Transfection of primary central and peripheral nervous system neurons by electroporation.
Neurons are difficult cells to transfect. Many methods that work routinely for immortalized tissue culture cells or primary cultures of nonneuronal cells are ineffective, toxic, or both when applied to neurons. This chapter describes a protocol that optimizes electroporation-based transfection of chick embryonic peripheral and central neurons. The key features required for successful electroporation and recovery of transfected neurons are high cell density, correct applied voltage and pulse duration, and the presence of calcium ions in the electroporation medium. Less important features are temperature, postporation rest, and the general composition of the electroporation medium. We emphasize the rationale for each element in our method and provide information useful for optimizing the procedure for other neurons.
Martinez
Cecilia Y
CY
Department of Biological Sciences, Purdue University, West Lafayette, Indiana 47906, USA.
Hollenbeck
Peter J
PJ
D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D002478:Cells, Cultured / Q000166:cytology / Q000502:physiology*; D002490:Central Nervous System / Q000166:cytology / Q000502:physiology; D018274:Electroporation / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D017933:Peripheral Nervous System / Q000166:cytology / Q000502:physiology; D014162:Transfection / Q000295:instrumentation / Q000379:methods*