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Title: X-ray ptychographic and fluorescence microscopy of frozen-hydrated cells using continuous scanning

Abstract

X-ray microscopy can be used to image whole, unsectioned cells in their native hydrated state. It complements the higher resolution of electron microscopy for submicrometer thick specimens, and the molecule-specific imaging capabilites of fluorescence light microscopy. We describe here the first use of fast, continuous x-ray scanning of frozen hydrated cells for simultaneous sub-20 nm resolution ptychographic transmission imaging with high contrast, and sub-100 nm resolution deconvolved x-ray fluorescence imaging of diffusible and bound ions at native concentrations, without the need to add specific labels. Here, by working with cells that have been rapidly frozen without the use of chemical fixatives, and imaging them under cryogenic conditions, we are able to obtain images with well preserved structural and chemical composition, and sufficient stability against radiation damage to allow for multiple images to be obtained with no observable change.

Authors:
 [1];  [2];  [3];  [4];  [3];  [3];  [3]; ORCiD logo [5]
  1. Northwestern Univ., Evanston, IL (United States); Argonne National Lab. (ANL), Argonne, IL (United States)
  2. Argonne National Lab. (ANL), Argonne, IL (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
  3. Argonne National Lab. (ANL), Argonne, IL (United States)
  4. Northwestern Univ., Evanston, IL (United States)
  5. Argonne National Lab. (ANL), Argonne, IL (United States); Northwestern Univ., Evanston, IL (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22); National Institutes of Health (NIH)
OSTI Identifier:
1363812
Alternate Identifier(s):
OSTI ID: 1411588
Grant/Contract Number:
AC02-06CH11357; AC02-05CH11231
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Scientific Reports
Additional Journal Information:
Journal Volume: 7; Journal Issue: 1; Journal ID: ISSN 2045-2322
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
73 NUCLEAR PHYSICS AND RADIATION PHYSICS; microscopy; optical techniques

Citation Formats

Deng, Junjing, Vine, David J., Chen, Si, Jin, Qiaoling, Nashed, Youssef S. G., Peterka, Tom, Vogt, Stefan, and Jacobsen, Chris. X-ray ptychographic and fluorescence microscopy of frozen-hydrated cells using continuous scanning. United States: N. p., 2017. Web. doi:10.1038/s41598-017-00569-y.
Deng, Junjing, Vine, David J., Chen, Si, Jin, Qiaoling, Nashed, Youssef S. G., Peterka, Tom, Vogt, Stefan, & Jacobsen, Chris. X-ray ptychographic and fluorescence microscopy of frozen-hydrated cells using continuous scanning. United States. doi:10.1038/s41598-017-00569-y.
Deng, Junjing, Vine, David J., Chen, Si, Jin, Qiaoling, Nashed, Youssef S. G., Peterka, Tom, Vogt, Stefan, and Jacobsen, Chris. Mon . "X-ray ptychographic and fluorescence microscopy of frozen-hydrated cells using continuous scanning". United States. doi:10.1038/s41598-017-00569-y. https://www.osti.gov/servlets/purl/1363812.
@article{osti_1363812,
title = {X-ray ptychographic and fluorescence microscopy of frozen-hydrated cells using continuous scanning},
author = {Deng, Junjing and Vine, David J. and Chen, Si and Jin, Qiaoling and Nashed, Youssef S. G. and Peterka, Tom and Vogt, Stefan and Jacobsen, Chris},
abstractNote = {X-ray microscopy can be used to image whole, unsectioned cells in their native hydrated state. It complements the higher resolution of electron microscopy for submicrometer thick specimens, and the molecule-specific imaging capabilites of fluorescence light microscopy. We describe here the first use of fast, continuous x-ray scanning of frozen hydrated cells for simultaneous sub-20 nm resolution ptychographic transmission imaging with high contrast, and sub-100 nm resolution deconvolved x-ray fluorescence imaging of diffusible and bound ions at native concentrations, without the need to add specific labels. Here, by working with cells that have been rapidly frozen without the use of chemical fixatives, and imaging them under cryogenic conditions, we are able to obtain images with well preserved structural and chemical composition, and sufficient stability against radiation damage to allow for multiple images to be obtained with no observable change.},
doi = {10.1038/s41598-017-00569-y},
journal = {Scientific Reports},
number = 1,
volume = 7,
place = {United States},
year = {Mon Mar 27 00:00:00 EDT 2017},
month = {Mon Mar 27 00:00:00 EDT 2017}
}

Journal Article:
Free Publicly Available Full Text
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Cited by: 7works
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  • X-ray microscopy can be used to image whole, unsectioned cells in their native hydrated state. It complements the higher resolution of electron microscopy for submicrometer thick specimens, and the molecule-specific imaging capabilites of fluorescence light microscopy. We describe here the first use of fast, continuous x-ray scanning of frozen hydrated cells for simultaneous sub-20 nm resolution ptychographic transmission imaging with high contrast, and sub-100 nm resolution deconvolved x-ray fluorescence imaging of diffusible and bound ions at native concentrations, without the need to add specific labels. Here, by working with cells that have been rapidly frozen without the use of chemicalmore » fixatives, and imaging them under cryogenic conditions, we are able to obtain images with well preserved structural and chemical composition, and sufficient stability against radiation damage to allow for multiple images to be obtained with no observable change.« less
  • Bionanoprobe (BNP), a hard x-ray fluorescence sample-scanning nanoprobe at the Advanced Photon Source of Argonne National Laboratory, has been used to quantitatively study elemental distributions in biological cells with sub-100 nm spatial resolution and high sensitivity. Cryogenic conditions enable biological samples to be studied in their frozen-hydrated state with both ultrastructure and elemental distributions more faithfully preserved compared to conventional chemical fixation or dehydration methods. Furthermore, radiation damage is reduced in two ways: the diffusion rate of free radicals is decreased at low temperatures; and the sample is embedded in vitrified ice, which reduces mass loss.
  • Trace metals play important roles in normal and in disease-causing biological functions. X-ray fluorescence microscopy reveals trace elements with no dependence on binding affinities (unlike with visible light fluorophores) and with improved sensitivity relative to electron probes. However, X-ray fluorescence is not very sensitive for showing the light elements that comprise the majority of cellular material. Here we show that X-ray ptychography can be combined with fluorescence to image both cellular structure and trace element distribution in frozen-hydrated cells at cryogenic temperatures, with high structural and chemical fidelity. Ptychographic reconstruction algorithms deliver phase and absorption contrast images at a resolutionmore » beyond that of the illuminating lens or beam size. Using 5.2-keV X-rays, we have obtained sub-30-nm resolution structural images and similar to 90-nm-resolution fluorescence images of several elements in frozen-hydrated green algae. This combined approach offers a way to study the role of trace elements in their structural context.« less
  • Trace metals play important roles in normal and in disease-causing biological functions. X-ray fluorescence microscopy reveals trace elements with no dependence on binding affinities (unlike with visible light fluorophores) and with improved sensitivity relative to electron probes. However, X-ray fluorescence is not very sensitive for showing the light elements that comprise the majority of cellular material. Here we show that X-ray ptychography can be combined with fluorescence to image both cellular structure and trace element distribution in frozen-hydrated cells at cryogenic temperatures, with high structural and chemical fidelity. Ptychographic reconstruction algorithms deliver phase and absorption contrast images at a resolutionmore » beyond that of the illuminating lens or beam size. Using 5.2-keV X-rays, we have obtained sub–30-nm resolution structural images and ~90-nm–resolution fluorescence images of several elements in frozen-hydrated green algae. This combined approach offers a way to study the role of trace elements in their structural context.« less