Two-colour live-cell nanoscale imaging of intracellular targets

Francesca Bottanelli, Emil B. Kromann, Edward S. Allgeyer, Roman S. Erdmann, Stephanie Wood Baguley, George Sirinakis, Alanna Schepartz, David Baddeley, Derek K. Toomre, James E. Rothman and Joerg Bewersdorf ()
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Francesca Bottanelli: Yale University School of Medicine
Emil B. Kromann: Yale University School of Medicine
Edward S. Allgeyer: Yale University School of Medicine
Roman S. Erdmann: Yale University School of Medicine
Stephanie Wood Baguley: Yale University School of Medicine
George Sirinakis: Yale University School of Medicine
Alanna Schepartz: Yale University
David Baddeley: Yale University School of Medicine
Derek K. Toomre: Yale University School of Medicine
James E. Rothman: Yale University School of Medicine
Joerg Bewersdorf: Yale University School of Medicine

Nature Communications, 2016, vol. 7, issue 1, 1-5

Abstract: 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.

Date: 2016
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DOI: 10.1038/ncomms10778

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