Practical FRET measurement - R.Truant

This section will involve the theory and practical application of the use of Forster Resonant Energy Transfer, or FRET, to detect direct molecular interactions in vivo, or in use as biosensors. We will discuss the theory behind FRET, and the detection of FRET by multiple methods including stimulated emission, acceptor photobleaching and FRET detection by fluorescence lifetime imaging, or FLIM. Advantages and disadvantages of this method will be discussed, as well as some well characterized uses of FRET as biosensors for calcium measurement and protein conformation, and the use of FRET for protein-nuclei acid or protein-lipid interactions in vivo. Some typical fluorescent protein donor-acceptor pairs will be discussed, as well as typical fluorescent protein-fluorescent dye donor-acceptor pairs. Focus will be on two molecule FRET detection.