10 February 2007 Multi-dimensional fluorescence lifetime measurements
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In this study, we present two different approaches that allow multi-wavelength fluorescence lifetime measurements in the time domain. One technique is based on a streak camera system, the other technique is based on a time-correlated singlephoton- counting (TCSPC) approach. The setup consists of a confocal laser-scanning microscope (LSM 510, Zeiss) and a Titanium:Sapphire-laser (Mira 900D, Coherent) that is used for pulsed one- and two-photon excitation. Fluorescence light emitted by the sample is dispersed by a polychromator (250is, Chromex) and recorded by a streak camera (C5680 with M5677 sweep unit, Hamamatsu Photonics) or a 16 channel TCSPC detector head (PML-16, Becker & Hickl) connected to a TCSPC imaging module (SPC-730/SPC-830, Becker & Hickl). 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 over fluorescence techniques that are already well established in the field of confocal microscopy, such as spectrally resolved intensity measurements or single-wavelength fluorescence lifetime measurements.
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Christoph Biskup, Christoph Biskup, Birgit Hoffmann, Birgit Hoffmann, Laimonas Kelbauskas, Laimonas Kelbauskas, Thomas Zimmer, Thomas Zimmer, Sascha Dietrich, Sascha Dietrich, Wolfgang Becker, Wolfgang Becker, Axel Bergmann, Axel Bergmann, Nikolaj Klöcker, Nikolaj Klöcker, Klaus Benndorf, Klaus Benndorf, "Multi-dimensional fluorescence lifetime measurements", Proc. SPIE 6442, Multiphoton Microscopy in the Biomedical Sciences VII, 64420V (10 February 2007); doi: 10.1117/12.699870; https://doi.org/10.1117/12.699870


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