Perspective: Biochemical and Physical Constraints Associated With Preparing Thin Specimens for Single-Particle Cryo-EM

Han, Bong-Gyoon; Armstrong, Max; Fletcher, Daniel A.; Glaeser, Robert M.

doi:10.3389/fmolb.2022.864829

Perspective: Biochemical and Physical Constraints Associated With Preparing Thin Specimens for Single-Particle Cryo-EM

Journal Article · Tue Apr 26 00:00:00 EDT 2022 · Frontiers in Molecular Biosciences

DOI:https://doi.org/10.3389/fmolb.2022.864829· OSTI ID:1895336

Han, Bong-Gyoon ^[1]; Armstrong, Max ^[2]; Fletcher, Daniel A. ^[3]; Glaeser, Robert M. ^[1]

Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Univ. of California, Berkeley, CA (United States)
Univ. of California, Berkeley, CA (United States); Chan Zuckerberg Biohub, San Francisco, CA (United States)

While many aspects of single-particle electron cryo-microscopy (cryo-EM) of biological macromolecules have reached a sophisticated level of development, this is not yet the case when it comes to preparing thin samples on specimen grids. As a result, there currently is considerable interest in achieving better control of both the sample thickness and the amount of area that is useful, but this is only one aspect in which improvement is needed. This Perspective addresses the further need to prevent the macromolecular particles from making contact with the air-water interface, something that can result in preferential orientation and even structural disruption of macromolecular particles. This unwanted contact can occur either as the result of free diffusion of particles during the interval between application, thinning and vitrification of the remaining buffer, or—when particles have been immobilized—by the film of buffer becoming too thin prior to vitrification. An opportunity now exists to apply theoretical and practical insights from the fields of thin-film physical chemistry and interfacial science, in an effort to bring cryo-EM sample preparation to a level of sophistication that is comparable to that of current data collection and analysis.

View Accepted Manuscript (DOE)

Research Organization:: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)

Sponsoring Organization:: National Institutes of Health (NIH); USDOE Office of Science (SC), Biological and Environmental Research (BER)

Grant/Contract Number:: AC02-05CH11231

OSTI ID:: 1895336

Journal Information:: Frontiers in Molecular Biosciences, Journal Name: Frontiers in Molecular Biosciences Vol. 9; ISSN 2296-889X

Publisher:: Frontiers Media S.A.Copyright Statement

Country of Publication:: United States

Language:: English

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