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Updated: May 16, 2025

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Solid-State Nanopore Sizing for cfDNA Sample Quality Control in Point-of-Need Sequencing.

Muhammad Asad Ullah Khalid1, Md Ahasan Ahamed1,2, Anthony J Politza3

  • 1Department of Intelligent System Engineering, Indiana University, Bloomington, Indiana 47408, United States.

Biorxiv : the Preprint Server for Biology
|April 1, 2025
PubMed
Summary

Portable nanopore sequencing needs on-site DNA quality control. This study introduces a solid-state nanopore device for label-free quantification of cell-free DNA (cfDNA), enabling accurate measurements at the point of need.

Keywords:
cell-free DNAnanoporeplasmapoint-of-needquality controlsequencing

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Area of Science:

  • Biotechnology
  • Genomics
  • Analytical Chemistry

Background:

  • DNA sequencing, including portable nanopore sequencing, is vital for disease diagnosis.
  • Current DNA sample quality control (QC) is lab-bound, limiting the utility of portable sequencers.
  • On-site QC is essential for personalized healthcare applications of portable sequencing.

Purpose of the Study:

  • To develop and validate a solid-state nanopore device for label-free quantification and qualification of cell-free DNA (cfDNA).
  • To enable accurate cfDNA assessment at the point of need, complementing portable sequencing technologies.

Main Methods:

  • Utilized a solid-state nanopore device for label-free cfDNA analysis.
  • Employed a 1 kbp double-stranded DNA internal marker for controlled quantification.
  • Investigated nanopore diameters (6-19 nm) and acquisition times for measurement consistency.
  • Compared nanopore assay results with capillary electrophoresis (CE) for plasma cfDNA samples.

Main Results:

  • Demonstrated controlled quantification of cfDNA using a DNA internal marker.
  • Achieved consistent measurements with nanopores (6-19 nm) with <15% CV.
  • Reduced measurement uncertainty to <10% CV by analyzing data from multiple nanopores over longer times.
  • Showed highly correlated results between the nanopore QC assay and CE for plasma cfDNA.

Conclusions:

  • The developed solid-state nanopore device offers a robust method for label-free cfDNA quantification and qualification.
  • This nanopore QC assay is suitable for point-of-need applications, enhancing the utility of portable DNA sequencing.
  • The technology shows significant potential for effective cfDNA assessment in personalized healthcare settings.