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Enhanced labeling density and whole-cell 3D dSTORM imaging by repetitive labeling of target proteins
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| Veröffentlicht in: | Scientific reports 8(2018) Artikel-Nummer 5507, 7 Seiten |
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| Personen und Körperschaften: | , , , , |
| Titel: | Enhanced labeling density and whole-cell 3D dSTORM imaging by repetitive labeling of target proteins/ Varun Venkataramani, Markus Kardorff, Frank Herrmannsdörfer, Ralph Wieneke, Alina Klein, Robert Tampé, Mike Heilemann & Thomas Kuner |
| Format: | E-Book-Kapitel |
| Sprache: | Englisch |
| veröffentlicht: |
2018
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| Gesamtaufnahme: |
: Scientific reports, 8(2018) Artikel-Nummer 5507, 7 Seiten
, volume:8 |
| Quelle: | Verbunddaten SWB Lizenzfreie Online-Ressourcen |
| Zusammenfassung: | With continuing advances in the resolving power of super-resolution microscopy, the inefficient labeling of proteins with suitable fluorophores becomes a limiting factor. For example, the low labeling density achieved with antibodies or small molecule tags limits attempts to reveal local protein nano-architecture of cellular compartments. On the other hand, high laser intensities cause photobleaching within and nearby an imaged region, thereby further reducing labeling density and impairing multi-plane whole-cell 3D super-resolution imaging. Here, we show that both labeling density and photobleaching can be addressed by repetitive application of trisNTA-fluorophore conjugates reversibly binding to a histidine-tagged protein by a novel approach called single-epitope repetitive imaging (SERI). For single-plane super-resolution microscopy, we demonstrate that, after multiple rounds of labeling and imaging, the signal density is increased. Using the same approach of repetitive imaging, washing and re-labeling, we demonstrate whole-cell 3D super-resolution imaging compensated for photobleaching above or below the imaging plane. This proof-of-principle study demonstrates that repetitive labeling of histidine-tagged proteins provides a versatile solution to break the ‘labeling barrier’ and to bypass photobleaching in multi-plane, whole-cell 3D experiments. |
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| Beschreibung: |
Published online: 03 April 2018 Gesehen am 14.06.2018 |
| Umfang: |
Illustrationen 7 |
| ISSN: |
2045-2322
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| DOI: | 10.1038/s41598-018-23818-0 |