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- Title
Prolonged irradiation of enhanced cyan fluorescent protein or Cerulean can invalidate Fo¨rster resonance energy transfer measurements.
- Authors
Birgit Hoffmann; Thomas Zimmer; Nikolaj Klo¨cker; Laimonas Kelbauskas; Karsten Ko¨nig; Klaus Benndorf; Christoph Biskup
- Abstract
Since its discovery, green fluorescent protein (GFP) and its variants have proven to be a good and convenient fluorescent label for proteins: GFP and other visible fluorescent proteins (VFPs) can be fused selectively to the protein of interest by simple cloning techniques and develop fluorescence without additional cofactors. Among the steadily growing collection of VFPs, several pairs can be chosen that can serve as donor and acceptor fluorophores in Fo¨rster resonance energy transfer (FRET) experiments. Among them, the cyan fluorescent proteins (ECFP/Cerulean) and the enhanced yellow fluorescent protein (EYFP) are most commonly used. We show that ECFP and Cerulean have some disadvantages despite their common use: Upon irradiation with light intensities that are commonly used for intensity- and lifetime-based FRET measurements, both the fluorescence intensity and the fluorescence lifetime of ECFP and Cerulean decrease. This can hamper both intensity- and lifetime-based FRET measurements and emphasizes the need for control measurements to exclude these artifacts.
- Subjects
PROTEINS; ENERGY transfer; IRRADIATION; FLUORESCENCE
- Publication
Journal of Biomedical Optics, 2008, Vol 13, Issue 3, p31205
- ISSN
1083-3668
- Publication type
Article
- DOI
10.1117/1.2937829