This chemical is widely used as a substrate in enzymatic assays, particularly for alkaline phosphatase activity. It serves as a chromogenic agent, where the enzyme catalyzes the hydrolysis of the compound, producing a yellow-colored product, 4-nitrophenol, which can be quantified spectrophotometrically. This application is crucial in biochemical research, clinical diagnostics, and pharmaceutical studies for measuring enzyme activity and screening inhibitors. Additionally, it is employed in immunoassays and histochemical staining to detect specific biomolecules or cellular components. Its stability and sensitivity make it a reliable tool in various laboratory settings.