Strong Self-Off Biosensor Based on Proximity-Induced DNA Strand Displacement Triggered Controlled Release for Dual-Mode Sensitive Analysis of Neuron Specific Enolase

The effective improvement of sensitivity and accuracy is crucial for the immunoassay of biological targets. Based on this, an ultrasensitive self-off biosensor using Au-Ag nanoclusters (Au-Ag NCs) as luminophores was developed by integrating proximity-induced DNA strand displacement (PISD), controlled release, and resonance energy transfer (RET) strategies for trace analysis of neuron specific enolase. Because of the synergistic properties between bimetallic atoms, Au-Ag NCs showed a higher electrochemiluminescence (ECL) efficiency. At the same time, the cyclic conversion between Fe²⁺/Mn³⁺ and Fe³⁺/Mn⁴⁺ greatly catalyzed the reduction of S₂O₈^2-, thereby producing a large amount of SO₄^•- for ECL emissions. Second, the specific binding between the target antigens and antibodies could drive PISD to open the blocked pores, leading to the release of quenching probes and achieving signal self-off through the RET effect. Third, the driven PISD also triggered the detachment of the electrochemical signal probes, resulting in the self-off of the differential pulse voltammetry (DPV) signal. The PISD enhanced the anti-interference ability of the sensing system through target recognition and proximity triggering. Meanwhile, by combining controlled release to achieve signal self-change, background noise was suppressed, thus improving the detection sensitivity. In addition, the induced DNA strand displacement reaction could connect multiple signal output modes, effectively improving the accuracy of the detection results. The detection limits of the constructed biosensor in ECL and DPV modes were 3.23 and 31.8 fg/mL, respectively (<i>S</i>/<i>N</i> = 3), which provides a reference for the construction of sensitive and reliable biosensing platforms and the immunoassay of multiple biomarkers.