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  1. Article ; Online: Plasma FIB milling for the determination of structures in situ

    Casper Berger / Maud Dumoux / Thomas Glen / Neville B.-y. Yee / John M. Mitchels / Zuzana Patáková / Michele C. Darrow / James H. Naismith / Michael Grange

    Nature Communications, Vol 14, Iss 1, Pp 1-

    2023  Volume 12

    Abstract: The authors harness plasma focused ion beams for pseudo-atomic structure determination, reporting increased throughput and automation in in situ structural biology to elucidate structure-function relationships inside cells and tissues. ...

    Abstract The authors harness plasma focused ion beams for pseudo-atomic structure determination, reporting increased throughput and automation in in situ structural biology to elucidate structure-function relationships inside cells and tissues.
    Keywords Science ; Q
    Language English
    Publishing date 2023-02-01T00:00:00Z
    Publisher Nature Portfolio
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Cryo-plasma FIB/SEM volume imaging of biological specimens

    Maud Dumoux / Thomas Glen / Jake LR Smith / Elaine ML Ho / Luis MA Perdigão / Avery Pennington / Sven Klumpe / Neville BY Yee / David Andrew Farmer / Pui YA Lai / William Bowles / Ron Kelley / Jürgen M Plitzko / Liang Wu / Mark Basham / Daniel K Clare / C Alistair Siebert / Michele C Darrow / James H Naismith /
    Michael Grange

    eLife, Vol

    2023  Volume 12

    Abstract: Serial focussed ion beam scanning electron microscopy (FIB/SEM) enables imaging and assessment of subcellular structures on the mesoscale (10 nm to 10 µm). When applied to vitrified samples, serial FIB/SEM is also a means to target specific structures in ...

    Abstract Serial focussed ion beam scanning electron microscopy (FIB/SEM) enables imaging and assessment of subcellular structures on the mesoscale (10 nm to 10 µm). When applied to vitrified samples, serial FIB/SEM is also a means to target specific structures in cells and tissues while maintaining constituents’ hydration shells for in situ structural biology downstream. However, the application of serial FIB/SEM imaging of non-stained cryogenic biological samples is limited due to low contrast, curtaining, and charging artefacts. We address these challenges using a cryogenic plasma FIB/SEM. We evaluated the choice of plasma ion source and imaging regimes to produce high-quality SEM images of a range of different biological samples. Using an automated workflow we produced three-dimensional volumes of bacteria, human cells, and tissue, and calculated estimates for their resolution, typically achieving 20–50 nm. Additionally, a tag-free localisation tool for regions of interest is needed to drive the application of in situ structural biology towards tissue. The combination of serial FIB/SEM with plasma-based ion sources promises a framework for targeting specific features in bulk-frozen samples (>100 µm) to produce lamellae for cryogenic electron tomography.
    Keywords HeLa cells ; Vero cells ; mouse tissue (brain) ; mouse tissue (cardiac) ; S. cerevisiae ; R. rubrum ; Medicine ; R ; Science ; Q ; Biology (General) ; QH301-705.5
    Subject code 500
    Language English
    Publishing date 2023-02-01T00:00:00Z
    Publisher eLife Sciences Publications Ltd
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article: Structural Basis of Vesicle Formation at the Inner Nuclear Membrane

    Hagen, Christoph / Kyle C. Dent / Tzviya Zeev-Ben-Mordehai / Michael Grange / Jens B. Bosse / Cathy Whittle / Barbara G. Klupp / C. Alistair Siebert / Daven Vasishtan / Felix J.B. Bäuerlein / Juliana Cheleski / Stephan Werner / Peter Guttmann / Stefan Rehbein / Katja Henzler / Justin Demmerle / Barbara Adler / Ulrich Koszinowski / Lothar Schermelleh /
    Gerd Schneider / Lynn W. Enquist / Jürgen M. Plitzko / Thomas C. Mettenleiter / Kay Grünewald

    Cell. 2015 Dec. 17, v. 163

    2015  

    Abstract: Vesicular nucleo-cytoplasmic transport is becoming recognized as a general cellular mechanism for translocation of large cargoes across the nuclear envelope. Cargo is recruited, enveloped at the inner nuclear membrane (INM), and delivered by membrane ... ...

    Abstract Vesicular nucleo-cytoplasmic transport is becoming recognized as a general cellular mechanism for translocation of large cargoes across the nuclear envelope. Cargo is recruited, enveloped at the inner nuclear membrane (INM), and delivered by membrane fusion at the outer nuclear membrane. To understand the structural underpinning for this trafficking, we investigated nuclear egress of progeny herpesvirus capsids where capsid envelopment is mediated by two viral proteins, forming the nuclear egress complex (NEC). Using a multi-modal imaging approach, we visualized the NEC in situ forming coated vesicles of defined size. Cellular electron cryo-tomography revealed a protein layer showing two distinct hexagonal lattices at its membrane-proximal and membrane-distant faces, respectively. NEC coat architecture was determined by combining this information with integrative modeling using small-angle X-ray scattering data. The molecular arrangement of the NEC establishes the basic mechanism for budding and scission of tailored vesicles at the INM.
    Keywords X-radiation ; capsid ; coated vesicles ; image analysis ; membrane fusion ; models ; nuclear membrane ; nucleocytoplasmic transport ; progeny ; viral proteins
    Language English
    Dates of publication 2015-1217
    Size p. 1692-1701.
    Publishing place Elsevier Inc.
    Document type Article
    ZDB-ID 187009-9
    ISSN 1097-4172 ; 0092-8674
    ISSN (online) 1097-4172
    ISSN 0092-8674
    DOI 10.1016/j.cell.2015.11.029
    Database NAL-Catalogue (AGRICOLA)

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