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TripleFRET measurements in flow cytometry
Author(s) -
Fábián Ákos,
Horváth Gábor,
Vámosi György,
Vereb György,
Szöllősi János
Publication year - 2013
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.22267
Subject(s) - förster resonance energy transfer , fluorescence , energy transfer , biological system , biophysics , nanotechnology , chemistry , chemical physics , materials science , physics , optics , biology
A frequently used method for viewing protein interactions and conformation, Förster (fluorescence) resonance energy transfer (FRET), has traditionally been restricted to two fluorophores. Lately, several methods have been introduced to expand FRET methods to three species. We present a method that allows the determination of FRET efficiency in three‐dye systems on a flow cytometer. TripleFRET accurately reproduces energy transfer efficiency values measured in two‐dye systems, and it can indicate the presence of trimeric complexes, which is not possible with conventional FRET methods. We also discuss the interpretation of energy transfer values obtained with tripleFRET in relation to spatial distribution of labeled molecules, specifically addressing the limitations of using total energy transfer to determine molecular distance. © 2013 International Society for Advancement of Cytometry