Dependent signal quenching and enhancing triggered by bipedal DNA walker for ultrasensitive photoelectrochemical biosensor

Herein, by utilizing bipedal DNA walker as booster to adjust the distance of quencher ferrocene (Fc) and sensitizer methylene blue (MB) to photoactive material perylene-3,4,9,10-tetracarboxylic acid (PTCA), a novel “on-off-super on” photoelectrochemical (PEC) biosensor was proposed for ultrasensitive detection of thrombin (TB). Firstly, the PTCA matrix on electrode could provide a high initial PEC signal assisted by depAu. Upon the Fc labeled on hairpin DNA 1 (H1-Fc) proximate to PTCA, the PEC signal could obviously decrease to reduce the background signal. Interestingly, the target TB-related bipedal DNA walker implemented the opening of H1-Fc for the departure of Fc toward PTCA, which achieved the recovery of PEC signal and exposed the prelocked toehold domain for the hybridization with hairpin DNA 2 labeled with MB (H2-MB), thereby making the MB approach to PTCA for achieving the “super on” signal. As a result, this proposed strategy showed a wide linear range from 0.5 fM to 100 nM with a low detection limit down to 0.17 fM for TB detection, providing an efficient and available avenue for sensitive detection of biomolecules in bioanalysis and disease diagnosis. •The MB and Fc could assemble on signal-DNA strands for implementing the signal quenching and enhancing via DNA regulator.•Both the departure of Fc to PTCA and proximity of MB to PTCA could enhance the second “on” signal to achieve “super on” state.•A target-recycling amplification strategy by releasing the bipedal DNA walker was applied to achieve the higher detection sensitivity of TB.