Orthogonal signal-transducing electrochemical dual-aptasensor for ultrasensitive parallel detection of GP73 and AFP...

Golgi protein 73 (GP73) has emerged as a promising serological biomarker for hepatocellular carcinoma (HCC), with its combination with alpha-fetoprotein (AFP) substantially improving diagnostic precision in early-stage HCC detection. To address this clinical need, we constructed an electrochemical dual-aptasensor for parallel quantification of GP73&AFP.

GP73 aptamer (GP73_Apt) functionalized with reduced graphene oxide-ferrocene-polyaniline (rGO-Fc-PANi) nanocomposites and AFP aptamer (AFP_Apt) conjugated with nitrogen-doped reduced graphene oxide-cuprous oxide (NrGO-Cu₂O) nanostructures were employed as the dual-signal probes. Upon GP73&AFP binding, the GP73_Apt-GP73 interaction induced double-helix dissociation of aptamer-complementary DNA (cDNA) complexes, stabilizing the Fc-mediated electrochemical structure and amplifying Fc redox current. Conversely, AFP_Apt-AFP interaction induced steric hindrance through aptamer-protein complex formation, synergistically combined with dielectric property changes to suppress Cu₂O redox signals. Under optimal conditions, the aptasensor demonstrated logarithmic linear ranges of 0.001-100.0 ng/mL for both biomarkers, achieving lower LOD of 0.48 pg/mL (GP73) and 1.77 pg/mL (AFP). Additionally, GP73&AFP were directly measured in human serum samples and demonstrated excellent reproducibility with RSD of 1.15-3.52 % for GP73 and 3.96-5.97 % for AFP.

This work presents a groundbreaking advancement in HCC diagnostics through the development of a dual-mode electrochemical aptasensor. The system's sensitive simultaneous detection capability, combined with operational stability in complex biological matrices, positions it as a transformative tool for early-stage HCC screening and management.