A Comparison of the Sensitivity of Photodamage Assays in Rat Basophilic Leukemia Cells

Abstract

Many aspects of cellular function or physiology can be used to indicate the level of damage resulting from the application of potentially deleterious agents such as drugs, solvents or even light. The dose required to reach a specific biological endpoint will necessarily depend on the characteristics of the damage induced by the agent. By using multiple biological probes, it is possible to get a more complete description of the type of damage induced. Photodamage was induced in rat basophilic leukemia cells by either 254-nm UVC light exposure or rose bengal photosensitization. Damage was measured by three quantitative assays employing fluorescent probes: calcein, to measure nonspecific esterase activity, propidium iodide (PI), to measure loss of plasma membrane integrity, rhodamine 123 (R123) to measure mitochondrial depolarization, and the incorporation of 5'-bromodeoxyuridine (BrdU), to measure the progress of cell replication. BrdU incorporation was found to be the most sensitive indicator for both forms of photodamage. For UVC photodamage, the BrdU assay was 330 times more sensitive than the other two assays. For rose bengal photosensitization, the BrdU assay was 48 or 62 times more sensitive than either the R123 or calcein/PI assays, respectively.