Intracellular calcium, in particular the cytosolic free ionized calcium concentration [Ca(2+)](i), is tightly regulated under physiological conditions. Stimulation of receptors, belonging to almost all the classes so far described, will result in changes in [Ca(2+)](i). These changes might be directly induced by either Ca(2+)-influx or Ca(2+)-mobilization from intracellular stores, or indirectly by a number of other mechanisms. The development of fluorescent indicators of free [Ca(2+)] that could be loaded into intact cells has contributed enormously to the understanding of cellular Ca(2+) homeostasis, especially dyes that respond to Ca(2+) with shifts of excitation or emission spectra. By measuring at two selected wavelengths (either dual emission or dual excitation), it is possible to calculate the proportion of dye in the Ca(2+)-bound and Ca(2+)-free forms.
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