An electroactive RNA-cleaving DNAzyme based electrochemical biosensor for enzyme-free detection of microRNA

Cancer remains among the leading causes of mortality globally, where the detection of biomarkers is essential for early diagnosis and accurate prognosis. Among them, microRNA-21 (miR-21) is a clinically significant oncogenic biomarker that is dysregulated across various cancers, highlighting the need for rapid, sensitive, and cost-effective detection strategies. In this study, we introduced a unique electroactive RNA-cleaving DNAzyme (e-RCD)-based electrochemical biosensor for enzyme-free detection of miR-21. An Mg²⁺ dependent 8-17 DNAzyme architecture was engineered to enable direct, target-specific recognition of miR-21. The target binding in the presence of Mg²⁺ activates the e-RCD probe's catalytic cleavage mechanism, releasing the methylene blue-labelled fragment from the electrode surface and generating a concentration-dependent electrochemical signal. Under optimised sensing conditions, the biosensor demonstrated a broader linear detection range from 500 aM to 50 nM with a low limit of detection of 107 aM. Furthermore, the biosensor demonstrated higher selectivity against closely related miRNAs, and the target recovery studies in serum samples showed satisfactory analytical accuracy. These results support the platform's potential applicability to complex biological samples, and it can be extended to detect other miRNA targets.