Syntheses and Characterizations of 4-(3,17 β-Dihydroxyestra-1,3,5(10)-trien-6α- and 6β-yl)amino-7-nitro-2,1,3-benzoxadiazoles as Fluorescent Probes

4-(3,17β-Dihydroxyestra-1,3,5(10)-trien-6α- and 6β-yl)amino-7-nitro-2,1,3-benzoxadiazoles have been synthesized and characterized as fluorescent probes for use in a receptor assay and/or a homogeneous immunoassay for estradiol. The fluorescence intensities are strongly dependent upon the solvent polarity used. The intensities in water were reduced to less than 1% of those in ethyl acetate, and a blue shift was also observed in polar solvents. The quenched fluorescence in aqueous solution was recovered by adding bovine serum albumin or an anti-estradiol antibody. Adding intact estradiol inhibited the fluorescence recovered by the antibody.