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Plasma FIB milling for the determination of structures in situ

Author

Listed:
  • Casper Berger

    (Harwell Science & Innovation Campus
    University of Oxford)

  • Maud Dumoux

    (Harwell Science & Innovation Campus)

  • Thomas Glen

    (Harwell Science & Innovation Campus)

  • Neville B.-y. Yee

    (Harwell Science & Innovation Campus)

  • John M. Mitchels

    (Thermo Fisher Scientific Brno s.r.o)

  • Zuzana Patáková

    (Thermo Fisher Scientific Brno s.r.o)

  • Michele C. Darrow

    (Harwell Science & Innovation Campus)

  • James H. Naismith

    (Harwell Science & Innovation Campus
    University of Oxford)

  • Michael Grange

    (Harwell Science & Innovation Campus
    University of Oxford)

Abstract

Structural biology studies inside cells and tissues require methods to thin vitrified specimens to electron transparency. Until now, focused ion beams based on gallium have been used. However, ion implantation, changes to surface chemistry and an inability to access high currents limit gallium application. Here, we show that plasma-coupled ion sources can produce cryogenic lamellae of vitrified human cells in a robust and automated manner, with quality sufficient for pseudo-atomic structure determination. Lamellae were produced in a prototype microscope equipped for long cryogenic run times (> 1 week) and with multi-specimen support fully compatible with modern-day transmission electron microscopes. We demonstrate that plasma ion sources can be used for structural biology within cells, determining a structure in situ to 4.9 Å, and characterise the resolution dependence on particle distance from the lamella edge. We describe a workflow upon which different plasmas can be examined to further streamline lamella fabrication.

Suggested Citation

  • Casper Berger & Maud Dumoux & Thomas Glen & Neville B.-y. Yee & John M. Mitchels & Zuzana Patáková & Michele C. Darrow & James H. Naismith & Michael Grange, 2023. "Plasma FIB milling for the determination of structures in situ," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-36372-9
    DOI: 10.1038/s41467-023-36372-9
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    References listed on IDEAS

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    Cited by:

    1. Nikita Balyschew & Artsemi Yushkevich & Vasilii Mikirtumov & Ricardo M. Sanchez & Thiemo Sprink & Mikhail Kudryashev, 2023. "Streamlined structure determination by cryo-electron tomography and subtomogram averaging using TomoBEAR," Nature Communications, Nature, vol. 14(1), pages 1-11, December.
    2. Benjamin C. Creekmore & Kathryn Kixmoeller & Ben E. Black & Edward B. Lee & Yi-Wei Chang, 2024. "Ultrastructure of human brain tissue vitrified from autopsy revealed by cryo-ET with cryo-plasma FIB milling," Nature Communications, Nature, vol. 15(1), pages 1-12, December.
    3. Zhen Hou & Frank Nightingale & Yanan Zhu & Craig MacGregor-Chatwin & Peijun Zhang, 2023. "Structure of native chromatin fibres revealed by Cryo-ET in situ," Nature Communications, Nature, vol. 14(1), pages 1-7, December.

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