ELECTROPORATION BY USING BIPOLAR OSCILLATING ELECTRIC-FIELD - AN IMPROVED METHOD FOR DNA TRANSFECTION OF NIH 3T3 CELLS

Using the plasmid DNA pSV2-neo (which, when integrated into the cellular genome confers resistance to the antibiotic G418 for selection), we examined and compared the transfection efficiency on NIH 3T3 cells electro-permeabilized by applying a sequence of high-frequency unipolar or bipolar square waves or a single square pulse. Results show that a bipolar square wave is, at least, 1.7- and 5.5-fold more efficient than the unipolar square wave and single square pulse, respectively. In the range of electric field strength used for optimum transfection, the survivability of electro-permeabilized cells was comparable between the unipolar and bipolar square waves but fell considerably with the single square pulse. Qualitative comparison of cell permeabilization induced by the three types of wave forms and monitored by ethidium bromide uptake revealed that only the bipolar square wave permeabilizes the cell membrane symmetrically at the two hemispheres facing the electrodes. With unipolar square wave or single square pulse, the membrane is permeabilized either on one side or asymmetrically. Taken together, our results suggests that permeabilization of the membrane at multiple sites without affecting cell survivability may account for the improvements in transfection efficiency observed with bipolar oscillating electric fields.