This study assessed whether electroporation enhances transport across intact intestinal epithelial monolayers that mimic the intestinal epithelium. Confluent Caco-2 monolayers were exposed to electroporation pulses and then monitored over time for transepithelial transport of calcein, a small fluorescent tracer, or fluorescein-labeled bovine serum albumin, a large protein. Cumulative transport of both molecules across the monolayers increased significantly (up to 34-fold) after electroporation and depended on electroporation voltage and pulse length and on molecular size. Increased transport was accompanied by a decrease in the transepithelial electrical resistance of the monolayers. Further analysis of these results suggests that the increase in transport observed after electroporation is due, at least in part, to the killing of a small fraction of cells, which increased transport across "leaky" dead cells that remained adherent and increased transport through small, temporary holes left by dead cells that detached, but appeared to reseal within minutes by monolayer restitution. These findings could form the basis for the development of electroporation as a clinical tool to increase intestinal permeability and, thereby, increase the absorption of poorly absorbed drugs.