Nonfluorescent Single‐Molecule Tracking: Revealing Nanoscale Kinetics of Individual Molecular Binding Events

Authors:
Publisher website
Google Scholar
Abstract
Single‐molecule tracking (SMT) has revolutionized our understanding of molecular interactionsby revealing nanoscale behaviors and dynamic processes obscured in ensemble‐averaged measurements. While fluorescent SMT has been instrumental, its utility is constrained by fundamental limitations such as photobleaching and finite photon budgets, which restrict temporal resolution and observation periods. Furthermore, the fluorescent labels themselves can perturb native molecular interactions. The recent emergence of nonfluorescent SMT techniques offers a powerful alternative, enabling long‐term or label‐free observation with high temporal resolution for a more direct view into nanoscale binding kinetics. This review comprehensively summarizes recent advances in nonfluorescent SMT methodologies. We describe the typical optical devices, detail the tracking algorithms, and showcase their applications in elucidating previously inaccessible aspects of molecular binding. We further critically discuss persistent technical hurdles and outline promising future directions, underscoring the interdisciplinary synergy between advanced optical techniques and data science required to unlock the full potential of these next‐generation tools.
Funding Information
  • National Natural Science Foundation of China (22404164)

This publication has 83 references indexed in Scilit: