;Abstract In this study, polyethylenimine (PEI) and DNA were deposited onto conductive polypyrrole (PPy) substrate using layer-by-layer assembly. The formed polyelectrolyte multilayers (PEMs) films were released under electric field. The release results suggested that the electric field perpendicular to the substrate can efficiently release PEI and DNA from PEMs. Voltages were the main factors controlled the PEMs disassembly that higher potential released more PEI and DNA. The treatment period demonstrated similar effects. The surface morphologies of PEMs after electric field treatment were observed using atomic force microscopy (AFM). Compared to the nontreated group, the roughness of the films significantly increased after treating electric field, which was more obvious as the potential was high. These results suggested that the polyelectrolytes may be easily released from thin regions of the films due to the driving force of electric field as well as the dissolution resulted from electrochemical reaction on electrodes. Finally, the release DNA and PEI were used for transfection. The released material from the films with electric field treatment demonstrated superior transgene expression, suggesting that the electric assisted gene delivery from PEMs should be beneficial to gene delivery.