The detection of cyclobutane thymine dimers, (6-4) photolesions and the Dewar photoisomers in sections of UV-irradiated human skin using specific antibodies, and the demonstration of depth penetration effects

Abstract

Ultraviolet irradiation of skin induces various DNA photolesions. Here we demonstrate that irradiation of human buttock skin with 300 nm UVR in situ induces thymine dimers and 6-4 photoproducts. Irradiation with 260 nm immediately followed by UVA (320 nm) induces the Dewar photoisomers of the 6-4 lesions. All three lesions can be detected in methanol-fixed paraffin sections using specific monoclonal antibodies. The sections have been analysed in an automated image analysis system (Discovery) and the level of immuno-DAB-peroxidase measured in individual epidermal cell nuclei as absorption at 460 nm (integrated optical density). The staining patterns with the antibodies showed no detectable change with epidermal depth by eye after 300 nm irradiation, however, the machine detected a fall off with depth of about 2.5% per cell layer. Following irradiation with a shorter wavelength (260 nm) there was a rapid fall off in staining with depth easily detectable by eye and machine (39% per cell layer).