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Photoconversion of YFP into a CFP-like species during acceptor photobleaching FRET experiments

Nature Methods volume 2, page 801 (2005)Cite this article

To the editor:

Fluorescence resonance energy transfer (FRET) is a useful technique to detect protein-protein interactions in cells1,2. Acceptor photobleaching approaches with CFP- and YFP-tagged proteins are simple and prevent problems associated with variable expression levels: when FRET occurs, the fluorescence of the CFP donor increases after bleaching the YFP acceptor chromophore, and this is recognized as a signature for FRET2.

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Figure 1: YFP can yield false FRET signals with the acceptor photobleaching method.

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Authors and Affiliations

  1. IGMM-CNRS UMR 5535, 1919, route de Mende, Montpellier, 34293, Cedex 5, France

    Guillaume Valentin, Céline Verheggen, Henry Neel & Edouard Bertrand

  2. IJM-CNRS Université Paris VI/VII, 2, pl. Jussieu, Paris, 75251, Cedex 5, France

    Tristan Piolot & Maïté Coppey-Moisan

Authors
  1. Guillaume Valentin
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  2. Céline Verheggen
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  3. Tristan Piolot
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  4. Henry Neel
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  5. Maïté Coppey-Moisan
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  6. Edouard Bertrand
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Supplementary information

Supplementary Fig. 1

CFP-like species induced by YFP photobleaching appear at sites of intracellular YFP accumulation. (PDF 87 kb)

Supplementary Fig. 2

Venus and Citrine generate false FRET signals with the acceptor photo-bleaching method. (PDF 292 kb)

Supplementary Fig. 3

YFP proteins become photo-converted into CFP-like species during photo-bleaching at 514 nm. (PDF 381 kb)

Supplementary Methods (PDF 32 kb)

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Valentin, G., Verheggen, C., Piolot, T. et al. Photoconversion of YFP into a CFP-like species during acceptor photobleaching FRET experiments. Nat Methods 2, 801 (2005). https://doi.org/10.1038/nmeth1105-801

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