医学科技有限公司