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    Precise DNA Concentration Measurements with Nanopores by Controlled Counting
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    • Martin Charron
      Martin Charron
      Department of Physics, University of Ottawa, 150 Louis-Pasteur, Ottawa, Ontario, Canada K1N 6N5
      More by Martin Charron
    • Kyle Briggs
      Kyle Briggs
      Department of Physics, University of Ottawa, 150 Louis-Pasteur, Ottawa, Ontario, Canada K1N 6N5
      More by Kyle Briggs
      Orcidhttp://orcid.org/0000-0002-0183-6585
    • Simon King
      Simon King
      Department of Physics, University of Ottawa, 150 Louis-Pasteur, Ottawa, Ontario, Canada K1N 6N5
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    • Matthew Waugh
      Matthew Waugh
      Department of Physics, University of Ottawa, 150 Louis-Pasteur, Ottawa, Ontario, Canada K1N 6N5
      More by Matthew Waugh
    • Vincent Tabard-Cossa*
      Vincent Tabard-Cossa
      Department of Physics, University of Ottawa, 150 Louis-Pasteur, Ottawa, Ontario, Canada K1N 6N5
      *E-mail: [email protected]
      More by Vincent Tabard-Cossa
      Orcidhttp://orcid.org/0000-0003-4375-717X
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    Analytical Chemistry

    Cite this: Anal. Chem. 2019, 91, 19, 12228–12237
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    https://doi.org/10.1021/acs.analchem.9b01900
    Published August 22, 2019
    Copyright © 2019 American Chemical Society
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    Abstract

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    Using a solid-state nanopore to measure the concentration of clinically relevant target analytes, such as proteins or specific DNA sequences, is a major goal of nanopore research. This is usually achieved by measuring the capture rate of the target analyte through the pore. However, progress is hindered by sources of systematic error that are beyond the level of control currently achievable with state-of-the-art nanofabrication techniques. In this work, we show that the capture rate process of solid-state nanopores is subject to significant sources of variability, both within individual nanopores over time and between different nanopores of nominally identical size, which are absent from theoretical electrophoretic capture models. We experimentally reveal that these fluctuations are inherent to the nanopore itself and make nanopore-based molecular concentration determination insufficiently precise to meet the standards of most applications. In this work, we present a simple method by which to reduce this variability, increasing the reliability, accuracy, and precision of single-molecule nanopore-based concentration measurements. We demonstrate controlled counting, a concentration measurement technique, which involves measuring the simultaneous capture rates of a mixture of both the target molecule and an internal calibrator of precisely known concentration. Using this method on linear DNA fragments, we show empirically that the requirements for precisely controlling the nanopore properties, including its size, height, geometry, and surface charge density or distribution, are removed while allowing for higher-precision measurements. The quantitative tools presented herein will greatly improve the utility of solid-state nanopores as sensors of target biomolecule concentration.

    Copyright © 2019 American Chemical Society

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    Supporting Information

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    The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem.9b01900.

    • S1, methods of capture rate analysis; S2, analyzing capture rates of time-varying capture processes; S3, controlled counting with time-varying capture rates; S4, electric field near the pore; S5, capture radius; S6, review of existing nanopore capture rate theory; and S7, scaling of translocation times with DNA length (PDF)

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    This article is cited by 40 publications.

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    Analytical Chemistry

    Cite this: Anal. Chem. 2019, 91, 19, 12228–12237
    Click to copy citationCitation copied!
    https://doi.org/10.1021/acs.analchem.9b01900
    Published August 22, 2019
    Copyright © 2019 American Chemical Society
    Request reuse permissions

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