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Setup and Troubleshooting of Volta Phase Plate Cryo-EM Data Collection

Ottilie von Loeffelholz 1 Bruno Klaholz 1, * 
* Corresponding author
1 Inserm U964 - CNRS UMR7104 - IGBMC - Centre for Integrative Biology - CBI
IGBMC - Institut de Génétique et de Biologie Moléculaire et Cellulaire
Abstract : Cryo electron microscopy (cryo-EM) has become a method of choice in structural biology to analyze isolated complexes and cellular structures. This implies adequate imaging of the specimen and advanced image-processing methods to obtain high-resolution 3D reconstructions. The use of a Volta phase plate in cryo-EM drastically increases the image contrast while being able to record images at high acceleration voltage and close to focus, i.e., at conditions where high-resolution information is best preserved. During image processing, higher contrast images can be aligned and classified better than lower quality ones resulting in increased data quality and the need for less data. Here, we give step-by-step guidelines on how to set up high-quality VPP cryo-EM single particle data collections, as exemplified by human ribosome data acquired during a one-day data collection session. Further, we describe specific technical details in image processing that differ from conventional single particle cryo-EM data analysis.
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https://www.hal.inserm.fr/inserm-03376627
Contributor : Bruno Klaholz Connect in order to contact the contributor
Submitted on : Monday, December 6, 2021 - 6:04:05 PM
Last modification on : Sunday, June 26, 2022 - 3:23:03 AM
Long-term archiving on: : Monday, March 7, 2022 - 7:47:36 PM

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Ottilie von Loeffelholz, Bruno Klaholz. Setup and Troubleshooting of Volta Phase Plate Cryo-EM Data Collection. Structural Proteomics, 2305, Springer US, pp.291-299, 2021, Methods in Molecular Biology, ⟨10.1007/978-1-0716-1406-8_14⟩. ⟨inserm-03376627⟩

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