A dual signal ratio electrochemical sensor based on DNA tetrahedrons and bimetallic nanocomposites for AFB₁ analysis

Given the hepatotoxicity and widespread contamination of Aflatoxin B₁ (AFB₁), developing ultra-sensitive, anti-interference analytical platforms is paramount for public health. Herein, a dual signal ratio electrochemical sensing platform for accurate AFB₁ analysis was constructed integrating DNA tetrahedrons (TNDA), a dynamic DNA walker, and nitrogen-doped graphene oxide-supported hollow silver-platinum bimetallic nanospheres (NGR-HP-AgPt). The NGR-HP-AgPt cavity provides a uniform microenvironment, synergistically promoting catalytic effects to significantly accelerate interfacial electron transfer. Simultaneously, rigid 3D TNDA prevent spatial probe entanglement, providing a well-oriented track. To avoid false-positive artifacts in complex matrices, a competitive binding strategy is employed. Upon target recognition, AFB₁ displaces a complementary sequence (DNA1) from the aptamer. The released DNA1 acts as a walking strand, hybridizing with signal probes (Cd²⁺-DNA2). Subsequently, Exonuclease III initiates the DNA walker, continuously cleaving probes to amplify the ratiometric signal variation for reliable self-calibration. Under optimal conditions, this sensor exhibits a broad linear range from 2 × 10^-4 to 20 ng/mL, with an ultra-low detection limit of 73.99 fg/mL. Furthermore, its practical utility and high accuracy were successfully validated in complex food and medicinal matrices, yielding consistent results with the standard HPLC-FL method. This work broadens the robust design paradigm of anti-interference biosensing.