Chitosan-platinum nanozyme-mediated ratiometric fluorescent immunosensor with dual catalytic cascades for sensitive detection of fenitrothion

Organophosphorus pesticide residues pose significant threats to food safety and environmental health, and their monitoring methods demand sensitivity and anti-interference capability. In this study, we developed a novel ratiometric fluorescent immunosensor for fenitrothion (FN) detection by employing chitosan-modified platinum nanoparticles (Ch-Pt NPs) as catalytic nanozymes. Ch-Pt NPs exhibit oxidase-mimicking activity, enabling the oxidation of ascorbic acid (AA) and o -phenylenediamine (OPD) to produce dehydroascorbic acid (DHAA) and 2,3-diaminophenazine (DAP, Em = 568 nm), respectively. Then, DHAA further reacted with the remaining OPD to generate quinoxaline derivatives (DFQ, Em = 430 nm). These two products further form ratiometric fluorescent signals. Taking AA as a breakthrough point, the dual-catalytic cascade of natural enzymes and nanozymes was achieved by the combination with the alkaline phosphatase (ALP) -based enzyme-linked immunosorbent assay (ALP-ELISA). Compared with the conventional colorimetric ELISA, the proposed sensor demonstrated a nearly 20-fold enhancement in sensitivity (detection limit: 0.48 ng/mL) and achieved satisfactory recoveries of 80.0–108.3 % in spiked samples. The ratiometric fluorescent immunosensor demonstrates outstanding performance and holds great potential for extensive applications in the field of pesticide monitoring.