Several double-chain ammonium amphiphiles were designed to have physicochemical properties, such as membrane fluidity and aggregate morphology, suitable for DNA transfection, according to our previous results (1, 2). Potency of the amphiphiles during the transfer of plasmid DNA into COS cells was examined. Of the amphiphiles tested, O,O'-ditetradecanolyl-N-(trimethylammonio acetyl)diethanolamine chloride (14Dea2) had the highest transfection activity. Optimal conditions for transfection of COS cells by 14Dea2 were also determined.