Single-Molecule Imaging of Signaling Molecules in Living Cells

Single-molecule imaging is an ideal technology to study molecular mechanisms of biological reactions in vitro. Recently, we extended this technology to real-time observation of complexes of fluorescent dye-labeled epidermal growth factor (EGF) and its receptor (EGFR). Detection of single molecules was confirmed by single-step photobleaching. Using the single-molecule technique, the process of dimerization, a key step of EGFR signal transduction was observed. In addition to EGFR, single-molecules of fluorescently labeled nerve growth factor, GFP fusion forms of small G proteins and Raf1 kinase, and a fluorescent analogue of cAMP were observed in living cells.