Mechanistic investigation of mEos4b reveals a strategy to reduce track interruptions in sptPALM

Green-to-red photoconvertible fluorescent proteins repeatedly enter dark states, causing interrupted tracks in single-particle-tracking localization microscopy (sptPALM). We identified a long-lived dark state in photoconverted mEos4b that results from isomerization of the chromophore and efficiently...

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Veröffentlicht in:Nature methods Jg. 16; H. 8; S. 707 - 710
Hauptverfasser: De Zitter, Elke, Thédié, Daniel, Mönkemöller, Viola, Hugelier, Siewert, Beaudouin, Joël, Adam, Virgile, Byrdin, Martin, Van Meervelt, Luc, Dedecker, Peter, Bourgeois, Dominique
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States Nature Publishing Group 01.08.2019
Schlagworte:
Blinking
Chromophores
Fluorescence
Isomerization
Lasers
Light
Localization
Microscopy
Proteins
Recording
Tracking
ISSN:1548-7091, 1548-7105, 1548-7105
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Zusammenfassung:Green-to-red photoconvertible fluorescent proteins repeatedly enter dark states, causing interrupted tracks in single-particle-tracking localization microscopy (sptPALM). We identified a long-lived dark state in photoconverted mEos4b that results from isomerization of the chromophore and efficiently absorbs cyan light. Addition of weak 488-nm light swiftly reverts this dark state to the fluorescent state. This strategy largely eliminates slow blinking and enables the recording of longer tracks in sptPALM with minimum effort.
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ISSN:1548-7091
1548-7105
1548-7105
DOI:10.1038/s41592-019-0462-3