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Cold Spring Harb Protoc. 2017 Feb 1;2017(2). doi: 10.1101/pdb.prot094904.

Single-Cell Electroporation of Neurons.

Author information

1
Institute for Synaptic Physiology, Center for Molecular Neurobiology (ZMNH), 20251 Hamburg, Germany.
2
Institute for Synaptic Physiology, Center for Molecular Neurobiology (ZMNH), 20251 Hamburg, Germany thomas.oertner@zmnh.uni-hamburg.de.

Abstract

Single-cell electroporation allows the transfection of a small number of neurons in an organotypic culture with a single plasmid or a defined mixture of plasmids. Desired protein expression levels can vary depending on the experimental goals (e.g., high expression levels are needed for optogenetic experiments); however, when too much protein is expressed, cellular toxicity and cell death may arise. To a large degree, protein expression can be controlled by adjusting the concentration of plasmid DNA in the electroporation pipette. Here, we present a protocol for transfecting individual neurons in hippocampal slice cultures by electroporation. Essentially, a patch-clamp setup is required that includes an upright microscope with infrared differential interference contrast or Dodt contrast with a camera and a specialized amplifier that is able to deliver large-voltage pulses to the electroporation pipette.

PMID:
28148856
DOI:
10.1101/pdb.prot094904
[Indexed for MEDLINE]

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