Research Papers

Enhanced yellow fluorescent protein photoconversion to a cyan fluorescent protein-like species is sensitive to thermal and diffusion conditions

[+] Author Affiliations
Merete K. Raarup

Aarhus University, Stereology and Electron Microscopy Research Laboratory, Histoinformatics and MIND Centres, Ole Worms Allé 1185, DK-8000 Aarhus C., Denmark

Anja W. Fjorback

Aarhus University, Stereology and Electron Microscopy Research Laboratory, Histoinformatics and MIND Centres, Ole Worms Allé 1185, DK-8000 Aarhus C., Denmark and Aarhus University Hospital, Centre for Psychiatric Research, Skovagervej 2, DK-8240 Risskov, Denmark

Stig M. R. Jensen

Aarhus University, Department of Molecular Biology, C.F. Møllers Allé 1130, DK-8000 Aarhus C., Denmark

Heidi K. Müller

Aarhus University Hospital, Centre for Psychiatric Research, Skovagervej 2, DK-8240 Risskov, Denmark

Maj M. Kjærgaard

Stereology and Electron Microscopy Research Laboratory, Histoinformatics and MIND Centres, Ole Worms Allé 1185, DK-8000 Aarhus C., Denmark

Hanne Poulsen

Aarhus University, Department of Molecular Biology, Gustav Wieds Vej 10 C, DK-8000 Aarhus C., Denmark

Ove Wiborg

Aarhus University Hospital, Centre for Psychiatric Research, Skovagervej 2, DK-8240 Risskov, Denmark

Jens R. Nyengaard

Stereology and Electron Microscopy Research Laboratory, Histoinformatics and MIND Centres, Ole Worms Allé 1185, DK-8000 Aarhus C., Denmark

J. Biomed. Opt. 14(3), 034039 (May 27, 2009). doi:10.1117/1.3103338
History: Received January 19, 2009; Accepted January 28, 2009; Published May 27, 2009
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Ongoing research efforts into fluorescent proteins continuously generates new mutation variants, some of which can become photoactivated or photoconverted to a red-shifted color upon intense UV or blue light illumination. We report a built-in propensity for enhanced yellow fluorescent protein (EYFP) to undergo irreversible photoconversion into a cyan fluorescent protein (CFP)–like species upon green-light illumination. The photoconversion is thermally activated, happens mainly in fixed, nonsealed cell samples, and may result in a very bright and relatively photostable CFP-like species. The photoconversion efficiency depends on the sample diffusivity and is much increased in dehydrated, oxygenated samples. Given the large variations in conversion efficiency observed among samples as well as within a sample, photoconversion cannot be appropriately accounted for in the analysis of acceptor photobleaching fluorescence resonance energy transfer (pbFRET) images and should rather be completely avoided. Thus, samples should always be checked and discarded if photoconversion is observed.

Figures in this Article
© 2009 Society of Photo-Optical Instrumentation Engineers

Citation

Merete K. Raarup ; Anja W. Fjorback ; Stig M. R. Jensen ; Heidi K. Müller ; Maj M. Kjærgaard, et al.
"Enhanced yellow fluorescent protein photoconversion to a cyan fluorescent protein-like species is sensitive to thermal and diffusion conditions", J. Biomed. Opt. 14(3), 034039 (May 27, 2009). ; http://dx.doi.org/10.1117/1.3103338


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