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目的:建立有效的人类神经干细胞分离纯化系统和方法。方法:无菌取孕24周流产胎儿的室下区脑组织,制备细胞悬液,用磁性微珠标记的CD133单克隆抗体与细胞孵育,通过磁分选器分离出CD133(+)和CD133(-)细胞,通过体外培养并诱导分化,观察CD133阳性细胞和阴性细胞的增殖情况及分化能力。结果:经免疫磁珠分选的室下区脑组织中,CD133(+)细胞占胚胎脑组织细胞的2％左右,该细胞体外扩增能力强,细胞呈球形生长,并易聚集成团,培养5 d、10 d、15 d、20 d进行活细胞计数,细胞增长率分别为1.79％、4.82％、7.21％、14.66％,诱导分化后的细胞经染色鉴定可分化为神经元、神经胶质等多种神经细胞;阴性细胞扩增能力弱,活细胞计数以死细胞数量居多,大约70％,另外一些细胞贴壁分化。结论:免疫磁珠法简便、有效,经免疫磁珠分离的神经干细胞在体外能进一步培养扩增并分化。
Objective: To establish an effective human neural stem cell isolation and purification system and method. Methods: The abdomen area brains of aborted fetus were prepared 24 weeks after aseptic implantation. The suspension of cells was prepared. The cells were incubated with CD133 monoclonal antibody labeled with magnetic microbeads. CD133 (+) and CD133 -) cells were cultured and induced to differentiate to observe the proliferation and differentiation ability of CD133 positive cells and negative cells. RESULTS: The CD133 (+) cells accounted for about 2% of the embryonic brain cells in the subventricular zone by magnetic beads sorting. The cells were capable of expanding in vitro and were spherical in shape. After cultured for 5, 10, 15 and 20 days, the viable cells were counted and the cell growth rates were 1.79%, 4.82%, 7.21% and 14.66% respectively. After differentiation, the differentiated cells could differentiate into neurons, Quality and other neural cells; negative cells with weak ability to expand, living cells count to the majority of dead cells, about 70%, adherent cells adherent differentiation. Conclusion: Immunomagnetic beads method is simple and effective. Neural stem cells isolated by immunomagnetic beads can be further cultured, expanded and differentiated in vitro.