The aim of the present investigation was to test the hypothesis that atrial natriuretic factor (ANF) is secreted into the proximal tubule lumen by the organic anion transport mechanism. The rationale for this hypothesis was the reported probenecid attenuation of the natriuretic effect of ANF. Probenecid is widely regarded as an inhibitor of organic acid transport in the proximal tubule. ANF was prepared in varying degrees of purity ranging from a relatively crude extract to a highly purified form. A commercially available form was also used. All forms were bioassayed using the anesthetized rat and a diuresis and natriuresis was observed in each case which was comparable to literature reports. Interaction of ANF with the organic acid transport system was evaluated using the renal cortical slice technique. Over a wide range of concentrations, there was no effect of ANF on cortical slice accumulation of either p-aminohippurate (PAH), the classical substrate of the organic anion transport system or tetraethylammonium (TEA), a typical organic cation. It is concluded that although ANF may indeed exert its effect at the luminal membranes of the nephron, access to the lumen is not mediated by the organic cation or anion transport system in the proximal tubule.