Artificially mineralized bio-cascade enzyme-catalyzed probes for immunoassay of neuron-specific enolase with neuroblastoma diagnostics.

A novel artificial mineralized bioprobe was developed through a templated strategy, enabling highly sensitive and rapid detection of neuron-specific enolase (NSE) in serum samples. This probe was based on a nitrogen-doped carbon porous hollow shell as a carrier, loaded with Fe-Cu nanoparticles exhibiting highly efficient peroxidase-like activity. The Fe-Cu nanoparticles demonstrated outstanding catalytic decomposition of hydrogen peroxide, while the unique porous structure provided an ideal confined space for immobilizing the native enzyme. The acidic microenvironment generated by the enzymatic products of anchored glucose oxidase activates adjacent Fe-Cu bimetallic nanozymes, enabling cascade catalytic reactions under near-neutral conditions. Furthermore, due to the confining effect of the porous carbon shell layer, immobilized natural enzymes showed superior stability in response to temperature changes (10–50 °C) and during storage (30 days). In the presence of NSE targets, immunocomplex formed and catalyzed the cascade decomposition of substrate glucose, which attacked the chromogenic substrate to induce color change. Under optimized conditions, the developed sensor exhibited an ultra-wide linear response range (0.1–100 ng mL− 1) and a low detection limit (LOD = 0.0486 ng mL− 1). This work provides new insights for developing highly efficient enzyme-catalyzed probes.