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TitleHigh-speed high-resolution data collection on a 200 keV cryo-TEM.
Journal, issue, pagesIUCrJ, Vol. 9, Issue Pt 2, Page 243-252, Year 2022
Publish dateMar 1, 2022
AuthorsJared V Peck / Jonathan F Fay / Joshua D Strauss /
PubMed AbstractLimitations to successful single-particle cryo-electron microscopy (cryo-EM) projects include stable sample generation, production of quality cryo-EM grids with randomly oriented particles embedded ...Limitations to successful single-particle cryo-electron microscopy (cryo-EM) projects include stable sample generation, production of quality cryo-EM grids with randomly oriented particles embedded in thin vitreous ice and access to microscope time. To address the limitation of microscope time, methodologies to more efficiently collect data on a 200 keV Talos Arctica cryo-transmission electron microscope at speeds as fast as 720 movies per hour (∼17 000 per day) were tested. In this study, key parameters were explored to increase data collection speed including: (1) using the beam-image shift method to acquire multiple images per stage position, (2) employing UltrAufoil TEM grids with R0.6/1 hole spacing, (3) collecting hardware-binned data and (4) adjusting the image shift delay factor in . Here, eight EM maps of mouse apoferritin at 1.8-1.9 Å resolution were obtained in the analysis with data collection times for each dataset ranging from 56 min to 2 h. An EM map of mouse apoferritin at 1.78 Å was obtained from an overnight data collection at a speed of 500 movies per hour and subgroup analysis performed, with no significant variation observed in data quality by image shift distance and image shift delay. The findings and operating procedures detailed herein allow for rapid turnover of single-particle cryo-EM structure determination.
External linksIUCrJ / PubMed:35371504 / PubMed Central
MethodsEM (single particle)
Resolution1.78 - 2.19 Å
Structure data

EMDB-25839:
1.78A reconstruction of mouse heavy chain apoferritin collected at 500 micrographs/hr
Method: EM (single particle) / Resolution: 1.78 Å

EMDB-25840:
2.19A reconstruction of mouse heavy chain apoferritin collected at 720 micrographs/hr
Method: EM (single particle) / Resolution: 2.19 Å

EMDB-25841:
1.83A reconstruction of mouse heavy chain apoferritin collected at 520 micrographs/hr
Method: EM (single particle) / Resolution: 1.83 Å

EMDB-25842:
1.85A reconstruction of mouse heavy chain apoferritin collected at 346 micrographs/hr
Method: EM (single particle) / Resolution: 1.85 Å

Source
  • Mus musculus (house mouse)

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