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Montage electron tomography of vitrified specimens

Peck, Ariana and Carter, Stephen D. and Mai, Huanghao and Chen, Songye and Burt, Alister and Jensen, Grant J. (2022) Montage electron tomography of vitrified specimens. Journal of Structural Biology, 214 (2). Art. No. 107860. ISSN 1047-8477. doi:10.1016/j.jsb.2022.107860. https://resolver.caltech.edu/CaltechAUTHORS:20211105-210618923

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Abstract

Cryo-electron tomography provides detailed views of macromolecules in situ. However, imaging a large field of view to provide more cellular context requires reducing magnification during data collection, which in turn restricts the resolution. To circumvent this trade-off between field of view and resolution, we have developed a montage data collection scheme that uniformly distributes the dose throughout the specimen. In this approach, sets of slightly overlapping circular tiles are collected at high magnification and stitched to form a composite projection image at each tilt angle. These montage tilt-series are then reconstructed into massive tomograms with a small pixel size but a large field of view. For proof-of-principle, we applied this method to the thin edge of HeLa cells. Thon rings to better than 10 Å were detected in the montaged tilt-series, and diverse cellular features were observed in the resulting tomograms. These results indicate that the additional dose required by this technique is not prohibitive to performing structural analysis to intermediate resolution across a large field of view. We anticipate that montage tomography will prove particularly useful for lamellae, increase the likelihood of imaging rare cellular events, and facilitate visual proteomics.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1016/j.jsb.2022.107860DOIArticle
https://doi.org/10.1101/2021.11.02.466666DOIDiscussion Paper
http://dx.doi.org/10.22002/D1.2096DOIData
http://dx.doi.org/10.22002/D1.2099DOIData
http://dx.doi.org/10.22002/D1.2103DOIData
https://github.com/apeck12/montageRelated ItemCode
https://etdb.caltech.edu/Related ItemCaltech Electron Tomography Database
ORCID:
AuthorORCID
Peck, Ariana0000-0002-5940-3897
Carter, Stephen D.0000-0002-4237-4276
Mai, Huanghao0000-0003-2278-0768
Chen, Songye0000-0001-5407-5049
Burt, Alister0000-0002-9341-2295
Jensen, Grant J.0000-0003-1556-4864
Additional Information:© 2022 Elsevier Inc. Received 15 November 2021, Revised 25 March 2022, Accepted 20 April 2022, Available online 26 April 2022, Version of Record 6 May 2022. Data collection and analysis were respectively performed at the Beckman Institute Resource Center for Transmission Electron Microscopy and the Resnick High Performance Computing Center at Caltech. We thank Wei Zhao for preliminary samples, David Mastronarde for valuable discussions, and Tom Morrell for help with the Caltech Data Repository. A.P. is The Mark Foundation for Cancer Research Fellow of the Damon Runyon Cancer Research Foundation (DRG 2361-19). This work was supported by NIH grant AI127401 to G.J.J. Data and code availability: Raw data are available at the Caltech Data Repository (https://data.caltech.edu) under accession IDs 2096, 2099, and 2103. The processed tilt-series binned to 5.3 Å can be found in the Caltech Electron Tomography Database (https://etdb.caltech.edu/). The code developed for data collection and processing is available at https://github.com/apeck12/montage. CRediT authorship contribution statement: Ariana Peck: Methodology, Software, Writing – original draft, Writing – review & editing. Stephen D. Carter: Conceptualization, Methodology, Investigation. Huanghao Mai: Methodology, Software. Songye Chen: Software, Methodology, Investigation. Alister Burt: Formal analysis, Writing – review & editing. Grant J. Jensen: Conceptualization, Writing – original draft, Writing – review & editing, Funding acquisition, Supervision. Declaration of Competing Interest: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Funders:
Funding AgencyGrant Number
Damon Runyon Cancer Research FoundationDRG 2361-19
NIHAI150464
Subject Keywords:Cryo-electron microscopy; Montage tomography; Image processing; Ultrastructural analysis; Cellular biology
Issue or Number:2
DOI:10.1016/j.jsb.2022.107860
Record Number:CaltechAUTHORS:20211105-210618923
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20211105-210618923
Official Citation:Ariana Peck, Stephen D. Carter, Huanghao Mai, Songye Chen, Alister Burt, Grant J. Jensen, Montage electron tomography of vitrified specimens, Journal of Structural Biology, Volume 214, Issue 2, 2022, 107860, ISSN 1047-8477, https://doi.org/10.1016/j.jsb.2022.107860. (https://www.sciencedirect.com/science/article/pii/S1047847722000302)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:111774
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:05 Nov 2021 21:51
Last Modified:17 May 2022 19:55

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