title: Intensity range based quantitative FRET data analysis to localize protein molecules in live cell nuclei.
abstract: Förster (fluorescence) resonance energy transfer (FRET) is an ideal technique to estimate the distance between interacting protein molecules in live specimens using intensity-based microscopy. The spectral overlap of donor and acceptor- essential for FRET-also generates a contamination of the FRET signal. There are a number of algorithms available to remove this spectral bleedthrough (SBT) contamination and in this paper we compare two popular algorithms to estimate the SBT element and to calculate a more precise level of energy transfer efficiency, and with that a more accurate distance estimate.
authors.affiliation: W.M. Keck Center for Cellular Imaging, University of Virginia, Gilmer Hall, Charlottesville, Virginia 22904, USA.
authors.forename: Ye
authors.initials: Y
authors.lastname: Chen
authors.forename: Ammasi
authors.initials: A
authors.lastname: Periasamy
mesh_terms: D000465:Algorithms; D000818:Animals; D015153:Blotting, Western; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D016764:Cell Polarity; D003627:Data Interpretation, Statistical; D004285:Dogs; D018274:Electroporation; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007074:Immunoglobulin G / Q000378:metabolism; D008024:Ligands; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D008956:Models, Chemical; D010902:Pituitary Gland / Q000166:cytology; D010957:Plasmids; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D011961:Receptors, Fc / Q000378:metabolism; D014162:Transfection
chemical_list: D022763:CCAAT-Enhancer-Binding Protein-alpha; D005456:Fluorescent Dyes; C045281:IgA receptor; D007074:Immunoglobulin G; D008024:Ligands; D008164:Luminescent Proteins; D011506:Proteins; D011961:Receptors, Fc; D049452:Green Fluorescent Proteins