Label-free electrochemical aptasensor for sensitive and selective recognition of N6-Methyladenosine: Applicability in human urine.

Abstract

The specific and accurate detection of ultra-low levels of N6-methyladenosine (m6A) in biological samples holds great potential in the clinical diagnosis of various cancer types and neurodegenerative diseases. In this study, a cost-effective, label-free electrochemical aptasensor was developed by self-assembling a thiol-modified aptamer onto the surface of a gold electrode (AuE). Target recognition was measured by differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS) using [Fe(CN)6]3-/4- redox probe. The biosensor demonstrated superior selectivity for m6A determination in diluted artificial human urine samples, effectively distinguishing it from structurally related nucleosides and endogenous electroactive compounds. Under optimized conditions, the 6-mercapto-1-hexanol (MCH)-blocked electrode gave a limit of detection (LOD) in urine of 0.510 pM over a linear range of 3.2 pM - 10 nM by DPV and 0.700 pM over the same range by EIS. For the MCH/Apt/AuE platform, recovery values of 95.70% by DPV and 96.20% by EIS were obtained in 1:25 diluted artificial human urine, with corresponding relative standard deviations (RSD) of 2.65% and 2.50%. These results confirm the satisfactory accuracy and reproducibility of the developed aptasensor in this matrix. Consequently, the proposed aptasensor has the potential to serve as a non-invasive diagnostic platform that enhances patient comfort in the early diagnosis, treatment, and monitoring of cancer and neurodegenerative diseases, offering a strong alternative to invasive biopsy procedures.