P-Doped Cu–N–C Single-Atom Catalysts Boost Cathodic Electrochemiluminescence of Luminol for MicroRNA-320d Detection

Compared with efficient anodic luminol electrochemiluminescence (ECL), the disadvantage of cathodic ECL is that luminol cannot be electrochemically oxidized in a direct manner, and the conversion efficiency of dissolved oxygen (DO) as the coreactant to reactive oxygen species (ROS) is poor, which limits its application. Therefore, it is necessary to develop a functional catalyst suitable for the luminol-DO ECL system to directly trigger cathodic ECL. In this study, a coordination microenvironment modulation strategy was proposed. The heteroatom P was doped into the Cu single-atom catalyst (P/Cu SAs@CN), significantly enhancing the cathodic ECL emission in the luminol-DO system. The P/Cu SAs@CN catalyst facilitates a three-electron oxygen reduction reaction pathway, generating abundant ROS, particularly hydroxyl radicals, that amplify the ECL emission by oxidizing luminol anions. Simultaneously, a dual-signal amplification biosensor is developed by integrating APE1 enzyme-mediated target recycling and DNAzyme-catalyzed cleavage cycles. The biosensor achieves ultrasensitive detection of miRNA-320d, a biomarker of metastatic colorectal cancer, with the detection limit of 0.158 fM. This work not only elucidates the mechanistic link between the ORR-driven ROS generation and the ECL enhancement but also provides a versatile platform for designing advanced coreaction accelerators, which can help to address the critical demands in clinical diagnostics.