Chemically Modified Hydrogel-Filled Nanopores: A Tunable Platform for Single-Molecule Sensing.

Area of Science:

• Biophysics
• Nanotechnology
• Molecular Diagnostics

Background:

• Label-free, single-molecule sensing is crucial for detecting low concentrations in complex biological samples.
• Solid-state nanopores are used for charged analyte detection, but fast nucleic acid transport limits sensing of small fragments.
• Existing methods struggle with the detection of small DNA fragments in physiological conditions.

Purpose of the Study:

• To develop a novel nanopore sensing platform for accurate detection of small nucleic acid fragments.
• To engineer hydrogel-filled nanopores (HFNs) capable of slowing down DNA transport for enhanced sensing.
• To demonstrate the potential of HFNs for molecular diagnostics, particularly for cell-free nucleic acids.

Main Methods:

• Fabrication of hydrogel-filled nanopores (HFNs) using quartz nanopipettes and poly(vinyl) alcohol hydrogels.
• Physical and chemical modification of hydrogels to control mesh size and composition.
• Characterization of DNA transport dynamics and sensing capabilities at physiological ionic strength.

Main Results:

• Hydrogel modification significantly slowed DNA transport by four orders of magnitude.
• Successfully detected DNA fragments as small as 100 base pairs (bp) using nanopores larger than 20 nm.
• Achieved sensing and size profiling of DNA fragments from 100 bp to >1 kbp under physiological conditions.

Conclusions:

• Hydrogel-filled nanopores (HFNs) offer a tunable and effective platform for label-free, single-molecule nucleic acid sensing.
• HFNs overcome limitations of standard nanopores in detecting small DNA fragments.
• This technology shows promise as a new generation of molecular diagnostic tools for applications like cancer diagnostics and prenatal testing.