Properties of membranes of intact P388 murine lymphoblastic leukemia and a sub-line selected for resistance to adriamycin (P388/ADR) were examined using two fluorescent probes: diphenylhexatriene (DPH) and trimethylammonium diphenylhexatriene (T-DPH). Time-dependent changes in dye accumulation, fluorescence anisotropy, and lifetimes were measured. Accumulation of DPH, which eventually labels all cellular lipids, increased with time. Uptake of T-DPH, a more membrane-specific probe, reached a maximum in less than 1 min. No alteration could be detected in fluorescence anisotropy or lifetime of either dye associated with anthracycline resistance or anthracycline treatment. The rate of uptake of the T-DPH was not different in P388 versus P388/ADR cell lines, suggesting no differences in membrane "traffic". Calcium-channel antagonists could partially reverse anthracycline resistance in P388/ADR, but this was not accompanied by alterations in fluorescence parameters.