The overuse of tetracycline (TC) antibiotics has raised significant environmental and health concerns, while traditional detection methods face challenges in real-time monitoring and intracellular analysis. In this study, sulfur-doped carbon dots (S-CDs) were synthesized through a one-step hydrothermal method, using biotin and o-phenylenediamine as precursors. The resulting S-CDs exhibit three key functional properties: (1) a highly sensitive fluorescent quenching response to TC, with a detection limit of 76.6 nM and a linear range of 0–200 μM, driven by specific molecular interactions (inner filter effect and static quenching); (2) pH-dependent fluorescence signaling (pH 2.0–8.0) attributed to protonation of surface acid-base groups; and (3) selective lysosomal targeting, with a Pearson correlation coefficient of 0.95, confirmed by confocal microscopy in HeLa cells and zebrafish models. The S-CDs demonstrated excellent recoveries (97.07–107.99 %) in real food samples (milk, beef, and pork) and exhibited negligible cytotoxicity. This work effectively integrates environmental pollutant detection with intracellular microenvironment analysis, providing a versatile tool for investigating TC-induced lysosomal dysfunction and advancing food safety monitoring and biomedical imaging.
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