This report deals with the in vitro dissolution of cholesterol gallstones by bile salts, as observed with the scanning electron microscope. Scanning electron microscopic techniques permit excellent visualization of the substructure of these stones and hence allow insight into the structural basis of gallstone dissolution. The small faceted, mixed cholesterol gallstone was used since this is the most common type clinically. Scanning electron microscopy revealed that the gallstone has four structurally different zones; an outer surface, a cortex of compactly layered cholesterol plates, an inner layer of cholesterol mixed with other components, and a center or nidus. The cortex is of particular importance in terms of gallstone dissolution with bile salts, since the interior of the stone remains unaffected until this zone is removed. The speed and completeness of dissolution of the stone depends mainly upon the thickness of the cortex and in part upon the content of noncholesterol components in the interior of the stone. Chenodeoxycholate and cholate behave similarly in terms of the dissolution process as observed by scanning electron microscopy. The solubilization process is very similar to that observed in vitro with solvents such as ether and ethanol. Stones treated with 400 mM cholate were completely solubilized in 3 weeks. The process described in this paper is probably similar to that which occurs in vivo in spontaneous gallstone dissolution in man and in patients treated with bile salts for gallstone dissolution.