Self-Powered Microfluidic Device with Laser-Converted Graphene Electrodes for Immobilization-Free Electrochemical Detection of MPOX Virus DNA via Mismatch-Driven Nanocomplexes

Abstract

The re-emergence of monkeypox virus (MPXV) underscores the urgent need for rapid and decentralized diagnostic tools. Herein, we present an innovative, immobilization-free, and label-free paper-based electrochemical fast-flow microfluidic device (eFMD) integrated with near-field communication (NFC) technology for point-of-care detection of MPXV. The platform utilizes a C─C mismatch-mediated Ag⁺ intercalation mechanism, generating distinct current shifts detectable via differential pulse voltammetry (DPV). A microfluidic device with optimized serpentine mixing channels ensures efficient Ag⁺ intercalation without external power sources. The biosensor incorporates laser-converted graphene (LCG) electrodes, offering eco-friendly, scalable, and solvent-free fabrication with rapid prototyping and high design flexibility, ideal for disposable diagnostics. Key assay parameters including buffer type, mismatch probe concentration, AgNO<inf>3</inf> levels, and detection time were systematically optimized to enhance sensitivity. The biosensor demonstrated excellent analytical performance with a low detection limit down to 1.4 pм for MPXV DNA and significant discrimination against single-base mismatched sequences. Robust performance was further demonstrated in complex matrices, including spiked biological samples and clinically relevant lesion swab specimens, with detection sensitivity down to 1.3 copies mL⁻¹ and complete concordance with PCR classification. Importantly, the system allows for wireless, on-site readout via smartphones within 35 min, making it suitable for rapid MPXV screening.