Single-molecule tracking of carbohydrate-binding modules on cellulose using fluorescence microscopy

Single-molecule fluorescence detection is an invaluable technique for the study of molecular behavior in biological systems, both in vitro and in vivo. In this chapter, we focus on detailed protocols that utilize Total Internal Reflection Fluorescence Microscopy (TIRF-M) to visualize single molecules of carbohydrate-binding module (CBM) labeled with green fluorescent protein (GFP). The content describes step-by-step sample preparation and data acquisition, processing, and analysis. These methods can also be further used to study interactions between domains of cellulase molecules and between cellulases and cellulose. © 2012 Springer Science+Business Media, LLC.