Trehalose improves survival of electrotransfected mammalian cells

Background: Electropermeabilization is widely used for introduction of DNA and other foreign molecules into eukaryotic cells. However, conditions yiel ding the greatest molecule uptake and gene expression can result in low cel l survival. In this study, we assessed the efficiency of trehalose for enha ncing cell viability after excessive electropermeabilization. This disaccha ride was chosen because of its capability of stabilizing cell membranes und er various stressful conditions, such as dehydration and freezing. Materials and Methods: Various mammalian cell lines were electropermeabiliz ed by single exponentially decaying electric Pulses of few kV/cm strength a nd of several-microsecond duration. Propidium iodide (PI) and a plasmid enc oding green fluorescent protein (GFP), respectively, served as reporter mol ecules. The effects of trehalose on PI-uptake, GFP gene expression, transfe ction yield, and short- and long-term viability were analyzed by flow cytom etry and electronic cell counting. Results: The substitution of inositol by trehalose in pulse media protected cells against field-induced cell lysis. The protection effect saturated at about 40 -50 mM trehalose. Transfection yield and gene expression were not significantly affected by trehalose. But the transfection efficiency was g enerally higher in the presence of trehalose, mainly because of the increas ed cell survival. Conclusions: We demonstrated that trehalose-substituted media are superior to standard trehalose-free pulse media for improving cell survival and achi eving higher electrotransfection efficiency. Cytometry 45:161-169, 2001. (C ) 2001 Wiley-Liss, Inc.