Analysis and Applications of Single-Molecule Fluorescence in Live Cell Membranes

Abstract

In the past decades, single-molecule fluorescence (SMF) techniques have been booming as they provide researchers with valuable molecular information that are unobtainable by using ensemble techniques. Applications of SMF techniques to live cell membranes have revolutionized our views on many cellular processes. In this chapter, we describe the basics of SMF techniques such as SMF imaging, single-molecule fluorescence resonance energy transfer (sm-FRET), and fluorescence correlation spectroscopy (FCS) and then discuss their contributions to the understanding of live cell membranes. We finally provide the practical protocols of applying SMF techniques to live cell membranes, including the choice of fluorescent probes, labeling strategies, cell sample preparation, instrumentation setup, and data analysis.