Microscopic analysis of fluorescence resonance energy transfer (FRET).

We describe here detailed practical procedures for implementing various approaches of fluorescence resonance energy transfer (FRET) in microscope-based measurements. A comprehensive theoretical formalism is developed and the different experimental procedures are outlined. A step-by-step protocol is provided for preparing the specimens for FRET measurements, data acquisition procedures, analysis, and quantification. Particular emphasis is given to exemplify the FRET applications in the study of protein-protein interactions.