1. A technique for the radioautographic identification, localization, and study of the turnover of cellular depots of 5-hydroxytryptamine (5-HT) has been evaluated. A light microscopic survey was made of the uptake and turnover of 5-HT, synthesized in vivo from its administered tritium-labelled precursor, 5-hydroxytryptophan (5-HTP).2. 5-HT was taken up rapidly and retained for long periods of time by adrenal medullary chromaffin cells, gastric enterochromaffin cells, blood platelets, thyroid parafollicular cells, beta cells of pancreatic islets, mast cells, and septal cells of the lung. Reticulo-endothelial cells of liver and spleen took up radioactive 5-HT more slowly but also retained it for days.3. Specific uptake, and a rapid turnover of radioactive 5-HT, was found in pancreatic exocrine cells, neurones of the superior cervical ganglion, terminal axons of gastro-intestinal myenteric plexus, carotid body cells, and pinealocytes.4. A supranuclear localization of radioactive 5-HT in renal proximal tubular cells may represent tubular secretion of 5-HT.5. The localization of 5-HT by radioautography generally corresponds to that found by formaldehyde-induced fluorescence. It makes possible the study of 5-HT turnover and is capable of greater resolution than the fluorescence technique. Freeze-drying may also be avoided.