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FRET excitation ratiometric zinc sensing scheme: In the presence of zinc, dapoxyl sulfonamide binds to holo-CAII and transfers energy to the acceptor, AlexaFluor 594. The ratio of the acceptor fluorescence emission with excitation of the donor to that with excitation of the acceptor -is measured. The FRET intensity ratio increases with zinc concentration as the fraction of zinc-bound CAII increases, producing a binding curve with picomolar affinity (inset).
