Brain microvascular endothelial cell extraction and purification method

A technology for endothelial cells and brain microvessels, which is applied in the directions of vascular endothelial cells, cell dissociation methods, biochemical equipment and methods, etc., can solve the problems of low activity and purity, and low yield, and achieves simple operation steps, high yield, and low stimulating effect

Active Publication Date: 2021-10-29
GUANGDONG NO 2 PROVINCIAL PEOPLES HOSPITAL
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Problems solved by technology

[0004] The present invention provides a method for extracting and purifying cerebral microvascular endothelial cells. In order to solve the defects of low yield, low activity and low purity in the current purification technology, the inventor adopted CD31 immunomagnetic beads for multiple sorting through repeated tests and improvements. , reduce the impact on the activity of brain microvascular endothelial cells, high separation efficiency, and successfully explored a relatively simple, high-yield, high-purity and high-viability purification method with stable repeatability

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  • Brain microvascular endothelial cell extraction and purification method
  • Brain microvascular endothelial cell extraction and purification method
  • Brain microvascular endothelial cell extraction and purification method

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Embodiment 1

[0056] Study on the extraction and purification method of high yield, high activity and high purity brain microvascular endothelial cells

[0057] 1. Main materials of the experiment:

[0058] 1) C57BL / 6 mice.

[0059] 2) Papain: brand Worthington, model LS003119, working solution concentration 100U / mL, used to break the extracellular matrix connecting cells.

[0060] 3) Endothelial cell culture medium, referred to as ECM, 50mL ECM includes the following components:

[0061] Heparin: brand Sigma, model H3393, 250 μL;

[0062] Glutamine: brand Lonza, model 17-605F, 500 μL;

[0063] F12 medium: brand Sigma, model 51651C, 43.75mL;

[0064] 100x penicillin-streptomycin: brand Corning, model 30-004, 500 μL;

[0065] FBS: brand Gibco, model C0232, 5mL;

[0066] Endothelial cell growth supplement: brand Millipore, model 02-102, 1.5 mg.

[0067] 4) CD31 immunomagnetic beads.

[0068] 5) F12 animal cell culture medium: F12 for short, brand Sigma, model 51651C.

[0069] 6) Panc...

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Abstract

The invention discloses a brain microvascular endothelial cell extraction and purification method, and relates to the technical field of cell purification. The extraction and purification method comprises the following steps: S1, coarsely extracting brain microvascular endothelial cells in brain tissues of a mouse; S2, carrying out primary CD31 immunomagnetic bead sorting on a brain microvascular endothelial cell mixture treated in the step S1 for purification; and S3, carrying out secondary CD31 immunomagnetic bead sorting on brain microvascular endothelial cells treated in the step S2 for purification so as to obtain the purified brain microvascular endothelial cells. According to the method, a mode of carrying out CD31 magnetic bead sorting for twice is adopted, damage to the activity of the brain microvascular endothelial cells is reduced, the separation and purification efficiency is high, the operation steps are relatively simple, and the brain microvascular endothelial cells with high purity, high yield and high activity can be obtained.

Description

technical field [0001] The invention relates to the technical field of cell extraction, in particular to a method for extracting and purifying cerebral microvascular endothelial cells. Background technique [0002] Brain microvascular endothelial cells, referred to as BMECs, are the main components of the blood-brain barrier and the target cells of various physiological and pathological factors. Separated from blood, restricts access of most polar molecules and proteins to brain tissue. Its culture was refined in the late 1970s, and it is also an important model for the study of blood-brain barrier and various diseases of brain neurology. The separation and purification of BMECs provides effective technical support for the study of the physiological and pathological characteristics of brain endothelial cells. BMECs cells have many characteristics, such as low cell activity, and the isolated cells are easily contaminated by fibroblasts, smooth muscle and other miscellaneous ...

Claims

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Application Information

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IPC IPC(8): C12N5/071
CPCC12N5/069C12N2509/10C12N2509/00C12N2501/91C12N2500/32C12N2529/00
Inventor 黄志新刘新通卢海克
Owner GUANGDONG NO 2 PROVINCIAL PEOPLES HOSPITAL
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