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  Integrative Approaches for Cellular Cryo-electron Tomography: Correlative Imaging and Focused Ion Beam Micromachining

Rigort, A., Villa, E., Bäuerlein, F. J. B., Engel, B. D., & Plitzko, J. M. (2012). Integrative Approaches for Cellular Cryo-electron Tomography: Correlative Imaging and Focused Ion Beam Micromachining. In Methods in Cell Biology; Vol. 111 (pp. 259-281). San Diego: Academic Press.

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 Creators:
Rigort, Alexander1, Author              
Villa, Elizabeth1, Author              
Bäuerlein, Felix Jan Benedict1, Author              
Engel, Benjamin D.1, Author              
Plitzko, Jürgen M.1, Author              
Affiliations:
1Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565142              

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 Abstract: The application of cryo-electron tomography to cells and tissues is commonly referred to as “cellular tomography,” and enables visualization of the supramolecular architecture of cells in a near-native state. However, in order to access structural features hidden deep inside cellular volumes, it is necessary to use hybrid techniques to identify and localize features of interest and prepare such regions for subsequent analysis by transmission electron microscopy. We present a workflow that integrates different approaches: (1) correlative cryo-fluorescence microscopy to localize features within frozen-hydrated cells, (2) focused ion beam milling to thin these specimens in a targeted manner, and (3) cryo-electron tomography to provide detailed information about the cellular ultrastructure of thinned samples. We describe the combined use of these techniques and the instrumentation required to enable cryo-electron tomography for a vast range of cellular samples.

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Language(s): eng - English
 Dates: 2012
 Publication Status: Published in print
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 Rev. Type: Peer
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Title: Methods in Cell Biology; Vol. 111
  Alternative Title : Chapter 14
Source Genre: Series
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Publ. Info: San Diego : Academic Press
Pages: - Volume / Issue: 111 Sequence Number: - Start / End Page: 259 - 281 Identifier: ISSN: 0091-679X