Unlike FDG that requires a two-step hydrolysis to generate maximum fluorescence, resorufin β-D-galactopyranoside requires only a single-step hydrolysis reaction to attain full fluorescence. This substrate is especially useful for sensitive enzyme measurements in ELISAs. The relatively low pKa (~6.0) of resorufin (the enzymatic hydrolysis product of resorufin galactoside) with Ex/Em=573/585 nm permits continuous measurement of enzymatic activity. Resorufin galactoside has also been used to quantitate β-galactosidase activity in single yeast cells by flow cytometry and to detect immobilized β-galactosidase activity.
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