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  A cryo-FIB lift-out technique enables molecular-resolution cryo-ET within native Caenorhabditis elegans tissue

Schaffer, M., Pfeffer, S., Mahamid, J., Kleindiek, S., Laugks, T., Albert, S., et al. (2019). A cryo-FIB lift-out technique enables molecular-resolution cryo-ET within native Caenorhabditis elegans tissue. Nature Methods, 16(8), 757-762. doi:10.1038/s41592-019-0497-5.

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Schaffer, Miroslava1, Autor           
Pfeffer, Stefan2, Autor
Mahamid, Julia2, Autor
Kleindiek, Stephan2, Autor
Laugks, Tim1, Autor           
Albert, Sahradha1, Autor           
Engel, Benjamin D.1, Autor           
Rummel, Andreas2, Autor
Smith, Andrew J.2, Autor
Baumeister, Wolfgang1, Autor           
Plitzko, Juergen M.1, Autor           
Affiliations:
1Baumeister, Wolfgang / Molecular Structural Biology, Max Planck Institute of Biochemistry, Max Planck Society, ou_1565142              
2external, ou_persistent22              

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Schlagwörter: SAMPLE PREPARATION; CRYOELECTRON TOMOGRAPHY; BEAM; CELLS; SOCIOLOGY; STEMBiochemistry & Molecular Biology;
 Zusammenfassung: Cryo-focused ion beam milling of frozen-hydrated cells has recently provided unprecedented insights into the inner space of cells. In combination with cryo-electron tomography, this method allows access to native structures deep inside cells, enabling structural studies of macromolecules in situ. However, this approach has been mainly limited to individual cells that can be completely vitrified by plunge-freezing. Here, we describe a preparation method that is based on the targeted extraction of material from high-pressure-frozen bulk specimens with a cryo-gripper tool. This lift-out technique enables cryo-electron tomography to be performed on multicellular organisms and tissue, extending the range of applications for in situ structural biology. We demonstrate the potential of the lift-out technique with a structural study of cytosolic 80S ribosomes in a Caenorhabditis elegans worm. The preparation quality allowed for subtomogram analysis with sufficient resolution to distinguish individual ribosomal translocation states and revealed significant cell-to-cell variation in ribosome structure.

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Sprache(n): eng - English
 Datum: 2019
 Publikationsstatus: Erschienen
 Seiten: 8
 Ort, Verlag, Ausgabe: -
 Inhaltsverzeichnis: -
 Art der Begutachtung: Expertenbegutachtung
 Identifikatoren: ISI: 000477857700033
DOI: 10.1038/s41592-019-0497-5
 Art des Abschluß: -

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Titel: Nature Methods
  Andere : Nature Methods
Genre der Quelle: Zeitschrift
 Urheber:
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Ort, Verlag, Ausgabe: New York, NY : Nature Pub. Group
Seiten: - Band / Heft: 16 (8) Artikelnummer: - Start- / Endseite: 757 - 762 Identifikator: ISSN: 1548-7091
CoNE: https://pure.mpg.de/cone/journals/resource/111088195279556