From Serum to Organ: The Amine-Naphthalene Modular Platform Enables Precise Identification and Quantification of Alkaline Phosphatase

Accurate detection of alkaline phosphatase (ALP) is critical for clinical diagnosis and mechanistic studies across diverse tissues. Here, we present a modular “off-on” fluorescent probe platform based on 1-naphthol phosphate sodium (1-NPP) and ethylenediamine (EDA), enabling precise, rapid, and cost-effective quantification of ALP activity from serum to whole organs. The probe’s mechanism relies on the ALP-catalyzed hydrolysis of 1-NPP to liberate 1-naphthol, which subsequently reacts with EDA to generate a robust fluorescent signal. This system demonstrates high sensitivity, with a detection limit of 0.068 mU/mL, and maintains excellent linearity and stability across complex biological matrices including serum, cell lysates, and tissue homogenates. Application in various cell lines and mice organs revealed organ- and disease-specific ALP activity distributions, highlighting its utility for distinguishing physiological and pathological states. The probe’s low cytotoxicity and straightforward preparation offer significant advantages over nanoparticle-based sensors. Overall, this work establishes a versatile and scalable platform for ALP analysis, whose remarkable ability to distinguish ALP activity across cells and organs underpins its broad potential for early disease surveillance, mechanistic exploration, and high-throughput screening.