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Title: In situ bipolar electroporation for localized cell loading with reporter dyes and investigating gap junctional coupling
Authors: De Vuyst, Elke ×
De Bock, Marijke
Decrock, Elke
Van Moorhem, Marijke
Naus, Christian
Mabilde, Cyriel
Leybaert, Luc #
Issue Date: Jan-2008
Publisher: Biophysical Society
Series Title: Biophysical Journal vol:94 issue:2 pages:469-79
Abstract: Electroporation is generally used to transfect cells in suspension, but the technique can also be applied to load a defined zone of adherent cells with substances that normally do not permeate the plasma membrane. In this case a pulsed high-frequency oscillating electric field is applied over a small two-wire electrode positioned close to the cells. We compared unipolar with bipolar electroporation pulse protocols and found that the latter were ideally suited to efficiently load a narrow longitudinal strip of cells in monolayer cultures. We further explored this property to determine whether electroporation loading was useful to investigate the extent of dye spread between cells coupled by gap junctions, using wild-type and stably transfected C6 glioma cells expressing connexin 32 or 43. Our investigations show that the spatial spread of electroporation-loaded 6-carboxyfluorescein, as quantified by the standard deviation of Gaussian dye spread or the spatial constant of exponential dye spread, was a reliable approach to investigate the degree of cell-cell coupling. The spread of reporter dye between coupled cells was significantly larger with electroporation loading than with scrape loading, a widely used method for dye-coupling studies. We conclude that electroporation loading and dye transfer is a robust technique to investigate gap-junctional coupling that combines minimal cell damage with accurate probing of the degree of cell-cell communication.
ISSN: 0006-3495
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Odisee General
× corresponding author
# (joint) last author

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