Quantitative intensity-based FRET approaches--a comparative snapshot.

Author: Zeug A, Woehler A, Neher E, Ponimaskin EG.
Cellular Neurophysiology, Center of Physiology, Hannover Medical School, Hannover, Germany.
Conference/Journal: Biophys J.
Date published: 2012 Nov 7
Other: Volume ID: 103 , Issue ID: 9 , Pages: 1821-7 , Special Notes: doi: 10.1016/j.bpj.2012.09.031 , Word Count: 167

Förster resonance energy transfer (FRET) has become an important tool for analyzing different aspects of interactions among biological macromolecules in their native environments. FRET analysis has also been successfully applied to study the spatiotemporal regulation of various cellular processes using genetically encoded FRET-based biosensors. A variety of procedures have been described for measuring FRET efficiency or the relative abundance of donor-acceptor complexes, based on analysis of the donor fluorescence lifetime or the spectrally resolved fluorescence intensity. The latter methods are preferable if one wants to not only quantify the apparent FRET efficiencies but also calculate donor-acceptor stoichiometry and observe fast dynamic changes in the interactions among donor and acceptor molecules in live cells. This review focuses on a comparison of the available intensity-based approaches used to measure FRET. We discuss their strengths and weaknesses in terms of FRET quantification, and provide several examples of biological applications.
Copyright © 2012 Biophysical Society. Published by Elsevier Inc. All rights reserved.
PMID: 23199910 [PubMed - indexed for MEDLINE] PMCID: PMC3491707 [Available on 2013/11/7]