Controllable Methylene Blue Release/Enrichment System for Ultrasensitive Electrochemical Detection of α-Fetoprotein

Tackling the critical challenge of low-abundance biomarker detection in electrochemical sensing, where traditional methods suffer from insufficient sensitivity and high background signals, we present a cascade signal amplification strategy. This approach integrates DNA-gated mesoporous silica nanoparticles (MSNs) for target-responsive release of methylene blue (MB) and G-Quadruplex DNA (G4)-functionalized nanoporous gold electrodes for MB enrichment via strong end stacking to form a stable G4/MB complex. RecJ_f exonuclease-mediated target recycling amplifies signals by enabling continuous MB release from MSNs. This sensing strategy achieves a detection limit of 2.14 fg/mL for α-fetoprotein across 10 fg/mL to 100 ng/mL, with high selectivity in serum samples. The universality of this strategy was confirmed by detecting carcinoembryonic antigen via a modular Anc/Apt design. Its practicability was validated by a clinical sample analysis. By combining controllable MB release and enhanced enrichment along with efficient electrochemical signal output, this low-background platform offers a universal strategy for ultrasensitive biomolecule detection, paving the way for its extension to diverse clinical biomarkers.