Nanozyme-Enhanced Paper-Based Bipolar Electrode Biosensor for Dual-Mode Detection for M918T in Serum with Single-Base Specificity

The RET M918T mutation is a critical biomarker for early screening of medullary thyroid carcinoma (MTC). However, its ultralow abundance amid excess wild-type sequences poses challenges for accurate detection and limits the suitability of conventional methods for rapid point-of-care testing (POCT). Herein, a paper-based bipolar electrode-electrochemiluminescence (BPE-ECL) and colorimetry dual-modal biosensor integrating 2′-fluorinated nucleotide (2′-FN)-modified Y-shaped junctions with g-C 3 N 4 @FeMOFs nanozyme-mediated amplification was designed to enable sensitive and single-base resolution detection of M918T. The rigid 2′-FN Y-shaped junctions enhanced mismatch discrimination, while the g-C 3 N 4 @FeMOFs nanozyme exhibited strong peroxidase-like catalytic activity to simultaneously strengthen the ECL emission of Ru(bpy) 3 2+ /tripropylamine (Ru(bpy) 3 2+ /TPrA) and the colorimetric image of oxidizing 3,3′,5,5′-tetramethylbenzidine (TMB) to produce a blue chromogenic product in paper-based device readily quantified by smartphone imaging. Under optimal conditions, the proposed dual-mode strategy enabled detection of M918T with limits of 0.76 pM (ECL, S/N = 3) and 1.4 pM (colorimetry, S/N = 3) and demonstrated successful applicability in human serum samples. Both signals served as mutual self-validation for avoiding false-positives. This integrated, low-cost and portable sensing platform offers a rapid and reliable strategy for clinical biomarker detection, showing great potential for early diagnosis and on-site screening of MTC and related diseases.