Overcoming the Sensitivity-Accuracy Dilemma in Point-of-Care Testing via Cascade-Amplified Multidimensional Signal Integration Strategy Using Tetra-Enzyme-Mimicking Fe 2 O 3 /CoFe 2 O 4 Nanocubes

Despite the critical importance of a biomarker test for disease diagnosis and clinical management, existing strategies largely remain fundamentally constrained by an unresolved trade-off between analytical accuracy and independent of sophisticated instrumentation, limiting their applicability for rapid, user-friendly, and onsite biomarker monitoring. Herein, Fe 2 O 3 /CoFe 2 O 4 nanocubes (Fe 2 O 3 /CoFe 2 O 4 NCs) with exceptional tetra-enzyme-mimicking activities were prepared via a facile one-pot strategy and integrated as signal amplification elements into a trypsin-responsive hydrogel. By employing standard EP tubes and sample vials as miniature laboratories, a breakthrough tetramodal (barometric, colorimetric, thermal, and RGB) point-of-care testing (POCT) platform for trypsin detection was constructed. The sensing mechanism relies on a cascade catalytic process involving target-induced hydrogel degradation followed by nanozyme-mediated signal amplification. Noteworthily, trypsin-triggered gelatin hydrolysis releases Fe 2 O 3 /CoFe 2 O 4 NCs, thereby activating their multienzyme-mimetic functions. The pronounced peroxidase-like activity of Fe 2 O 3 /CoFe 2 O 4 NCs catalyzes the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB), generating a distinct colorimetric response accompanied by a near-infrared laser-induced photothermal effect, enabling dual readout via a smartphone and a portable thermometer. Concurrently, the catalase-like activity of Fe 2 O 3 /CoFe 2 O 4 NCs promotes H 2 O 2 decomposition in sealed vials, producing O 2 and resulting in a rapid increase in internal pressure that can be quantitatively monitored. Our strategy integrates cascade-amplified catalysis, multisignal transduction, and portable quantitation within a minimalist design, enhancing analytical sensitivity and reliability while enabling dose-customized detection. Crucially, this platform is readily generalizable, it can be adapted to other biomolecular targets by simply altering the responsive hydrogel, providing a versatile and robust paradigm for rapid and accurate biomarker sensing.