A disposable electrochemiluminescence biosensor sensitized with multipedal DNA walker for in situ quenching detection of DNA methylation

Herein, a disposable electrochemiluminescence (ECL) biosensor sensitized with target-triggered multi-DNA release and multipedal DNA walker was designed for in situ quenching detection of methylated target DNA. In this work, zirconium(IV)-based metal organic frameworks encapsulated with Ru(bpy)32+ emitter (UiO-66-NH2@Ru) were attached onto indium tin oxide (ITO) electrode as sensing substrate. With the assistance of magnetic bead separation, one methylated DNA released three kinds of DNAs, achieving a one-to-multiple signal transduction and amplification. Then, these multi-DNA strands hybridized with swing arm immobilized on ITO surface as DNA walking strands. The further hybridization between multi-DNA strands and DNA tracks resulted in the formation of cleavage sites, which were digested by a restriction endonuclease, causing the cleavage of DNA tracks into two short fragments. Subsequently, the dissociated DNA walking strands explored adjacent DNA tracks, and the multipedal walking procedure was conducted. Relying on the in situ quenching of H2O2 for ECL emission of Ru(bpy)32+ system, this method achieved the quantification of methylated target DNA. Moreover, taking the advantages of inexpensiveness, high walking kinetics, great amplification efficiency, good reproducibility and practicability, this disposable biosensor held great potential for analyzing DNA methylation levels in resource-limited environments.