Direct Visualization and Quantification of the Actin Nucleation andElongation Events in vitro by TIRF Microscopy

Total internal reflection fluorescence (TIRF) microscopy is a powerful tool for visualizing the dynamics of actin filaments at single-filament resolution in vitro. Thanks to the development of various fluorescent probes, we can easily monitor all kinds of events associated with actin dynamics, including nucleation, elongation, bundling, fragmentation and monomer dissociation. Here we present a detailed protocol regarding the visualization and quantification of actin nucleation and filament elongation events by TIRF microscopy in vitro, which is based on the methods previously reported ( Liu et al., 2015 ; Yang et al., 2011 ).