Nanometer-scale pores have demonstrated potential for the electrical detection, quantification, and characterization of molecules for biomedical applications and the chemical analysis of polymers. Despite extensive research in the nanopore sensing field, there is a paucity of theoretical models that incorporate the interactions between chemicals (i.e., solute, solvent, analyte, and nanopore). Here, we develop a model that simultaneously describes both the current blockade depth and residence times caused by individual poly(ethylene glycol) (PEG) molecules in a single α-hemolysin ion channel. Modeling polymer-cation binding leads to a description of two significant effects: a reduction in the mobile cation concentration inside the pore and an increase in the affinity between the polymer and the pore. The model was used to estimate the free energy of formation for K þ -PEG inside the nanopore (≈ − 49.7 meV) and the free energy of PEG partitioning into the nanopore (≈0.76 meV per ethylene glycol monomer). The results suggest that rational, physical models for the analysis of analyte-nanopore interactions will develop the full potential of nanopore-based sensing for chemical and biological applications.alpha-hemolysin | nanopore-based sensing | polymer confinement | polymer analysis P olymers play a fundamental role in life (1) and are central to many emerging technologies (2). Many of these applications require a detailed understanding of the structure, morphology, and chemical interactions of polymers under confinement in either 2-dimensional films (3) or narrow tubes (4). The ability to isolate and study single molecules has shown promise in overcoming the limitations of measurements with ensemble averages and permits probing the inter-and intra-molecular forces, structural changes, and dynamics of polymers (for a detailed review of single-molecule polymer analysis see refs. 5 and 6).Molecules partition into a nanopore and alter the flow of ions resulting in distinct current blockades that can be used to detect, characterize, and quantify a wide range of polymer types (7). These include single-stranded RNA and DNA (8-10), proteins (11-14), biowarfare agents (15), therapeutic agents against anthrax toxins (15, 16), and chemically synthesized molecules (14,(17)(18)(19)(20). More recently, single nanopores were used to determine the size distribution of polymers in a manner akin to mass spectrometry (19).To fully realize the potential of nanopore-based sensors, it is important to develop a detailed understanding of the physical and chemical interactions of polymers with the nanopore, solvent, electrolyte, and other components. In this work, the interaction between poly(ethylene glycol) (PEG) and the α-hemolysin (αHL) channel in a high ionic strength electrolyte was used to develop a unique model of polymers confined within single nanopores.Previous attempts to describe the magnitude of PEG-induced nanopore current blockades focused on volume exclusion (21) and/or microviscosity (22), which required adjustable ad hoc...

There are thousands of different nanometer-scale pores in biology, many of which act as sensors for specific chemical agents. Recent work suggests that protein and solid-state nanopores have many potential uses in a wide variety of analytical applications. In this review we survey this field of research and discuss the prospects for advances that could be made in the near future.

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We describe a novel single molecule nanopore-based sequencing by synthesis (Nano-SBS) strategy that can accurately distinguish four bases by detecting 4 different sized tags released from 5′-phosphate-modified nucleotides. The basic principle is as follows. As each nucleotide is incorporated into the growing DNA strand during the polymerase reaction, its tag is released and enters a nanopore in release order. This produces a unique ionic current blockade signature due to the tag's distinct chemical structure, thereby determining DNA sequence electronically at single molecule level with single base resolution. As proof of principle, we attached four different length PEG-coumarin tags to the terminal phosphate of 2′-deoxyguanosine-5′-tetraphosphate. We demonstrate efficient, accurate incorporation of the nucleotide analogs during the polymerase reaction, and excellent discrimination among the four tags based on nanopore ionic currents. This approach coupled with polymerase attached to the nanopores in an array format should yield a single-molecule electronic Nano-SBS platform.