Widely used as a substrate in enzyme assays, particularly for detecting and quantifying alkaline phosphatase activity. When dephosphorylated by alkaline phosphatase, it produces 4-nitrophenol, which is yellow in color under basic conditions, allowing for easy spectrophotometric detection at around 405 nm. This reaction makes it valuable in clinical diagnostics, research laboratories, and enzyme-linked immunosorbent assays (ELISAs). Its high sensitivity and clear color change enhance its utility in colorimetric detection systems. Commonly included in diagnostic kits and biochemical test kits due to its stability and reliable performance.