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15 February 2008 Multidimensional fluorescence lifetime measurements
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Abstract
In this study, we present two different approaches that can be used for multi-wavelength fluorescence lifetime measurements in the time domain. One technique is based on a streak-camera system, the other technique is based on the time-correlated-single-photon-counting (TCSPC) approach. The setup consists of a confocal laser-scanning microscope and a Titanium:Sapphire-laser that is used for pulsed one- and two-photon excitation. Fluorescence light emitted by the sample is fed back through the scan head and guided to one of the confocal channels, where it is coupled into an optical fiber and directed to a polychromator. The polychromator disperses the emitted light according to its wavelength and focuses the resulting spectrum on the entrance slit of a streak camera or a 16 channel PMT array, which is connected to a TCSPC imaging module. With these techniques it is possible to acquire fluorescence decays in several wavelength regions simultaneously. We applied these methods to Förster resonance energy transfer (FRET) measurements and discuss the advantages and pitfalls of fluorescence lifetime measurements.
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Christoph Biskup, Birgit Hoffmann, Laimonas Kelbauskas, Thomas Zimmer, Nikolaj Klöcker, Wolfgang Becker, and Klaus Benndorf "Multidimensional fluorescence lifetime measurements", Proc. SPIE 6860, Multiphoton Microscopy in the Biomedical Sciences VIII, 68601I (15 February 2008); https://doi.org/10.1117/12.762834
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