Highly sensitive and accurate detection of chloramphenicol is of paramount importance for food safety. Herein, an enzyme-modulated photothermal immunosensor that uses a self-calibrated thermal imaging system (SCTIS) as signal read-out was developed for detecting chloramphenicol. In this immunosensor, alkaline phosphatase was used as a modulator of the photothermal conversion. It could hydrolyze the substrate into ascorbic acid, thereby reducing oxidized 3,3′,5,5′-tetramethylbenzidine, which exhibited a near-infrared laser-driven photothermal effect. For precise temperature measurement, the SCTIS was designed by using the temperature compensation of a ceramic chip to enable real-time self-calibration of the temperature. This SCTIS-based immunosensor could detect chloramphenicol with a LOD of 9 pg/mL in 2 h, and relative standard derivations from 3.95% to 13.58%. The average recoveries in milk and egg samples ranged from 76% to 114%. This versatile sensing strategy can detect various targets by altering recognition elements, thus has wide applicability in food safety testing and monitoring.
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