We measured the
glutathione content of a panel of human
malignant melanoma cell lines by flow cytometry after staining with
mercury orange, using visible light (488 nm) excitation, and compared the values obtained with those measured biochemically using a modified Tietze assay.
Glutathione levels were depleted by culturing cells with various concentrations of
buthionine sulphoximine to provide a suitable spread of
glutathione concentrations. The two assays showed good correlation (r = 0.93). We found a number of technical factors to be critically important. In particular, the conditions of staining, cell number, and method of mixing media, cells and
stain were responsible for wide variations in fluorescence intensity. We applied the flow cytometric technique to cell
suspensions obtained by fine needle aspiration biopsy of human
malignant melanoma deposits, and observed a proportion of cells with high
glutathione levels in many samples. The results confirm the feasibility of measuring
glutathione content by visible light flow cytometry, and raise the possibility of monitoring
glutathione content as an
indicator of drug resistance in clinical
therapy of human
malignant melanoma.