Coherent X-ray diffractive imaging on the singel-cell-level of microbial samples

254 Pages·2015·14.79 MB·English

Vol.
14
Since its fi rst experimental demonstration in 1999, Coherent X-Ray Diffractive Göttingen Series in
Imaging has become one of the most promising high resolution X-Ray imaging
techniques using coherent radiation produced by brilliant synchrotron storage X-ray Physics
rings. The ability to directly invert diffraction data with the help of advanced algo-
rithms has paved the way for microscopic investigations and wave-fi eld analyses
on the spatial scale of nanometres without the need for ineffi cient imaging lenses.
X-Ray phase contrast which is a measure of the electron density is an important
contrast mode of soft biological specimens. For the case of many dominant ele-
ments of soft biological matter, the electron density can be converted into an Robin Niklas Wilke
effective mass density offering a unique quantitative information channel which
Coherent X-ray diffractive imaging
may shed light on important questions such as DNA compaction in the bacterial
nucleoid through ‚weighing with light‘.
on the single-cell-level of microbial samples
In this work X-Ray phase contrast maps have been obtained from different bio-
logical samples by exploring different methods. In particular, the techniques Pty-
chography and Waveguide-Holographic-Imaging have been used to obtain two- Ptychography, tomography, nano-diffraction and
dimensional and three-dimensional mass density maps on the single-cell-level of waveguide-imaging
freeze-dried cells of the bacteria Deinococcus radiodurans, Bacillus subtilis and
Bacillus thuringiensis allowing, for instance, to estimate the dry weight of the
bacterial genome in a near native state. On top of this, reciprocal space informa-
g
tion from coherent small angle X-Ray scattering (cellular Nano-Diffraction) of the n
gi
fi ne structure of the bacterial cells has been recorded in a synergistic manner and a
m
has been analysed down to a resolution of about 2.3/nm exceeding current limits of -i
of direct imaging approaches. Furthermore, the dynamic range of present detector el de
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ISBN 978-3-86395-190-0 Universitätsverlag Göttingen Universitätsverlag Göttingen
ISSN 2191-9860N:

Robin Niklas Wilke
Coherent X-ray diffractive imaging on the single-cell-level of microbial samples:
ptychography, tomography, nano-diffraction and waveguide-imaging

This work is licensed under a Creative Commons
Attribution-ShareAlike 4.0 International License.
Published in 2015 by Universitätsverlag Göttingen
as volume 14 in the series „Göttingen series in X-ray physics“
Robin Niklas Wilke

Coherent X-ray
diffractive imaging
on the single-cell-level
of microbial samples:

ptychography, tomography,
nano-diffraction and
waveguide-imaging

Göttingen series in X-ray physics
Volume 14

Universitätsverlag Göttingen
2015
Bibliographische Information der Deutschen Nationalbibliothek
Die Deutsche Nationalbibliothek verzeichnet diese Publikation in der Deutschen
Nationalbibliographie; detaillierte bibliographische Daten sind im Internet über
<http://dnb.ddb.de> abrufbar.

This work has been financially supported by the DFG Collaborative Research Center
SFB 755 “Nanoscale Photonic Imaging”
Address of the Author
Dr. Robin Niklas Wilke
e-mail: [email protected]

Dissertation
for the award of the degree
“Doctor rerum naturalium”
of the Georg-August-Universität Göttingen
within the doctoral program PROPHYS
of the Georg-August University School of Science (GAUSS)

Thesis Committee:
Prof. Dr. Tim Salditt
Prof. Dr. Claus Ropers

This work is protected by German Intellectual Property Right Law.
It is also available as an Open Access version through the publisher’s homepage and
the Online Catalogue of the State and University Library of Goettingen
(http://www.sub.uni-goettingen.de).
The conditions of the license terms of the online version apply.

