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Two-colour live-cell nanoscale imaging of intracellular targets

Author

Listed:
  • Francesca Bottanelli

    (Yale University School of Medicine)

  • Emil B. Kromann

    (Yale University School of Medicine
    Yale University)

  • Edward S. Allgeyer

    (Yale University School of Medicine)

  • Roman S. Erdmann

    (Yale University School of Medicine
    Yale University)

  • Stephanie Wood Baguley

    (Yale University School of Medicine)

  • George Sirinakis

    (Yale University School of Medicine
    Gurdon Institute, University of Cambridge)

  • Alanna Schepartz

    (Yale University
    Cellular and Developmental Biology, Yale University)

  • David Baddeley

    (Yale University School of Medicine
    Nanobiology Institute, Yale University)

  • Derek K. Toomre

    (Yale University School of Medicine
    Nanobiology Institute, Yale University)

  • James E. Rothman

    (Yale University School of Medicine
    Nanobiology Institute, Yale University)

  • Joerg Bewersdorf

    (Yale University School of Medicine
    Yale University
    Nanobiology Institute, Yale University)

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.

Suggested Citation

  • 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 & Jo, 2016. "Two-colour live-cell nanoscale imaging of intracellular targets," Nature Communications, Nature, vol. 7(1), pages 1-5, April.
    • Handle: RePEc:nat:natcom:v:7:y:2016:i:1:d:10.1038_ncomms10778
    DOI: 10.1038/ncomms10778
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    Cited by:

    1. Hannes Gonschior & Christopher Schmied & Rozemarijn Eva Van der Veen & Jenny Eichhorst & Nina Himmerkus & Jörg Piontek & Dorothee Günzel & Markus Bleich & Mikio Furuse & Volker Haucke & Martin Lehmann, 2022. "Nanoscale segregation of channel and barrier claudins enables paracellular ion flux," Nature Communications, Nature, vol. 13(1), pages 1-20, December.
    2. Cátia Silva Janota & Andreia Pinto & Anna Pezzarossa & Pedro Machado & Judite Costa & Pedro Campinho & Cláudio A. Franco & Edgar R. Gomes, 2022. "Shielding of actin by the endoplasmic reticulum impacts nuclear positioning," Nature Communications, Nature, vol. 13(1), pages 1-13, December.

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