Rhodamine WT, a xanthene dye used in the tracing of pollutants in water and in related studies, was tested for its mutagenicity in a battery of in vitro and in vivo mammalian assays. Using Chinese hamster ovary cells in the absence of metabolic activation mix, small dose-related increases in cytotoxicity, DNA damage (as detected by alkaline sucrose-gradient sedimentation) and sister-chromatid exchanges were detected, but an increase in the level of chromosomal damage was not seen. In the presence of metabolic activation a small, but statistically significant dose-related increase in sister-chromatid exchanges was evident, with no increase in cytotoxicity, DNA damage or chromosome aberrations. Furthermore, no increase in bone marrow micronuclei or sperm abnormalities was observed in male B6C3F1 mice. The data from all these mammalian assays, although involving different end-points, are in contrast to the mutagenic effects previously seen in Salmonella.