Highly Sensitive m6A Electrochemical Biosensor Based on Laser-Induced Graphene Electrodes Functionalized with DNA Tetrahedron/Gold-antibody Nanoparticle

Background N6-methyladenosine (m6A), the most abundant internal modification of eukaryotic mRNA, post-transcriptionally regulates gene expression, and its dysregulation is intimately linked to tumorigenesis and multiple human diseases. However, existed detection methods including high-performance liquid chromatography (HPLC) suffer from obvious drawbacks such as site specificity constraints, high cost, reliance on sophisticated instruments and tedious operation, making them unsuitable for rapid on-site detection. Therefore, the development of a novel, rapid, on-site and cost-effective detection strategy for m6A is of great significance and urgent demand. Results This study developed an electrochemical biosensor based on DNA tetrahedrons (TDN) and gold-antibody nanoparticle (AuNPs-ab) for highly sensitive detection of m6A-RNA modifications. After enhancing conductivity through gold nanoparticle modification, TDN was stably immobilized on the electrode surface via Au-S bonds. The rigid structure of TDN was utilized to optimize surface flatness and spatial architecture, enabling precise anchoring of the antibody probe. Target m6A-RNA captured by antibodies undergoes biotin-streptavidin-mediated horseradish peroxidase (HRP) conjugation, catalyzing a hydrogen peroxide–hydroquinone (H 2 O 2 –HQ) redox cycle to generate amplified current signals. The sensor exhibits excellent linear response within the 0.001–100 nM range, and the detection limit is 0.105 pM, demonstrating high specificity and stability (signal retention rate >88% after 15 days). Furthermore, this method has been successfully applied to detect target samples spiked into human serum and total RNA extracted from hepatocellular carcinoma cells. Significance This sensing platform combines the compact size and scalable batch-production advantages of LIG electrodes, offering a novel approach for precise detection of tumor-associated m6A modifications and rational design of disposable, portable point-of-care testing (POCT) devices. It demonstrates significant potential for rapid screening of key epigenetic biomarkers and real-time monitoring across diverse clinical and biological samples.