DISCRETE CHANGES OF THE FLUORESCENCE YIELD FROM CELLS VITALLY STAINED WITH ETHIDIUM-BROMIDE (EB), AS DETERMINED BY FLOW CYTOMETRY

Cultured L cells were scraped from the glass surface and directly suspended in solutions of ethidium bromide (EB) in Tris buffer. After different staining times the fluorescence distributions were analysed with a flow cytometer. At 5 min after staining the cells yielded a normal DNA-specific histogram, but then a second peak at about 30-50% beyond the normal G1 fluorescence appeared and grew with increasing staining time. This effect of a discrete transition into a state of higher fluorescence was not dependent on the proliferative state of the cultures but was influenced by the temperature of the suspension and by serum and RNase. The effect proved not to be correlated with a G0-G1 transition and shows the possibility of artefacts in the determination of G0 G1 ratios by vital staining. © 1979.