Caffeine interaction with fluorescent calcium indicator dyes

We report that caffeine, in millimolar concentrations, interacts strongly w ith four common calcium indicator dyes: mag-fura-2, magnesium green, fura-2 , and fluo-3. Fluorescence intensities are either noticeably enhanced (mag- fura-2, fura-2) or diminished (magnesium green, fluo-3). The caffeine-induc ed changes in the fluorescence spectra are clearly distinct from those of m etal ion binding at the indicator chelation sites. Binding affinities for c alcium of either mag-fura-2 or magnesium green increased only slightly in t he presence of caffeine. Caffeine also alters the fluorescence intensities of two other fluorescent dyes lacking a chelation site, fluorescein and sul forhodamine 101, implicating the fluorophore itself as the interaction site for caffeine. In the absence of caffeine, variation of solution hydrophobi city by means of water/dioxane mixtures yielded results similar to those fo r caffeine. These observations suggest that hydrophobic substances, in gene ral, can alter dye fluorescence in a dye-specific manner. For the particula r case of caffeine, and perhaps other commonly used pharmacological agents, the dye interactions can seriously distort fluorescence measurements of in tracellular ion concentrations with metal indicator dyes.