Abstract: Disclosed are sensitive methods for detecting a target substance in low concentrations. The method can include immobilizing a target substance to a solid phase; incubating the immobilized target substance with a reporter enzyme detection probe in solution under conditions for forming target: enzyme detection probe complexes; washing the solid phase to remove any unbound reporter enzyme detection probe; incubating the target: enzyme detection probe complex with a reporter enzyme detection probe substrate in substrate reaction solution to generate one or more ionizable products; and detecting one or more of the one or more ionizable products using mass spectrometry (MS).