Interrogating nanopores with light: optipore sensing for single molecule analyses

Nanopore sensors are at the heart of a technological shift, driven by their ability to detect biologically and medically relevant molecules one at a time. Electrical nanopore measurements have already transformed the field, yet they can struggle to distinguish analytes with similar ionic current signatures or to provide robust quantification in complex samples. Combining electrical and optical readouts addresses these limitations. Electro-optical nanopores enable multimodal single-molecule measurements, delivering reliable quantification alongside richer qualitative information.

This review focuses on synergetic integration of solid-state nanopores with advanced optical modalities, also referred to as optipore technology, as a unified framework that combines ionic current measurements with optical approaches such as fluorescence and scattering for synchronized, single-molecule analysis. Rather than treating electrical and optical methods independently, we emphasize their integration as a hybrid sensing paradigm that enables multidimensional characterization of molecular events. We examine key design considerations for nanopore membranes and microfluidic architectures, along with optical configurations ranging from confocal to wide-field microscopy, optimized to maximize photon-collection efficiency and enable high-throughput detection. Attention is given to strategies for background suppression, event validation, and molecular selectivity enabled by synchronized electro-optical readout. Recent developments in single-photon detectors and imaging technologies that improve sensitivity, throughput, and scalability are also highlighted. We analyze how these integrated platforms enable quantitative analysis of DNA, proteins, and protein-DNA complexes through chemo-selective labeling.

Electro-optical nanopores provide a versatile framework for multimodal single-molecule analysis, offer enhanced selectivity, improved confidence in event identification, and access to richer molecular information than purely electrical methods. By summarizing current methodologies, technological advances, and emerging applications, this review outlines the opportunities and remaining challenges for optipore technologies and their potential impact on next-generation single-molecule biosensing and diagnostic platforms.