Measurement of Fluorescence Resonance Energy Transfer in the Optical Microscope
Fluorescence resonance energy transfer (FRET) can be used as a spectroscopic ruler to study and quantify the interactions of cellular components with each other, as well as the conformational changes within individual molecules at the molecular level (Herman, 1998). FRET is a process by which a fluorophore (donor) in an excited state may transfer its excitation energy to a neighboring chromophore (acceptor) nonradiatively through dipole—dipole interactions. This energy transfer manifests itself as both quenching of donor fluorescence intensity and lifetime (in the presence of acceptor) as well as an increase in the emission of acceptor fluorescence (sensitized emission). Because FRET decreases in proportion to the inverse sixth power of the distance between the donor and acceptor, this phenomenon is effective at measuring separation of the donor- and acceptor-labeled molecules when they are within 10–100 Å of each other.
KeywordsFluorescence Resonance Energy Transfer Forster Resonance Energy Transfer Fluorescence Lifetime Imaging Microscopy Donor Emission Bioluminescence Resonance Energy Transfer
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