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Improved applicability and robustness of fast cryo-electron tomography data acquisition

The power of cryo-electron tomography (cryoET) lies in its capability to characterize macromolecules in their cellular context. Structure determination by cryoET, however, is time-consuming compared to single particle approaches. A recent study reported significant acceleration of data acquisition b...

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Detalles Bibliográficos
Autores principales: Eisenstein, Fabian, Danev, Radostin, Pilhofer, Martin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Academic Press 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6839400/
https://www.ncbi.nlm.nih.gov/pubmed/31425790
http://dx.doi.org/10.1016/j.jsb.2019.08.006
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author Eisenstein, Fabian
Danev, Radostin
Pilhofer, Martin
author_facet Eisenstein, Fabian
Danev, Radostin
Pilhofer, Martin
author_sort Eisenstein, Fabian
collection PubMed
description The power of cryo-electron tomography (cryoET) lies in its capability to characterize macromolecules in their cellular context. Structure determination by cryoET, however, is time-consuming compared to single particle approaches. A recent study reported significant acceleration of data acquisition by a fast-incremental single-exposure (FISE) tilt series scheme. Here we improved the method and evaluated its efficiency and performance. We show that (1) FISE combined with the latest generation of direct electron detectors speeds up collection considerably, (2) previous generation (pre-2017) double-tilt axis Titan Krios holders are also suitable for FISE data acquisition, (3) x, y and z-specimen shifts can be compensated for, and (4) FISE tilt series data can generate averages of sub-nanometer resolution. These advances will allow for a widespread adoption of cryoET for high-throughput in situ studies and high-resolution structure determination across different biological research disciplines.
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spelling pubmed-68394002019-11-12 Improved applicability and robustness of fast cryo-electron tomography data acquisition Eisenstein, Fabian Danev, Radostin Pilhofer, Martin J Struct Biol Article The power of cryo-electron tomography (cryoET) lies in its capability to characterize macromolecules in their cellular context. Structure determination by cryoET, however, is time-consuming compared to single particle approaches. A recent study reported significant acceleration of data acquisition by a fast-incremental single-exposure (FISE) tilt series scheme. Here we improved the method and evaluated its efficiency and performance. We show that (1) FISE combined with the latest generation of direct electron detectors speeds up collection considerably, (2) previous generation (pre-2017) double-tilt axis Titan Krios holders are also suitable for FISE data acquisition, (3) x, y and z-specimen shifts can be compensated for, and (4) FISE tilt series data can generate averages of sub-nanometer resolution. These advances will allow for a widespread adoption of cryoET for high-throughput in situ studies and high-resolution structure determination across different biological research disciplines. Academic Press 2019-11-01 /pmc/articles/PMC6839400/ /pubmed/31425790 http://dx.doi.org/10.1016/j.jsb.2019.08.006 Text en © 2019 The Author(s) http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
spellingShingle Article
Eisenstein, Fabian
Danev, Radostin
Pilhofer, Martin
Improved applicability and robustness of fast cryo-electron tomography data acquisition
title Improved applicability and robustness of fast cryo-electron tomography data acquisition
title_full Improved applicability and robustness of fast cryo-electron tomography data acquisition
title_fullStr Improved applicability and robustness of fast cryo-electron tomography data acquisition
title_full_unstemmed Improved applicability and robustness of fast cryo-electron tomography data acquisition
title_short Improved applicability and robustness of fast cryo-electron tomography data acquisition
title_sort improved applicability and robustness of fast cryo-electron tomography data acquisition
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6839400/
https://www.ncbi.nlm.nih.gov/pubmed/31425790
http://dx.doi.org/10.1016/j.jsb.2019.08.006
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