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Nat Commun. 2016 Mar 4;7:10778. doi: 10.1038/ncomms10778.

Two-colour live-cell nanoscale imaging of intracellular targets.

Author information

1
Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
2
Department of Biomedical Engineering, Yale University, New Haven, Connecticut 06520, USA.
3
Department of Chemistry, Yale University, New Haven, Connecticut 06520, USA.
4
Gurdon Institute, University of Cambridge, Cambridge CB2 1QN, UK.
5
Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut 06520, USA.
6
Nanobiology Institute, Yale University, West Haven, Connecticut 06516, USA.

Abstract

Stimulated emission depletion (STED) nanoscopy allows observations of subcellular dynamics at the nanoscale. Applications have, however, been severely limited by the lack of a versatile STED-compatible two-colour labelling strategy for intracellular targets in living cells. Here we demonstrate a universal labelling method based on the organic, membrane-permeable dyes SiR and ATTO590 as Halo and SNAP substrates. SiR and ATTO590 constitute the first suitable dye pair for two-colour STED imaging in living cells below 50 nm resolution. We show applications with mitochondria, endoplasmic reticulum, plasma membrane and Golgi-localized proteins, and demonstrate continuous acquisition for up to 3 min at 2-s time resolution.

PMID:
26940217
PMCID:
PMC4785223
DOI:
10.1038/ncomms10778
[Indexed for MEDLINE]
Free PMC Article
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