New Thiol Active Fluorophores for Intracellular Thiols and Glutathione Measurement

Cellular thiols, especially glutathione (GSH), act as scavenger nucleophiles and can protect cells against: hyperoxia, toxicity, many carcinogens and mutagenicity or transformation by radiation. A technique which combines the use of a cell membrane permeable thiol reactive fluorophore with that for cell sorting by flow cytometry (FACS) has been used as a means of quantitating glutathione in individual cells or in cell subpopulations1,2. These studies are aiding the understanding of the role of GSH and other protein thiols in viable cells. Although many fluorescent thiol probes exist, few are useable in viable cells. We report here the results of tests of six newly synthesized naphthopyranone fluorophore based thiol probes3,4 in V79 Chinese hamster lung fibroblasts. Time and concentration dependence of dye uptake was evaluated, as were subcellular localization, dye cytotoxicity, and specificity of the dyes for GSH relative to other cellular thiols.