Single Molecule Biosensing with Nanopores: Analyte quantification & Electric-field effects

Licentiatavhandling, 2026

Solid-state nanopores enable label-free single-molecule analysis by converting the passage of analytes through the pore into electrical signals. This thesis examines quantitative analyte sensing with nanopores while addressing a central challenge: the exceptionally strong electric field in thin solid-state pores. The first part of the work demonstrates that analyte concentration can be extracted directly from the voltage dependence of translocation event frequency. For double-stranded DNA in sufficiently large pores, the event frequency increases linearly with voltage. By extending measurements to high bias and introducing robust signal-processing routines for unstable baselines, the study establishes a calibration-free approach for concentration determination from nanopore data. The second part demonstrates that the electric field inside a solid-state nanopore is not merely a passive readout mechanism, but rather a perturbative environment that actively modifies biomolecular interactions. Using the biotin-avidin interaction as a model system,
applied bias shortens the effective lifetime of this strongly bound complex by several orders of magnitude. This finding has critical implications for affinity-based nanopore sensing, because binding kinetics measured in the pore can deviate substantially from field-free conditions. The thesis further includes studies on thermoresponsive PNIPAM-grafted nanostructures for trapping enzymes and controlling ionic transport, which demonstrates the viability of developing integrated platforms that simultaneously provide electrical detection, optical readout, and dynamically gated access to confined biomolecules, enabling real-time monitoring of individual enzyme kinetics in controlled nanoscale environments. Overall, these studies establish solid-state nanopores as versatile platforms for quantitative single-molecule biosensing, while revealing critical field-induced effects that must be
considered in future bioanalytical applications.