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Determination of Small-Sized α-Synuclein Oligomers Using Solid-State Nanopore Assisted with Circular Single-Stranded

Siying Heng1,2, Yao Tong1, Xiaoyan Xu1

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This study introduces a novel nanopore sensing method using a DNA frame to quantify alpha-synuclein (αS) oligomers, crucial in Parkinson's disease (PD) pathogenesis. The technique enables single-molecule analysis of αS oligomer species.

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Area of Science:

  • Biophysics
  • Nanotechnology
  • Neuroscience

Background:

  • Misfolded alpha-synuclein (αS) oligomers are key drivers of Parkinson's disease (PD) neurodegeneration.
  • Small αS oligomers exhibit higher cytotoxicity than mature aggregates.
  • Solid-state nanopores offer high-resolution single-molecule sensing but struggle with heterogeneous biomolecules like αS oligomers.

Purpose of the Study:

  • To develop a quantitative method for determining αS oligomers of varying sizes using nanopore sensing.
  • To enhance αS oligomer identification and classification using a circular single-stranded DNA (CssDNA) frame.
  • To assess the selectivity of the CssDNA frame for αS oligomers against other proteins.

Main Methods:

  • Utilized 10 nm silicon nitride (SiNx) nanopores for resistive pulse sensing.
  • Employed a CssDNA frame functionalized with an aptamer to capture αS oligomers.
  • Analyzed translocation events of CssDNA-αS complexes to identify and classify oligomer species.

Main Results:

  • Successfully identified and classified αS oligomers from dimers to heptamers based on translocation event analysis.
  • Detected primarily dimeric αS oligomers after 0.5 h incubation with the CssDNA frame.
  • Demonstrated high selectivity of the CssDNA frame by distinguishing αS oligomers from β-lactoglobulin, Aβ1-42, and tau proteins.

Conclusions:

  • The developed nanopore sensing approach, assisted by a CssDNA frame, enables quantitative analysis of αS oligomers at the single-molecule level.
  • This method provides a promising alternative for studying αS oligomer heterogeneity relevant to Parkinson's disease.
  • The specificity of the CssDNA frame highlights its potential for targeted biomolecular detection.