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Approaches to altering particle distributions in cryo-electron microscopy sample preparation

Cryo-electron microscopy (cryo-EM) can now be used to determine high-resolution structural information on a diverse range of biological specimens. Recent advances have been driven primarily by developments in microscopes and detectors, and through advances in image-processing software. However, for...

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Autores principales: Drulyte, Ieva, Johnson, Rachel M., Hesketh, Emma L., Hurdiss, Daniel L., Scarff, Charlotte A., Porav, Sebastian A., Ranson, Neil A., Muench, Stephen P., Thompson, Rebecca F.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: International Union of Crystallography 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6096488/
https://www.ncbi.nlm.nih.gov/pubmed/29872006
http://dx.doi.org/10.1107/S2059798318006496
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author Drulyte, Ieva
Johnson, Rachel M.
Hesketh, Emma L.
Hurdiss, Daniel L.
Scarff, Charlotte A.
Porav, Sebastian A.
Ranson, Neil A.
Muench, Stephen P.
Thompson, Rebecca F.
author_facet Drulyte, Ieva
Johnson, Rachel M.
Hesketh, Emma L.
Hurdiss, Daniel L.
Scarff, Charlotte A.
Porav, Sebastian A.
Ranson, Neil A.
Muench, Stephen P.
Thompson, Rebecca F.
author_sort Drulyte, Ieva
collection PubMed
description Cryo-electron microscopy (cryo-EM) can now be used to determine high-resolution structural information on a diverse range of biological specimens. Recent advances have been driven primarily by developments in microscopes and detectors, and through advances in image-processing software. However, for many single-particle cryo-EM projects, major bottlenecks currently remain at the sample-preparation stage; obtaining cryo-EM grids of sufficient quality for high-resolution single-particle analysis can require the careful optimization of many variables. Common hurdles to overcome include problems associated with the sample itself (buffer components, labile complexes), sample distribution (obtaining the correct concentration, affinity for the support film), preferred orientation, and poor reproducibility of the grid-making process within and between batches. This review outlines a number of methodologies used within the electron-microscopy community to address these challenges, providing a range of approaches which may aid in obtaining optimal grids for high-resolution data collection.
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spelling pubmed-60964882018-08-24 Approaches to altering particle distributions in cryo-electron microscopy sample preparation Drulyte, Ieva Johnson, Rachel M. Hesketh, Emma L. Hurdiss, Daniel L. Scarff, Charlotte A. Porav, Sebastian A. Ranson, Neil A. Muench, Stephen P. Thompson, Rebecca F. Acta Crystallogr D Struct Biol Research Papers Cryo-electron microscopy (cryo-EM) can now be used to determine high-resolution structural information on a diverse range of biological specimens. Recent advances have been driven primarily by developments in microscopes and detectors, and through advances in image-processing software. However, for many single-particle cryo-EM projects, major bottlenecks currently remain at the sample-preparation stage; obtaining cryo-EM grids of sufficient quality for high-resolution single-particle analysis can require the careful optimization of many variables. Common hurdles to overcome include problems associated with the sample itself (buffer components, labile complexes), sample distribution (obtaining the correct concentration, affinity for the support film), preferred orientation, and poor reproducibility of the grid-making process within and between batches. This review outlines a number of methodologies used within the electron-microscopy community to address these challenges, providing a range of approaches which may aid in obtaining optimal grids for high-resolution data collection. International Union of Crystallography 2018-05-18 /pmc/articles/PMC6096488/ /pubmed/29872006 http://dx.doi.org/10.1107/S2059798318006496 Text en © Drulyte et al. 2018 http://creativecommons.org/licenses/by/2.0/uk/ This is an open-access article distributed under the terms of the Creative Commons Attribution (CC-BY) Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.http://creativecommons.org/licenses/by/2.0/uk/
spellingShingle Research Papers
Drulyte, Ieva
Johnson, Rachel M.
Hesketh, Emma L.
Hurdiss, Daniel L.
Scarff, Charlotte A.
Porav, Sebastian A.
Ranson, Neil A.
Muench, Stephen P.
Thompson, Rebecca F.
Approaches to altering particle distributions in cryo-electron microscopy sample preparation
title Approaches to altering particle distributions in cryo-electron microscopy sample preparation
title_full Approaches to altering particle distributions in cryo-electron microscopy sample preparation
title_fullStr Approaches to altering particle distributions in cryo-electron microscopy sample preparation
title_full_unstemmed Approaches to altering particle distributions in cryo-electron microscopy sample preparation
title_short Approaches to altering particle distributions in cryo-electron microscopy sample preparation
title_sort approaches to altering particle distributions in cryo-electron microscopy sample preparation
topic Research Papers
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6096488/
https://www.ncbi.nlm.nih.gov/pubmed/29872006
http://dx.doi.org/10.1107/S2059798318006496
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