DNAzyme-driven tripedal DNA walker mediated signal-on and label-free for electrochemical detection of α-synuclein oligomers

In the case of Parkinson’s disease (PD), the soluble α-synuclein (α-syn) oligomer is considered to be a reliable molecular biomarker. Developing easy-to-prepare, cheap, sensitive, and reliable techniques for the detection of α-synuclein oligomers is therefore a vital task. In this work, based on Mg2+-dependent DNAzyme (MNAzyme) -driven tripedal DNA walker tactics, we designed an innovative label-free and signal-on electrochemical aptasensor for α-syn oligomer quantification. α-syn oligomer-triggered tripedal DNA walkers outperformed common DNA walkers with the larger area of walking and accelerated walking kinetics, and facilitated greater amplification efficiency. With the assistance of MNAzyme, free-running tripedal DNA walkers could rapidly and continually scissor DNA hairpins (HPs), thereby unlocking numerous active G-quadruplex-forming sequences. Afterwards, G-quadruplex/hemin complexes could be formed by hemin further recognizing these G-quadruplexes and generated an enhanced current signal. Benefiting from tripedal DNA walker and MNAzyme dual signal amplification, we achieved a superior detection limit of 0.46 fM as well as a broad linear range of 1 fM to 10 pM. After being applied to α-syn oligomer detection in human serum samples, this novel electrochemical aptasensor demonstrated a high degree of sensitivity, anti-interference, and reproducibility, indicating its great potential for bioanalysis in complex biological environments.