The use of fura-2 for estimating Ca buffers and Ca fluxes.

The compound fura-2 (Grynkiewicz et al., J. Biol. Chem. 260, 3440-3450, 1985) is generally known as an indicator dye for measuring the concentration of free calcium ([Ca2+]) inside living cells. It should be appreciated, however, that this is not what it actually is. More accurately, it is a divalent metal ion chelator which changes its fluorescence properties upon complexation. Thus, [Ca2+] has to be inferred indirectly by means of the law of mass action. As a chelator, fura-2 may influence the quantity of interest, the Ca signal. On the other hand, the chelator action may be used for a number of other purposes, some of them more directly related to its molecular properties: as a chelator, competing with endogenous Ca buffers, it can be used to estimate endogenous buffers and their properties. When present at sufficiently high concentration, such that it outcompetes endogenous buffers, fura-2 reports total Ca changes and is a probe for Ca fluxes across the membrane. Here, theory and methodological considerations of such applications of fura-2 will be summarized and results on Ca buffer and Ca flux measurements derived from various methods will be compared.