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Spatially-Resolved Chemical Detection with a Nanoneedle-Probe-Supported Biological Nanopore Kan Shoji, Ryuji Kawano, and Ryan J. White ACS Nano, Just Accepted Manuscript • DOI: 10.1021/acsnano.8b09667 • Publication Date (Web): 06 Feb 2019 Downloaded from http://pubs.acs.org on February 8, 2019
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ACS Nano
Spatially-Resolved Chemical Detection with a Nanoneedle-Probe-Supported Biological Nanopore Kan Shoji , Ryuji Kawano and Ryan J. White a, b
a
b,c*
a Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16 Naka-cho, Koganei-shi, Tokyo 184-8588, Japan b Department of Chemistry, University of Cincinnati, Cincinnati, Ohio 45221-0172, United States c Department of Electrical Engineering and Computer Science, University of Cincinnati, Cincinnati, Ohio 45221-0172, United States *Corresponding author: e-mail
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ABSTRACT
In this manuscript, we describe the quantitative characterization of a gold nanoneedle ion channel probe and demonstrate the utility of this probe for spatially-resolved detection of a small molecule using ion channel activity. Our report builds off of recent reports of Ide and coworkers, who reported on the use of and etched gold wire modified with a poly(ethylene) glycol (PEG) monolayer as a support for a lipid bilayer and subsequent single ion channel recordings. While this nanoneedle electrode approach has been reported previously, in our report we investigate the effects of several operational parameters on the performance of the ion channel measurement and electrochemical phenomenon that occur in the nano-confined space between the supported bilayer and the gold electrode. More specifically, we address the effects the length of the supporting monolayer and the composition of the electrolyte baths on channel current measurements, and provide a quantitative description of what carries current at the working electrode (double layer charging). In addition, we demonstrate the ability to control the direction of protein insertion (tipside vs. bath-side) with freely diffusing protein which has not been previously reported, with the former method (tip-side) enabling single molecule detection of β-cyclodextrin (βCD) using a reconstituted α-hemolysin channel. Finally, anticipating future use of a nanoneedle-based biological nanopore probe in a scanned-probe microscopy, we demonstrate the ability to quantify and spatially resolve the concentration of βCD molecules in a microfluidic channel. We believe, in the long term, the described nanoneedle-based biological nanopore probe can be employed in, for example, scanning ion conductance microscopy using ion channels.
KEYWORDS nanopore, resistive pulse, a-hemolysin, imaging, nanoneedle, lipid bilayer membrane
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ACS Nano
Resistive-pulse nanopore sensors that measure blockage currents caused by single molecules binding to, or translocating through, a nanopore provide single molecule sensing with high sensitivity, temporal resolution, signal-to-noise ratio, and molecular selectively.1-6 These sensors have been applied as DNA sequencers7-9 and sensors for biomolecules including miRNAs5,10-13 and proteins.14-17 Solid-state nanopores18,19 typically fabricated by exposing thin membranes such as silicon nitride and carbon to a high-energy electron beam or biological nanopores20 which comprise pore-forming proteins reconstituted into a lipid bilayer membrane are key examples of the types of nanopores utilized in this sensing strategy. While both show promise for sensing applications, biological nanopores (i.e., protein channels) provide reproducible pore sizes dictated by the protein structure and have thus found widespread as resistive pulse detectors. The employ biological nanopores for sensing purposes, a stable lipid bilayer membrane is required. While the painting method21 and the Montal-Mueller method22, are generally used, they result in the formation of a membrane spanning across a pore in a hydrophobic substrate these methods are time consuming, and the resulting membrane is unstable to vibrational and high voltage (>~250 mV) perturbations. As such, numerous developments of methods for the formation of lipid bilayer membranes have been reported for channel measurements. For example, a droplet contact method (DCM)23-25 and droplet interface bilayers (DIB)26,27 that form bilayer membranes by contacting droplets that are surrounded by a lipid monolayer have been reported as the simple and easy method to prepare stable lipid bilayer membrane. Glass micro- and nanopore membranes28-30 also provide a stable platform to support lipid bilayers across small (
