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 conce ntration of free calcium ([Ca2+]) inside living cells. It should be ap preciated, however, that this is not what it actually is. More accurat ely; it is a divalent metal ion chelator which changes its fluorescenc e properties upon complexation. Thus, [Ca2+] has to be inferred indire ctly by means of the law of mass action. As a chelator, fura-2 may inf luence 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 t hem more directly related to its molecular properties: as a chelator, competing with endogenous Ca buffers, it can be used to estimate endog enous buffers and their properties. When present at sufficiently high concentration, such that it outcompetes endogenous buffers, fura-2 rep orts 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 meas urements derived from various methods will be compared.