Biophysical properties of ergosterol-enriched lipid rafts in yeast and tools for their study: characterization of ergosterol/phosphatidylcholine membranes with three fluorescent membrane probes

► Three fluorescent membrane probes were used to study ergosterol-containing membranes. ► Fluorescent probes reflect the hydration behavior of ergosterol/phospholipid mixtures. ► Di-4-ANEPPS fluorescence properties are sensitive to sterol type and content. ► Di-4-ANEPPS fluorescence lifetime responds uniquely to sterol-lipid interactions. ► Several probe applications in the study of yeast membrane rafts are presented.