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Method of purifying pluripotent vascular progenitor cells from perinatal tissue

A tissue cell and perinatal technology, applied in the field of biomedical research, can solve the problems of good repeatability and fast purification of multipotent vascular progenitor cells, achieve good expansion potential, and promote the effect of cardiovascular system repair

Active Publication Date: 2021-01-01
至仁生命科技(天津)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no fast and reproducible method of using immunomagnetic bead separation technology to enrich the target cell population through negative expression, and then further purify the pluripotent vascular progenitor cells by positive expression

Method used

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  • Method of purifying pluripotent vascular progenitor cells from perinatal tissue
  • Method of purifying pluripotent vascular progenitor cells from perinatal tissue
  • Method of purifying pluripotent vascular progenitor cells from perinatal tissue

Examples

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Embodiment 1

[0042] This embodiment exemplifies the method for purifying pluripotent vascular progenitor cells from perinatal tissues, including a pretreatment step and an acquisition step. The pretreatment step refers to processing the perinatal tissues to obtain perinatal tissue cells Suspension, the obtaining step includes: step A. adding the first immunomagnetic beads to the perinatal tissue cell suspension to label and enrich the pretreated perinatal tissue cells, and obtain Negatively expressed cell subpopulation suspension; wherein, the first immunomagnetic beads are CD31 immunomagnetic beads; step B. adding the second immunomagnetic beads to the cell subpopulation suspension prepared in step A to The cell subpopulation obtained in A is marked, enriched, and the positively expressed cell subpopulation suspension is obtained; wherein, the second immunomagnetic beads are CD146 immunomagnetic beads; the CD31 immunomagnetic beads and CD146 immunomagnetic beads are both Commercially avai...

Embodiment 2

[0045] This example exemplifies the method for purifying pluripotent vascular progenitor cells from perinatal tissue. On the basis of implementation 1, the perinatal tissue is selected from the umbilical cord, and the first immunomagnetic beads are CD31 immunomagnetic beads produced by Miltenyi. . Specifically: take 1 cm of the umbilical cord and separate the umbilical cord cells, place them in a PBS buffer system containing EDTA, add 20 μL CD31 immunomagnetic beads per 10 million cells, and shake well to make the umbilical cord cells and CD31 immunomagnetic beads complex; Then, the suspension containing the "umbilical cord cell-CD31 immunomagnetic bead complex" flows through the commercially available magnetic separator produced by Miltenyi, so that the "umbilical cord cell-CD31 immunomagnetic bead complex" is magnetically attracted, The umbilical cord cells that are not complexed with CD31 immunomagnetic beads exist in the suspension and flow out; add NG2 immunomagnetic bead...

Embodiment 3

[0048] This example exemplifies the method for purifying pluripotent vascular progenitor cells from perinatal tissue. On the basis of implementation 1, the perinatal tissue is selected from the umbilical cord, and the first immunomagnetic beads are CD31 immunomagnetic beads produced by Miltenyi. . Specifically: take 1 cm of the umbilical cord and rinse it with 75% ethanol for 1 minute, then wash it three times with PBS buffer containing 1% (v / v) penicillin; cut the washed umbilical cord to an average volume of 1.0 mm³; add 1 mg / ml type A collagenase (Roche) digested the chopped umbilical cord continuously at 37°C for 1.5 hours, passed through a 100um pore size filter to make a suspension containing 100,000-200,000 placental cells per 1mL, and press Add 20 μL of CD31 immunomagnetic beads containing 1 mmol / L EDTA per 10 million cells and shake well to make the umbilical cord cells and CD31 immunomagnetic beads complex; The suspension flows through the commercially available mag...

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Abstract

The invention provides a method for purifying pluripotent vascular progenitor cells from a perinatal tissue. The method comprises the steps of pretreatment and acquisition, wherein in the acquisitionstep, immunomagnetic beads of two different ligands are used for marking and purifying the perinatal tissue cells, the first immunomagnetic beads are used for marking firstly, through positive expression, non-pluripotent vascular progenitor cells are recognized, and impurity cells are removed to obtain a high-concentration cell population containing the pluripotent vascular progenitor cells, thenthe impurity-removed perinatal tissue cells are marked through the second immunomagnetic beads, and recognition is carried out through positive expression to obtain the pluripotent vascular progenitorcells. The pluripotent vascular progenitor cells prepared through the method have phenotypes similar to phenotypes of bone marrow mesenchymal stem cells and perivascular cells at the same time, havethe same mesoderm cell differentiation function as the bone marrow mesenchymal stem cells, further have the functions of differentiating into mature vascular cells and promoting cardiovascular systemrepair and regeneration, and can maintain the cell morphology and the amplification capability.

Description

technical field [0001] The invention belongs to the field of biomedical research, in particular to a method for purifying and obtaining multipotent vascular progenitor cells from perinatal tissues. Background technique [0002] Perinatal tissue refers to the accessory organs and tissues related to fetal development produced during the period from 28 weeks of pregnancy to the birth of the baby, mainly including cord blood, umbilical cord and placenta. The perinatal tissue after the fetus is delivered at full term has a small chronological age and a low gene mutation rate. It is usually discarded as biological waste and can be obtained without any risk to the fetus and mother. The acquisition method will not cause harm to the human body, etc. advantage. Perinatal tissue has become the focus of attention in the field of regenerative medicine because it contains a variety of stem cells and specific tissue progenitor cell subsets, and has many advantages such as abundant resourc...

Claims

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Application Information

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IPC IPC(8): C12N5/074C12N5/0789
CPCC12N5/0607C12N5/0647C12N2509/00
Inventor 贾惠东崔占峰
Owner 至仁生命科技(天津)有限公司
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