Spatially and potential-resolved anti-interference ECL biosensor for simultaneous detection of ochratoxin A and aflatoxin B1.

The simultaneous detection of multiple targets using electrochemiluminescence (ECL) is often hampered by the interference between lumiphores. To address this challenge, a spatially and potential-resolved ECL biosensor was developed for the detection of ochratoxin A (OTA) and aflatoxin B1 (AFB1). Specially, graphitic carbon nitride @ silicon dioxide composite (C3N4@SiO2) and gadolinium-based metal-organic framework (Gd-MOF) were selected as cathodic and anodic emitters, respectively, and covalently immobilized in separate electrode zones to prevent cross-interference. In addition, tetrahedral DNA (TDN) scaffolds were anchored to both emitters, followed by conjugation with Fc-modified aptamer (for AFB1) and Au NP-modified aptamer (for OTA), creating a dual-functional detection interface. In the presence of targets, in the cathode zone, Fc quenched C3N4@SiO2 ECL, while AFB1 binding reversed this quenching, increasing the signal; in the anode zone, AuNPs enhanced Gd-MOF ECL via surface plasmon resonance (SPR) effect, while OTA binding caused signal reduction through aptamer complex dissociation. Optimized conditions enabled independent detection, with linear ranges of 0.01-120 μg/kg for both toxins (LODs: 1.5 × 10-4 μg/kg for OTA and 1.9 × 10-5 μg/kg for AFB1 at S/N = 3).