Generation of a defined and uniform population of CNS progenitors and neurons from mouse embryonic stem cells
Generation of a defined and uniform population of CNS progenitors and neurons from mouse embryonic stem cells
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DOI：
10.1038/nprot.2007.147
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发表时间：
2007-01-01
期刊：
影响因子：
14.8
通讯作者：
Barde, Yves-Alain
中科院分区：
文献类型：
--
作者：
Bibel, Miriam;Richter, Jens;Barde, Yves-Alain
一个详细的协议，允许从小鼠胚胎干细胞（ES）的神经元的基本上纯的群体的产生。它是基于ES细胞的培养，通过反复分裂保持未分化，随后扩增为非粘附细胞聚集体。用视黄酸处理导致这些ES细胞基本上成为具有Pax6阳性放射状胶质细胞特征的神经祖细胞。正如它们在体内所做的那样，这些祖细胞分化成形成功能性突触接触的突触能锥体神经元，并且可以在培养物中保持很长一段时间。该方案不需要使用表达抗性或荧光标记的ES系，因此原则上可应用于任何野生型或突变体ES系。从开始ES细胞培养到接种祖细胞和分化的神经元在约10天内建立突触传递需要至少2周。
A detailed protocol is described allowing the generation of essentially pure populations of glutamatergic neurons from mouse embryonic stem ( ES) cells. It is based on the culture of ES cells that are kept undifferentiated by repeated splitting and subsequently amplified as non-adherent cell aggregates. Treatment with retinoic acid causes these ES cells to essentially become neural progenitors with the characteristics of Pax6-positive radial glial cells. As they do in vivo, these progenitors differentiate in glutamatergic pyramidal neurons that form functional synaptic contacts and can be kept in culture for long periods of time. This protocol does not require the use of ES lines expressing resistance or fluorescent markers and can thus be applied in principle to any wild-type or mutant ES line of interest. At least 2 weeks are required from starting ES cell culture until plating progenitors and differentiating neurons establish synaptic transmission within about 10 days.