Isolation of neural stem cells from neural tissues using the neurosphere technique.
Isolation of neural stem cells from neural tissues using the neurosphere technique.
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DOI：
10.1002/9780470151808.sc02d06s15
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发表时间：
2010-11-01
影响因子：
--
通讯作者：
Vescovi, Angelo Luigi
中科院分区：
文献类型：
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作者：
Ferrari, Daniela;Binda, Elena;Vescovi, Angelo Luigi
本单元描述了从成年小鼠心室下区（mNSCs）、胚胎小鼠大脑和人胎儿大脑（hNSCs）中提取、表征和扩增神经干细胞（NSC）系的方法。通过酶切中枢神经系统（CNS）的特定区域（NSCs壁龛）可以分离出NSCs并在悬浮中生长。通过使用这种方法，NSCs形成称为神经球的球形簇，将其机械分离为单细胞悬浮液，并在选择性培养基中复制。去除生长因子并将细胞置于贴壁基质上，使细胞分化为神经元、星形胶质细胞和少突胶质细胞，这是中枢神经系统的主要细胞类型。根据这种方法，正确培养NSCs将允许细胞扩增超过100代，而不会改变细胞的核型、生长能力和分化潜力。
This unit describes protocols for the derivation, characterization, and expansion of neural stem cell (NSC) lines from the adult mouse subventricular zone (mNSCs), embryonic mouse brain and from the human fetal brain (hNSCs). NSCs can be isolated by enzymatic digestion of specific regions (NSCs niches) of the central nervous system (CNS) and grown in suspension. By using this methodology, NSCs form spherical clusters called neurospheres, which are mechanically dissociated to a single-cell suspension and replated in the selective culture medium. Removal of growth factors and plating cells on an adherent substrate allows cells to differentiate into neurons, astrocytes, and oligodendrocytes, the main cell type of the CNS. Correct culturing of NSCs, according to this methodology, will allow cells to expand over 100 passages without alteration of cell karyotype, growth ability, and differentiation potential.