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Intensity Range Based Quantitative FRET Data Analysis to Localize Protein Molecules in Live Cell Nuclei |
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| Authors: | Ye Chen Ammasi Periasamy |
| Affiliation: | (1) W.M. Keck Center for Cellular Imaging, University of Virginia, Gilmer Hall, Charlottesville, Virginia 22904., U.S.;(2) Departments of Biology and Biomedical Engineering, University of Virginia, Gilmer Hall, Charlottesville, Virginia 22904., U.S. |
| Abstract: | F?rster (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. |
| Keywords: | FRET spectral bleedthrough (SBT) protein– protein interactions green fluorescent proteins (GFPs) |
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