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Journal of Biomedical Optics

Spectrally resolved fluorescence lifetime imaging microscopy: Förster resonant energy transfer global analysis with a one- and two-exponential donor model
Author(s): Daniela Strat; Frank Dolp; Angelika C. Rueck; Bjorn von Einem; Cornelia Steinmetz; Christine A. F. von Arnim
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Paper Abstract

In many fields of life science, visualization of spatial proximity, as an indicator of protein interactions in living cells, is of outstanding interest. A method to accomplish this is the measurement of Förster resonant energy transfer (FRET) by means of spectrally resolved fluorescence lifetime imaging microscopy. The fluorescence lifetime is calculated using a multiple-wavelength fitting routine. The donor profile is assumed first to have a monoexponential time-dependent behavior, and the acceptor decay profile is solved analytically. Later, the donor profile is assumed to have a two-exponential time-dependent behavior and the acceptor decay profile is derived analytically. We develop and apply a multispectral fluorescence lifetime imaging microscopy analysis system for FRET global analysis with time-resolved and spectrally resolved techniques, including information from donor and acceptor channels in contrast to using just a limited spectral data set from one detector only and a model accounting only for the donor signal. This analysis is used to demonstrate close vicinity of β-secretase (BACE) and GGA1, two proteins involved in Alzheimer's disease pathology. We attempt to verify if an improvement in calculating the donor lifetimes could be achieved when time-resolved and spectrally resolved techniques are simultaneously incorporated.

Paper Details

Date Published: 1 February 2011
PDF: 19 pages
J. Biomed. Opt. 16(2) 026002 doi: 10.1117/1.3533318
Published in: Journal of Biomedical Optics Volume 16, Issue 2
Show Author Affiliations
Daniela Strat, Univ. Ulm (Germany)
Frank Dolp, Univ. Ulm (Germany)
Angelika C. Rueck, Univ. Ulm (Germany)
Bjorn von Einem, Univ. Ulm (Germany)
Cornelia Steinmetz, Univ. Ulm (Germany)
Christine A. F. von Arnim, Univ. Ulm (Germany)

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