Analogous modified DNA probe and immune competition method-based electrochemical biosensor for RNA modification

N6-methyladenosine (m6A), one of the most abundant RNA methylation which is ubiquitous in eukaryotic RNA, plays vital roles in many biological progresses. Therefore, the rapid and accurate quantitative detection of m6A is particularly important for its functional research. Herein, a label-free and highly selective electrochemical immunosensor was developed for the detection of m6A. The method is established on that the anti-m6A-Ab can recognize both m6A-RNA and m6A-DNA. An analogous modified DNA probe (L1) serves as a signal molecule, by competing with m6A-RNA for binding to Abs to broaden the linear range. The detection of m6A-RNA by this method is unaffected by the lengths and base sequences of RNA. Under optimal conditions, the proposed immunosensor presented a wide linear range from 0.05 to 200 nM with a detection limit as low as 0.016 nM (S/N = 3). The specificity and reproducibility of the method are satisfactory. Furthermore, the developed immunosensor was validated for m6A determination in human cell lines. Thus, the immunosensor provides a promising platform for m6A-RNA detection with simplicity, high specificity and sensitivity.