Styryl-based wavelength-ratiometric probes: a new class of fluorescent calcium probes with long wavelength emission and a large Stokes' shift.

We describe the chemical synthesis and fluorescence spectral characterization of a styryl-benzothiazole probe which contains the Ca2+ chelating group BAPTA as an integral part of the chromophoric system. The visible absorption spectrum of this probe displays a dramatic shift in the long wavelength maxima from 508 to 407 nm upon complexation with Ca2+, with a Ca2+ dissociation constant of 1.5 microM. The emission maximum centered at 615 nm is well shifted from the absorption. The emission spectrum displays a small blue shift upon binding Ca2+, allowing this probe to possibly be used as an emission wavelength-ratiometric probe using a single-excitation wavelength. This probe is likely to be the first of a series of long-wavelength ratiometric Ca2+ probes whose structure can be modified for improved quantum yield or altered Ca2+ affinity.