Protein localization in electron micrographs using fluorescence nanoscopy

Shigeki Watanabe, Annedore Punge, Gunther Hollopeter, Katrin I. Willig, Robert John Hobson, M. Wayne Davis, Stefan W. Hell, Erik M. Jorgensen

Research output: Contribution to journalArticle

Abstract

A complete portrait of a cell requires a detailed description of its molecular topography: proteins must be linked to particular organelles. Immunocytochemical electron microscopy can reveal locations of proteins with nanometer resolution but is limited by the quality of fixation, the paucity of antibodies and the inaccessibility of antigens. Here we describe correlative fluorescence electron microscopy for the nanoscopic localization of proteins in electron micrographs. We tagged proteins with the fluorescent proteins Citrine or tdEos and expressed them in Caenorhabditis elegans, fixed the worms and embedded them in plastic. We imaged the tagged proteins from ultrathin sections using stimulated emission depletion (STED) microscopy or photoactivated localization microscopy (PALM). Fluorescence correlated with organelles imaged in electron micrographs from the same sections. We used these methods to localize histones, a mitochondrial protein and a presynaptic dense projection protein in electron micrographs.

Original languageEnglish (US)
Pages (from-to)80-84
Number of pages5
JournalNature Methods
Volume8
Issue number1
DOIs
StatePublished - Jan 2011
Externally publishedYes

Fingerprint

Fluorescence
Electrons
Proteins
Organelles
Electron microscopy
Microscopy
Electron Microscopy
Microscopic examination
Stimulated emission
Fluorescence microscopy
Mitochondrial Proteins
Caenorhabditis elegans
Fluorescence Microscopy
Histones
Topography
Plastics
Antigens
Antibodies

ASJC Scopus subject areas

  • Biotechnology
  • Molecular Biology
  • Biochemistry
  • Cell Biology

Cite this

Watanabe, S., Punge, A., Hollopeter, G., Willig, K. I., Hobson, R. J., Davis, M. W., ... Jorgensen, E. M. (2011). Protein localization in electron micrographs using fluorescence nanoscopy. Nature Methods, 8(1), 80-84. https://doi.org/10.1038/nmeth.1537

Protein localization in electron micrographs using fluorescence nanoscopy. / Watanabe, Shigeki; Punge, Annedore; Hollopeter, Gunther; Willig, Katrin I.; Hobson, Robert John; Davis, M. Wayne; Hell, Stefan W.; Jorgensen, Erik M.

In: Nature Methods, Vol. 8, No. 1, 01.2011, p. 80-84.

Research output: Contribution to journalArticle

Watanabe, S, Punge, A, Hollopeter, G, Willig, KI, Hobson, RJ, Davis, MW, Hell, SW & Jorgensen, EM 2011, 'Protein localization in electron micrographs using fluorescence nanoscopy', Nature Methods, vol. 8, no. 1, pp. 80-84. https://doi.org/10.1038/nmeth.1537
Watanabe S, Punge A, Hollopeter G, Willig KI, Hobson RJ, Davis MW et al. Protein localization in electron micrographs using fluorescence nanoscopy. Nature Methods. 2011 Jan;8(1):80-84. https://doi.org/10.1038/nmeth.1537
Watanabe, Shigeki ; Punge, Annedore ; Hollopeter, Gunther ; Willig, Katrin I. ; Hobson, Robert John ; Davis, M. Wayne ; Hell, Stefan W. ; Jorgensen, Erik M. / Protein localization in electron micrographs using fluorescence nanoscopy. In: Nature Methods. 2011 ; Vol. 8, No. 1. pp. 80-84.
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