Transfection of rat or mouse neurons by biolistics or electroporation
Transfection of rat or mouse neurons by biolistics or electroporation
复制标题
DOI：
10.1038/nprot.2009.90
复制
发表时间：
2009-01-01
期刊：
影响因子：
14.8
通讯作者：
Waxman, Stephen G.
中科院分区：
文献类型：
--
作者：
Dib-Hajj, Sulayman D.;Choi, Jin Sung;Waxman, Stephen G.
离子通道的特性受表达它们的细胞的背景影响。因此，研究这些蛋白质在有丝分裂后神经元中的作用对于研究神经元功能具有重要意义。然而，有丝分裂后的神经元和许多细胞系难以通过标准方法检测。在这里，我们提供了详细的协议，两个不同的程序，生物枪和电穿孔，这已被用来从小鼠或大鼠的外周感觉神经元与电压门控钠通道的表达结构。可以在48小时内制备、转染神经元并记录电流。使用这些方法，可以以5- 20%的效率转染初级感觉神经元，这允许在天然神经元细胞背景中研究钠通道及其天然存在的突变体的生物物理性质。虽然我们在这里展示的例子中使用了钠通道，但这些方法也可以用于研究其他类型的分子。
Properties of ion channels are affected by the background of the cells in which they are expressed. Thus, it is important for investigators interested in neuronal function to study these proteins in post-mitotic neurons. However, post-mitotic neurons, and many cell lines, are difficult to transfect by standard methods. Here we provide detailed protocols for two different procedures, biolistic and electroporation, which have been used to transfect peripheral sensory neurons from mice or rats with expression constructs of voltage-gated sodium channels. Neurons can be prepared, transfected and currents recorded within 48 h. Using these methods, primary sensory neurons can be transfected with an efficiency of 5-20%, which has permitted studying biophysical properties of sodium channels and their naturally occurring mutants in a native neuronal cell background. Although we have used sodium channels for the examples that we show here, these methods can also be used to study other types of molecules.