Suberythemogenic exposure of human skin treated with aqueous 8-MOP to radiation greater than 380 nm prolongs photosensitization to subsequent UVA from 6 to 24-72 h. One hypothesis for prolonged photosensitization is that greater than 380 nm irradiation forms 8-MOP-DNA monoadducts, which are removed more slowly than free 8-MOP and serve as a substrate for crosslinking by further UVA exposure. Sufficient DNA crosslinking results in erythema. We have examined this hypothesis by measuring the action spectrum for induction of prolonged photosensitization. Skin of normal volunteers was treated with aqueous 8-MOP (0.003%) and immediately received suberythemogenic monochromatic exposures between 334 and 430 nm. twenty-four hours later, the presence of prolonged sensitization was tested by small exposures of UVA. Erythema was evaluated 3 and 5 d later, and an action spectrum for prolonged sensitization was determined. The minimum phototoxic dose (MPD) was also determined at each wavelength. The action spectrum for prolonged photosensitization declined gradually between 334 and 425 nm. The action spectrum for delayed erythema induced by a single exposure of 8-MOP-treated skin declined rapidly from 334-390 nm. These findings are consistent with prolonged binding of 8-MOP in the skin by an initial exposure, possibly as 8-MOP-DNA monoadducts, allowing the second exposure to induce an erythemogenic event, possibly crosslinking of DNA.