Abstract: We investigated gene electrotransfer using electrical short-circuiting via a cell suspension droplet in dielectric oil. An aqueous droplet of a few microliters placed between a pair of electrodes can be deformed by an intense DC electric field depending on the electric field intensity. When a droplet containing suspended cells and plasmid DNA elongates during deformation and connects the electrodes, the resulting short circuit can cause successful gene electrotransfection into various mammalian cells. We also investigated the influence of the electroporation medium on membrane permeabilization and the mechanisms of gene electrotransfection using short-circuiting via an aqueous droplet. One aim of this study was to investigate the influence of the conductivity of electroporation medium on gene electrotransfer stimulated by short-circuiting. It was found that low-conductivity medium with plasmid DNA resulted in a significant decrease in cell viability compared to the high-conductivity medium with plasmid DNA. Therefore, we demonstrated the influence of exogenous DNA on membrane damage stimulated by droplet electroporation using a low-conductivity medium. Thus, electrical stimulation with the combination of plasmid DNA and the low-conductivity medium resulted in tremendous membrane damage. Linearized plasmid DNA stimulated more significant membrane damage than circular DNA. However, the size of linear DNA did not influence the efflux of small intracellular molecules.
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