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Sci. Signal., 28 October 2008
Vol. 1, Issue 43, p. pl3
[DOI: 10.1126/scisignal.143pl3]

PROTOCOLS

High-Resolution Imaging of Redox Signaling in Live Cells Through an Oxidation-Sensitive Yellow Fluorescent Protein

Giuseppe Maulucci2, Valentina Labate1, Marina Mele1, Emiliano Panieri1, Giuseppe Arcovito2, Tommaso Galeotti1, Henrik Østergaard3, Jakob R. Winther4, Marco De Spirito2, and Giovambattista Pani1*

1 Institute of General Pathology, Catholic University Medical School, 00168 Rome, Italy.
2 Institute of Physics, Catholic University Medical School, 00168 Rome, Italy.
3 Novo Nordisk A/S, DK-2760 Måløv, Denmark.
4 Department of Molecular Biology, August Krogh Building, Universitetsparken 13, DK-2100 Copenhagen, Denmark.

Abstract: We present the application of a redox-sensitive mutant of the yellow fluorescent protein (rxYFP) to image, with elevated sensitivity and high temporal and spatial resolution, oxidative responses of eukaryotic cells to pathophysiological stimuli. The method presented, based on the ratiometric quantitation of the distribution of fluorescence by confocal microscopy, allows us to draw real-time "redox maps" of adherent cells and to score subtle changes in the intracellular redox state, such as those induced by overexpression of redox-active proteins. This strategy for in vivo imaging of redox signaling circumvents many of the technical limitations currently encountered in the study of complex redox-based phenomena and promises to contribute substantially to this expanding area of signal transduction.

* Corresponding author. E-mail, gpani{at}rm.unicatt.it

Citation: G. Maulucci, V. Labate, M. Mele, E. Panieri, G. Arcovito, T. Galeotti, H. Østergaard, J. R. Winther, M. De Spirito, G. Pani, High-Resolution Imaging of Redox Signaling in Live Cells Through an Oxidation-Sensitive Yellow Fluorescent Protein. Sci. Signal. 1, pl3 (2008).

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S. C. Ranieri, S. Fusco, E. Panieri, V. Labate, M. Mele, V. Tesori, A. M. Ferrara, G. Maulucci, M. De Spirito, G. E. Martorana, et al. (2010)
PNAS 107, 13420-13425
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