Localized delivery of coding nucleic acids into adherent cells by in situ electroporation: integrated impedance-based monitoring allows for loss-of-function or gain-of-function screening
TL;DR
The combination of ISE, specific NAs and impedance-based cell monitoring paves the way for a new class of gain-of-function or loss-of-function experiments.
Abstract
Coding nucleic acids (NAs) like DNA, mRNA or siRNA hold endless potential to engineer cells with respect to their genome and proteome. However, minimally invasive delivery of NAs across an intact plasma membrane into the cytosol of living cells remains a technical challenge. Electroporation creates transient pores in the plasma membrane by applying well-defined electric fields for fractions of a second allowing NA transfer across the membrane. In situ electroporation (ISE) applies the electric field to adherent cells growing on the surface of planar, thin-film electrodes deposited on the bottom of the culture well. This study reports on the localized delivery of plasmid DNA, mRNA, siRNA and aptamers into different mammalian cell lines by ISE. Fluorescence microscopy verified successful delivery of labeled NAs themselves (siRNA, aptamers) or expression of the proteins they encoded (DNA, mRNA). Impedance readings of the cell-covered electrodes before and after ISE allowed tracking the invasiveness of the pulse, the recovery of the cells within 30 min and the phenotypic impact of the NAs inside the cells. Loading the cells with siRNA specifically designed to knock down gene transcripts essential for survival, led to the onset of cell death within 20 h after the pulse. The combination of ISE, specific NAs and impedance-based cell monitoring paves the way for a new class of gain-of-function or loss-of-function experiments.