Method for culturing human neural stem cells

A neural stem cell and culture method technology, applied in nervous system cells, cell dissociation methods, microorganism-based methods, etc., can solve the problems of limited number of cases, many hybrid cells in neural stem cells, and insufficient sources of neural stem cells.

Pending Publication Date: 2022-04-19
北京银丰鼎诚生物工程技术有限公司 +1
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Problems solved by technology

[0004] At present, the following problems exist in the application of neural stem cells: most of the established neural stem cell lines are derived from mice, and there are obvious species differences between mice and humans; the number of cases is limited, the source of neural stem cells is insufficient, and cannot be isolated Pure neural stem cells; the sources of materials are often intact fetuses induced in the first month, and there are ethical issues; the embryonic brain tissue is small and very fragile, and it is difficult to obtain accurate materials during operation, resulting in more miscellaneous cells in the isolated neural stem cells

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  • Method for culturing human neural stem cells

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Embodiment 1

[0024] Example 1 isolation and culture of human neural stem cells

[0025] The steps are as follows:

[0026] (1) Select the artificial abortion embryo tissue that is intact and undamaged, no stasis, and no more than 4 hours away from the mother, peel off the brain tissue under sterile conditions, flush the residual blood, and then immerse the brain tissue in the cryopreservation protective fluid to quickly freeze it, so that the brain tissue is stereotyped.

[0027] The embryonic tissue is an embryonic tissue of 9 weeks to 10 weeks of gestation, the age of the mother is 20 to 35 years old, there are no contraindications to routine preoperative examination, and the mode of miscarriage is: mifepristone miscarriage two days in advance.

[0028] (2) Sterile forceps and scalpels are used to accurately separate the forebrain under the observation of the microscope, and then use sterile forceps to peel off the meninges to obtain pure forebrain tissue.

[0029](3) After the forebrain ti...

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Abstract

The invention discloses a method for culturing human neural stem cells, which comprises the following steps: (1) selecting an induced abortion embryonic tissue, stripping a brain tissue, immersing into a cryopreservation protective solution, and quickly freezing, so that the brain tissue is shaped; (2) accurately separating a forebrain part, and stripping a meninx to obtain pure forebrain tissues; (3) separating by a mechanical method to obtain pure neural stem cell tissues; (4) obtaining a single-cell suspension: inoculating the separated neural stem cell tissues into a complete neural dry culture medium, and culturing cells to obtain the single-cell suspension; and (5) subculture: changing the liquid once on the fourth day of culture, changing the liquid once on the eighth day, and subculturing on the 10th-12th day. According to the method, materials are taken from aborted embryos, ethical problems do not exist, the embryo forebrain can be accurately separated through the method, accurate material taking is achieved, and the method has important significance on establishment, research and application of human neural stem cell lines.

Description

Technical field [0001] The present invention relates to a method of culture of human neural stem cells, belonging to the field of stem cell technology. Background [0002] Stem cells are a class of multi-potential cells with the ability to self-replicate, and under certain conditions, they can differentiate into a variety of functional cells. Neural stem cells (NSCs) are a class of stem cells with multi-directional differentiation potential, which can self-renew for a long time and can differentiate into neurons, astrocytes and oligodendrocytes under certain conditions. [0003] It was previously thought that neurons in the central nervous system lost their ability to regenerate before or shortly after birth. However, some studies in recent years have shown that the brain tissue of adult mammals can still produce new neurons continuously, and NSCs are also present in adult brain tissue, mainly in the lateral subventricular layer (SVZ) and the hippocampal tooth gyrus. Neural stem...

Claims

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Application Information

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IPC IPC(8): C12N5/0797C12R1/91
CPCC12N5/0623C12N2509/00
Inventor 杨印祥陈小威周莹赵成
Owner 北京银丰鼎诚生物工程技术有限公司
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