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Purification method and identification method of peritoneal perivascular cells of mice

A purification method and perivascular technology, applied in the field of purification of peritoneal vascular pericytes in mice, can solve the problems of unreported peritoneal vascular pericyte extraction methods, complex labeled antibodies, high cost, etc., to prevent cell agglutination and save time , cost-saving effect

Inactive Publication Date: 2021-07-23
JIANGSU PROVINCIAL HOSPITAL OF TCM
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  • Abstract
  • Description
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Problems solved by technology

However, the extraction process of these cells has problems such as time-consuming, complicated antibody labeling, and high cost.
Moreover, in the field of peritoneal dialysis for renal diseases, the academic circle has not yet reported a mature method for extracting peritoneal peritoneal vascular pericytes.

Method used

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  • Purification method and identification method of peritoneal perivascular cells of mice
  • Purification method and identification method of peritoneal perivascular cells of mice
  • Purification method and identification method of peritoneal perivascular cells of mice

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Experimental program
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Effect test

Embodiment 1

[0085] 1. Experimental materials

[0086] 1.1 Mouse selection

[0087] C57BL / 6 mice (Victorian Lihua Experimental Animal Technology Co., Ltd., Beijing, China)

[0088] 1.2 Required reagents and experimental equipment

[0089] Cell source: ATCC cell bank ( CRL-2581 TM ), mouse vascular endothelial cell line (MVECs, C166-ATCC, USA), 1% penicillin-streptomycin (Invitrogen, USA), phosphate buffered saline (PBS, Invitrogen, USA), DMEM medium (Invitrogen, USA ), DMEM low glucose medium (Invitrogen, USA), type II collagenase (C6885, Sigma-Aldrich, USA), type I DNase (D5025, Sigma-Aldrich, USA), trypsin (25200072, Invitrogen, USA), EDTA (1340GR100, BioForxx, Germany), fetal bovine serum (FBS, Invitrogen, USA), pericyte growth supplement factor (PGS, Science Cell, USA), mouse FcR blocker (STEMCELL, Canada), CD140b / PDGFR- β monoclonal antibody (12-1402-81, Invitrogen, USA), Cocktail protease inhibitor (STEMCELL, Canada), Triton x-100 (ST795, Beyotime Biotechnology, China), α-SMA m...

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Abstract

The invention discloses a method for purifying peritoneal perivascular cells. The method comprises the following steps: (1) digesting and culturing mesentery through type II collagenase and type I deoxyribonuclease to obtain primary cells; and (2) digesting the primary cells obtained in the step (1) through trypsin, and sorting through immunomagnetic beads to obtain peritoneal perivascular cells. Compared with the prior art, the method provided by the invention is simple to operate, cost-saving, and time-consuming to obtain high-purity primary cells. Purity of the primary pericytes after conditioned culture reaches 88.5%, and the purity after combined MACS sorting reaches 97.8%. The method fills blank in the field of peritoneal pericytes and solves defects existing in other existing tissue pericyte extraction technologies.

Description

technical field [0001] The invention relates to the field of peritoneal vascular pericytes, in particular to a method for purifying and identifying mouse peritoneal vascular pericytes. Background technique [0002] Peritoneal dialysis (peritoneal dialysis, PD) is one of the main means of replacement therapy for end-stage renal disease. However, with the prolongation of dialysis time, stimulated by various pathological factors, the peritoneal structure remodels and peritoneal fibrosis (PF) progressively aggravates, leading to a gradual decline in dialysis efficiency, and eventually ultrafiltration failure, the patient has to withdraw from treatment , seriously restricting the popularization and development of PD technology. At present, the academic community generally believes that the activation and proliferation of myofibroblasts (MyoF), the main component of the mesothelial matrix, are the core factors in the pathological process of PF. [0003] Scholars at home and abro...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071G01N33/68G01N33/577G01N15/14
CPCC12N5/0679G01N33/74G01N33/68G01N33/6872G01N33/577G01N33/56966G01N15/14C12N2509/00
Inventor 盛梅笑史俊何伟明俞曼殊唐蕾单云
Owner JIANGSU PROVINCIAL HOSPITAL OF TCM
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