Single-molecule microscopy is a cutting-edge technology that has significantly advanced our understanding of individual molecule behavior in living cells and complex biological systems. Its improved spatial and temporal resolution has unveiled molecular information previously unknown by traditional ensemble measurements. Notable applications include single-particle tracking of plasma membrane proteins and studying molecules' structural organization and location within highly organized structures such as neurons, cardiomyocytes, and T cells. 

This JOVE Methods Collection aims to provide a comprehensive and detailed set of visual protocols for single-molecule imaging of plasma membrane proteins developed by experts. These protocols are designed to study the dynamic patterns, stoichiometry, and complex protein-protein interactions of individual molecules, revealing their functional consequences. Methods of interest include labeling strategies, sample preparation, high temporal resolution imaging by optical setups, data analysis, and precise quantification. This collection is a valuable resource that empowers researchers to advance their methodologies and foster the development of innovative therapeutic approaches.