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Proceedings Paper

Single molecule FRET using the FRET pair DRONPA/PhotoActivable mCherry
Author(s): Viviane Devauges; Elena Ortiz-Zapater; Christina Efthymiou; Melanie Keppler; Jody Barbeau; Daniel R. Matthews; James Monypenny of Pitmilly; Paul R. Barber; Daniel Rolfe; Tony Ng; Simon M. Ameer-Beg
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Paper Abstract

Photoswitchable and photoactivable proteins Dronpa and PhotoActivable mCherry (PA-mCherry) respectively, were used in order to perform FRET (Förster Resonance Energy Transfer) imaging at the single molecule level, using a FRET standard construct consisting of Dronpa and PA-mCherry separated by seven amino acids expressed in cells. Given Dronpa’s complex photophysical properties and the existence of a preswitched emissive state, irradiation conditions at 491 and 405 nm were optimised. We discuss strategies for observing FRET at the single molecule level with photoactivatable proteins by monitoring modifications in the donor and acceptors emissive states.

Paper Details

Date Published: 22 February 2013
PDF: 9 pages
Proc. SPIE 8590, Single Molecule Spectroscopy and Superresolution Imaging VI, 85900A (22 February 2013); doi: 10.1117/12.2002470
Show Author Affiliations
Viviane Devauges, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Elena Ortiz-Zapater, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Christina Efthymiou, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Melanie Keppler, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Jody Barbeau, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Daniel R. Matthews, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Queensland Brain Institute, The Univ. of Queensland (Australia)
James Monypenny of Pitmilly, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Paul R. Barber, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Univ. of Oxford (United Kingdom)
Daniel Rolfe, Rutherford Appleton Lab. (United Kingdom)
Tony Ng, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)
Simon M. Ameer-Beg, Richard Dimbleby Cancer Research Lab., King's College London (United Kingdom)


Published in SPIE Proceedings Vol. 8590:
Single Molecule Spectroscopy and Superresolution Imaging VI
Jörg Enderlein; Ingo Gregor; Zygmunt Karol Gryczynski; Rainer Erdmann; Felix Koberling, Editor(s)

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