Electroporation can be used as a tool for extracting or introducing molecules from or into a cell. The most important and promising applications of electroporation in medicine and biotechnology are described.

Gene electrotransfer is a complex phenomenon, where many factors mutually affect the process and the DNA-membrane interaction only comprises the first step. We showed that longer electric pulses are optimal for higher transfection efficiency but reduce viability, whereas shorter pulses enable moderate transfection efficiency and preserve viability. Thus, each application needs a careful choice of pulsing protocol.

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Knowledge of the parameters which influence the efficiency of gene electrotransfer has importance for practical implementation of electrotransfection for gene therapy as well as for better understanding of the underlying mechanism. The focus of this study was to analyze the differences in gene electrotransfer and membrane electropermeabilization between plated cells and cells in a suspension in two different cell lines (CHO and B16F1). Furthermore, we determined the viability and critical induced transmembrane voltage (ITV(c)) for both cell lines. In plated cells we obtained relatively little difference in electropermeabilization and gene electrotransfection between CHO and B16F1 cells. However, significant differences between the two cell lines were observed in a suspension. CHO cells exhibited a much higher gene electrotransfection rate compared to B16F1 cells, whereas B16F1 cells reached maximum electropermeabilization at lower electric fields than CHO cells. Both in a suspension and on plated cells, CHO cells had a slightly better survival rate at higher electric fields than B16F1 cells. Calculation of ITV(c) in a suspension showed that, for both electropermeabilization and gene electrotransfection, CHO cells have lower ITV(c) than B16F1 cells. In all cases, ITV(c) for electropermeabilization was lower than ITV(c) for gene electrotransfer, which is in agreement with other studies. Our results show that there is a marked difference in the efficiency of gene electrotransfer between suspended and plated cells.

Gene electrotransfer is a promising nonviral method that enables transfer of plasmid DNA into cells with electric pulses. Although many in vitro and in vivo studies have been performed, the question of the implied gene electrotransfer mechanisms is largely open. The main obstacle toward efficient gene electrotransfer in vivo is relatively poor mobility of DNA in tissues. Since cells are mechanically coupled to their extracellular environment and act differently compared to standard in vitro conditions, we developed a three-dimensional (3-D) in vitro model of CHO cells embedded in collagen gel as an ex vivo model of tissue to study electropermeabilization and different parameters of gene electrotransfer. For this purpose, we first used propidium iodide to detect electropermeabilization of CHO cells embedded in collagen gel. Then, we analyzed the influence of different concentrations of plasmid DNA and pulse duration on gene electrotransfer efficiency. Our results revealed that even if cells in collagen gel can be efficiently electropermeabilized, gene expression is significantly lower. Gene electrotransfer efficiency in our 3-D in vitro model had similar dependence on concentration of plasmid DNA and pulse duration comparable to in vivo studies, where longer (millisecond) pulses were shown to be more optimal compared to shorter (microsecond) pulses. The presented results demonstrate that our 3-D in vitro model resembles the in vivo situation more closely than conventional 2-D cell cultures and, thus, provides an environment closer to in vivo conditions to study mechanisms of gene electrotransfer.