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Diffusivity Measurement by Single-Molecule Recycling in a Capillary Microchannel.

Bo Wang1,2, Lloyd M Davis1,2

  • 1Center for Laser Application, University of Tennessee Space Institute, 411 B H Goethert Pkwy, Tullahoma, TN 37388, USA.

Micromachines
|August 6, 2021
PubMed
Summary

This study introduces a simpler, more cost-effective capillary microchannel method for single-molecule recycling. The new technique extends observation times by over 200 recycles, aiding high-throughput screening.

Keywords:
capillarymaximum likelihoodmicrofluidic devicesingle-molecule recycling

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Area of Science:

  • Biophysics
  • Analytical Chemistry
  • Microfluidics

Background:

  • Microfluidic devices are used in various analyses like DNA sequencing and ligand-binding.
  • Existing methods for single-molecule observation in microchannels face challenges such as surface sticking, high viscosity, and costly fabrication.

Purpose of the Study:

  • To develop a simplified and cost-effective single-molecule recycling technique using a capillary microchannel.
  • To overcome limitations of nanochannel-based methods, including complex fabrication and molecule localization difficulties.

Main Methods:

  • A 1-inch piece of a commercial fused-silica capillary (2-micron inner diameter) was modified with o-rings to serve as reservoirs and an observation window.
  • The capillary's inner surface was chemically treated to minimize non-specific sticking and enhance capillary effects.
  • A piezo stage translated the capillary for molecule recycling, with a confocal microscope for fluorescence collection and an FPGA-based system for real-time control and feedback.

Main Results:

  • Achieved over 200 recycles of a single molecule, significantly extending observation time.
  • Developed a maximum-likelihood estimation for molecule diffusivity, yielding results comparable to previous reports.
  • The capillary microchannel method demonstrated reduced non-specific sticking and improved molecule localization compared to nanochannels.

Conclusions:

  • The proposed capillary microchannel method offers a simplified, less expensive, and easier-to-prepare alternative for single-molecule recycling.
  • This technique effectively overcomes challenges associated with nanochannel-based approaches.
  • The method shows promise for applications in ligand-binding studies and high-throughput screening.