Widely used as a fluorogenic substrate in enzymatic assays, particularly for detecting β-galactosidase activity. Upon enzymatic hydrolysis, it releases 4-methylumbelliferone, a highly fluorescent compound that allows sensitive and quantitative detection. This makes it ideal for reporter gene assays, high-throughput screening, and monitoring gene expression in molecular biology and microbiology. Commonly applied in studies involving recombinant protein expression, especially with the lacZ gene system. Also used in diagnostic applications to detect enzyme activity in cells or enzyme-labeled immunoassays. Its high sensitivity and low background fluorescence enhance detection accuracy in both solution-based and plate-based assays.