Abstract

This chapter provides a comprehensive overview of how single-molecule imaging is achieved in live cells. The main focus is on fluorescent proteins, which are the most widely used fluorescent labels for live-cell imaging. The chromophore structures and the associated photochemical and photophysical properties of fluorescent proteins are discussed in detail, with a particular focus on how they influence single-molecule imaging in live cells. A few fluorescent proteins in the yellow-to-red spectral range, including newly discovered photoinducible ones, are selected for more detailed discussions due to their superior properties in single-molecule imaging. Special considerations for live-cell imaging and general instrumentations for single-molecule detection are also described. Finally, a few representative applications using single-molecule imaging in live cells are provided to illustrate how important biological knowledge can be obtained using this powerful technique.

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