Optimizing electrotransfection of Mammalian cells in vitro.

INTRODUCTION This protocol describes transfection of plasmid DNA into mammalian cell lines using electroporation, a process whereby external application of electric pulses induce cell membrane permeability. A number of factors can affect electrotransfection efficiency. In general, cells in suspension and small volume cells are difficult to transfect, whereas adherent cells and large volume cells are relatively easy. Regardless of cell size or phenotype, transfection efficiency increases with a high concentration of cells in a small volume. The use of plasmid DNA larger than 13 kb reduces transfection efficiency. One important variable is the choice of electroporation buffer--the appropriate buffer can increase cell transfection efficiency 50-98%. Generally, buffers with low ionic constants enhance transfection efficiency.