Transport of the prototypical organic cation tetraethylammonium (TEA) by the Malpighian tubules, ureters and gut of Drosophila melanogasterwas studied using two novel electrophysiological techniques. Both techniques exploited the high selectivity of the cation exchanger potassium tetra-p-chlorophenylborate for tetraalkylammonium compounds relative to inorganic cations such as K+. In the first technique, TEA fluxes were measured using a non-invasive self-referencing TEA-selective microelectrode positioned in the unstirred layer near the surface of each tissue. TEA fluxes from bath to lumen as large as 6 pmol cm–2s–1 were measured across the lower (reabsorptive) segment of the Malpighian tubule and the ureter bathed in saline containing 0.1 mmol l–1 TEA. Corresponding bath-to-lumen fluxes across the secretory main segment of the Malpighian tubule and the posterior midgut were∼1 pmol cm–2 s–1. TEA transport by the lower Malpighian tubule was enhanced by hyperpolarization of the basolateral membrane potential and was inhibited by cimetidine, quinidine, vinblastine and verapamil. In the second technique, TEA concentration was measured using a TEA-selective microelectrode positioned in droplets of fluid secreted by Malpighian tubules set up in saline droplets under oil in a Ramsay assay. Results from the Ramsay assay confirmed the dominant role of the lower Malpighian tubule in net transepithelial secretion of TEA and inhibition of TEA transport by cimetidine. Kinetic parameters (Jmax and Kt) were determined using both approaches.