Magnetic carbon dots nanozyme coupled in-situ growth strategy: Colorimetric/ratiometric fluorescent/photothermal multi-model detection of Microcystin-LR

Enzyme linked immunosorbent assay (ELISA) has become the gold standard assay format in multiple fields. Here, we develop a nano-catalyst-triggered cascade platform based on magnetic carbon dots nanozyme (MCDs) for colorimetric/ratiometric fluorescent/photothermal multi-model detection of Microcystin-LR (MC-LR). The multifunctional tracer (MCDs@PB) is first constructed by a simple in-suit shell-growing strategy. The synergy of MCDs and PB endows MCDs@PB with robust fluorescence emission, remarkable photothermal signal, favorable magnetic response, ultrahigh peroxidase mimicking activity along with superb antibody coupling efficiency. Notably, the Michaelis constant (K_m) and maximum reaction velocity (V_max) values of MCDs@PB are 0.56 mM and 11.12 × 10⁻⁸ M s⁻¹ for 3,3′,5,5′-tetramethylbenzidine (TMB), and 0.012 mM and 3.55 × 10⁻⁸ M s⁻¹ for hydrogen peroxide (H₂O₂), indicating that MCDs@PB have good affinity for both substrates. Leveraging this foundation, a triple-mode ELISA platform is designed. This platform consists of three parts: (1) catalytic amplification system triggered by TMB and H₂O₂, (2) ratiometric fluorescent analysis constructed by introducing rhodamine B into MCDs@PB-mediated ELISA system, and (3) photothermal assay achieved by the near-infrared laser-driven photothermal properties of oxidized TMB. The calculated limits of detection for MC-LR are 0.09 ng/mL in catalytic colorimetric mode, 0.12 ng/mL in ratiometric fluorescent mode, and 0.17 ng/mL in photothermal mode. The application feasibility of MCDs@PB-mediated ELISA is further verified in food and environmental samples with satisfactory recoveries. This work not only provides a new inspiration for the rational design of CDs-triggered signal tracers, but also exhibits their great potential in multimodal immunoassays.