Method for separating and enriching vascular endothelial progenitor cells from peripheral blood

A technology for the separation and enrichment of endothelial progenitor cells, which is applied in the field of vascular cell separation, can solve the problems of large differences in separation efficiency, proliferation potential, and unstable results of separation cells, and achieve less miscellaneous cells, high separation efficiency, and good separation effect Effect

Inactive Publication Date: 2016-06-22
SHANDONG MEIJIA SAIPEI BIOLOGICAL TECH CO LTD
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  • Abstract
  • Description
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Problems solved by technology

[0008] In order to solve the above existing problems in the separation efficiency and proliferation potential of EPCs in the prior art, the present invention aims to establish a standard, universal and efficient method for separating and enriching vascular endothelial progenitor cells from peripheral blood. The optimal conditions in the separation process have been explored to make the separation results comparable among various indicators, solve the problem of unstable separation cell results, improve the yield of separation and enrichment cells, and be used as the basis for the cell physiological characteristics of EPC Research and clinical applications for vascular repair and regeneration

Method used

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  • Method for separating and enriching vascular endothelial progenitor cells from peripheral blood
  • Method for separating and enriching vascular endothelial progenitor cells from peripheral blood
  • Method for separating and enriching vascular endothelial progenitor cells from peripheral blood

Examples

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Embodiment 1

[0037] A method for isolating and enriching vascular endothelial progenitor cells from peripheral blood, comprising the following steps:

[0038] (1) Obtain 30mL of peripheral blood samples to be separated and enriched (processed within 2 hours), invert and mix once in a centrifuge tube, and dilute 1:1 with PBS phosphate buffer solution of pH 7.2 to obtain diluted blood. Add PBS phosphate buffer solution to the tube, and then add peripheral blood sample;

[0039](2) Take four 50mL centrifuge tubes, add 15mL of hydroxyethyl starch solution with a mass / volume percentage concentration of 5.0% to each, and slowly add 20mL of diluted blood to the hydroxyethyl starch solution to ensure that the diluted blood remains in the hydroxyethyl starch solution. The upper layer of ethyl starch solution, at room temperature 375r / min, centrifuged for 35min, after the end, absorb the mononuclear cell fluid of the middle buffy coat cell layer, about 20mL, and combine the suction of each two tubes...

Embodiment 2

[0047] A method for isolating and enriching vascular endothelial progenitor cells from peripheral blood, comprising the following steps:

[0048] (1) Obtain 60mL of peripheral blood samples to be separated and enriched (processed within 2 hours), invert and mix once in a centrifuge tube, and dilute 1:1 with PBS phosphate buffer solution of pH 7.2 to obtain diluted blood. Add PBS phosphate buffer solution to the tube, and then add peripheral blood sample;

[0049] (2) Take four 50mL centrifuge tubes, add 15mL of hydroxyethyl starch solution with a mass / volume percentage concentration of 3.5% to each, and slowly add 20mL of diluted blood to the hydroxyethyl starch solution to ensure that the diluted blood remains in the hydroxyethyl starch solution. The upper layer of ethyl starch solution, room temperature 425r / min, centrifuged for 30min, after the end, absorb the mononuclear cell fluid of the middle buffy coat cell layer, about 20mL, and combine the suction of each two tubes i...

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Abstract

The invention relates to the technical field of vascular cell separation, in particular to a method for separating and enriching vascular endothelial progenitor cells from peripheral blood. The method comprises the steps of obtaining a peripheral blood sample, and using PBS phosphate buffer for dilution; adding diluted blood into a hydroxyethyl starch solution, performing centrifuging and sucking monocular cell sap of a middle albuginea cell layer; using the PBS phosphate buffer for dilution, performing centrifuging, abandoning a supernatant, and performing resuspension on residues through a nutrition solution; performing culture transferring. The method is high in separation efficiency, adopts few parenchyma cells, only needs a small number of experiment instruments, is good in separation effect and has the repeatability.

Description

technical field [0001] The invention relates to the technical field of vascular cell separation, in particular to a method for separating and enriching vascular endothelial progenitor cells from peripheral blood. Background technique [0002] EPC (endothelial progenitor cells, vascular endothelial progenitor cells) is a kind of precursor cells that can circulate, proliferate and differentiate into vascular endothelial cells, but have not yet expressed the phenotype of mature vascular endothelial cells, nor have they formed blood vessels. The physiological morphology of the cells cultured in vitro is a cobblestone-like endothelial cell layer, and the general surface markers identified by EPC are: CD45(-), CD34(+), KDR(+), CD31(+). [0003] After the identification and characterization of EPC cell populations derived from peripheral blood, angiogenesis in adults has also been reported. A large number of literature reports are emphasizing the importance of EPC to maintain the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/0692C12N2509/00
Inventor 韩发彬
Owner SHANDONG MEIJIA SAIPEI BIOLOGICAL TECH CO LTD
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