Simplified ratiometric electrochemiluminescence aptasensor for carcinoembryonic antigen detection based on unmodified...

Ratiometric electrochemiluminescence (ECL) aptasensors have garnered significant interest in bioanalysis by simultaneously measuring two distinct emission signals and utilizing their intensity ratio for quantitative output. However, conventional designs face critical limitations. Mandatory aptamer functionalization elevates fabrication complexity and costs. Additionally, dual-luminophore/coreactant systems often suffer from inherent signal mutual interference. To address these issues, we developed a novel ratiometric ECL aptasensing platform employing a single luminophore (luminol) and coreactant (H₂O₂) for carcinoembryonic antigen (CEA) detection. This simplified system utilized an unmodified aptamer-G-quadruplex (G4)-hemin complex, which played two roles: (1) suppressing cathodic ECL (E_c) through hydroxyl radical consumption; (2) enhancing anodic ECL (E_a) via catalytic acceleration of luminol and H₂O₂ oxidation. Target-induced CEA-aptamer binding triggered G4-hemin dissociation, generating inversely correlated E_c enhancement and E_a reduction. This dynamic enabled quantitative CEA detection through the E_c/E_a ratio, achieving an ultrabroad linear range (10 fg mL^-1-10 ng mL^-1) with a detection limit of 0.85 fg mL^-1. Notably, the ratiometric signal is independent of H₂O₂ concentration. This effectively eliminates interference from H₂O₂ self-decomposition, significantly improving detection reliability. Successful application in human serum analysis validated clinical utility. This work establishes a new strategy for developing robust ratiometric ECL biosensors, potentially expanding their applications in clinical diagnostics and point-of-care testing.