(233e) In Vitro Gene Delivery through Membrane Sandwich Electroporation

To avoid safety issues such as immune response and cytotoxicity associated with viruses and liposomes, physical methods have been widely used for either in vivo or ex vivo gene delivery. They are, however, very invasive and often provide limited efficiency. Here we demonstrate a new electroporation-based gene delivery method, called membrane sandwich electroporation. The nanoscale pores in the support membranes allow a focused electric field on the cell membrane, and thus enhance cell permeabilization at a low electric voltage. Using pEGFP and pSEAP plasmids and NIH 3T3 fibroblasts as models, the MSE method shows significant improvement over transgene expression of mammalian cells compared to current electroporation techniques. To explain why the MSE method promotes transgene delivery, we acquired multiple vertical z-stack images using a spinning disk confocal microscope. The results demonstrated that the membrane sandwich configuration is able to provide better gene confinement near the cell surface to facilitate genes transport into the cells. The applicability of the MSE method to primary cells and hard-to-transfect cells (such as mouse embryonic stem cells and human blood mononuclear cells) is currently under investigation.