Layout: Dr. Robin Niklas Wilke
Cover: Jutta Pabst
Cover image: Dr. Robin Niklas Wilke

© 2015 Universitätsverlag Göttingen
http://univerlag.uni-goettingen.de
ISBN: 978-3-86395-190-0
ISSN: 2191-9860
Preface of the series editor
s

The Göttingen series in x-ray physics is intended as a collection of research
monographs in x-ray science, carried out at the Institute for X-ray Physics at the
Georg-August-Universität in Göttingen, and in the framework of its related research
networks and collaborations.
It covers topics ranging from x-ray microscopy, nano-focusing, wave propagation,
image reconstruction, tomography, short x-ray pulses to applications of nanoscale x-
ray imaging and biomolecular structure analysis.
In most but not all cases, the contributions are based on Ph.D. dissertations. The
individual monographs should be enhanced by putting them in the context of related
work, often based on a common long term research strategy, and funded by the same
research networks. We hope that the series will also help to enhance the visibility of
the research carried out here and help others in the field to advance similar projects.

Prof. Dr. Sarah Köster
Prof. Dr. Tim Salditt
Editors
Göttingen June 2014

Contents
Nomenclature v
Introduction 1
Theory 7
1.1 Scalar Wave-Fields . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
1.1.1 Propagation of Scalar Wave-Fields . . . . . . . . . . . . . . 7
1.1.2 Remarks on the Numerical Propagation of Scalar Wave-Fields 12
1.1.3 Partial Coherence . . . . . . . . . . . . . . . . . . . . . . . 15
1.2 X-Rays in Matter . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
1.2.1 Scattering Theory . . . . . . . . . . . . . . . . . . . . . . . 20
1.2.2 The Projection Approximation . . . . . . . . . . . . . . . . 24
1.2.3 Dose vs. Resolution . . . . . . . . . . . . . . . . . . . . . . 30
1.3 Imaging with X-Rays . . . . . . . . . . . . . . . . . . . . . . . . . . 34
1.3.1 Direct Methods . . . . . . . . . . . . . . . . . . . . . . . . . 34
1.3.2 The Phase Problem . . . . . . . . . . . . . . . . . . . . . . 35
1.3.3 Coherent Di(cid:27)ractive Imaging . . . . . . . . . . . . . . . . . 36
1.3.4 Sampling in CDI . . . . . . . . . . . . . . . . . . . . . . . . 37
1.3.5 Phase Retrieval Algorithms for Isolated Objects . . . . . . 39
1.3.6 Ptychography . . . . . . . . . . . . . . . . . . . . . . . . . . 40
1.3.7 The ePIE algorithm . . . . . . . . . . . . . . . . . . . . . . 41
1.3.8 Tomography . . . . . . . . . . . . . . . . . . . . . . . . . . 43
1.3.9 The Fourier Slice Theorem . . . . . . . . . . . . . . . . . . 43
1.3.10 Filtered Back Projection Algorithm . . . . . . . . . . . . . 43
Bacterial Samples 47
2.1 Deinococcus radiodurans . . . . . . . . . . . . . . . . . . . . . . . . 47
2.2 Bacillus subtilis & Bacillus thuringiensis . . . . . . . . . . . . . . . 51
2.3 Sample Preparations . . . . . . . . . . . . . . . . . . . . . . . . . . 55
2.3.1 Cell Culture of D. radiodurans . . . . . . . . . . . . . . . . 55
2.3.2 Cell Culture of B. subtilis & B. thuringiensis . . . . . . . . 57
2.3.3 PreparationofUltra-thinSectionsforTransmissionElectron
Microscopy . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
2.3.4 Vitri(cid:28)cation . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
2.3.5 Lyophilisation. . . . . . . . . . . . . . . . . . . . . . . . . . 63
Experiments 69
3.1 Ptychographic Imaging and Nano-Di(cid:27)raction of
D. radiodurans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69
3.1.1 Experimental Set-up . . . . . . . . . . . . . . . . . . . . . . 70
3.1.2 Sample Preparation . . . . . . . . . . . . . . . . . . . . . . 71
3.1.3 Details and Terminology: Cellular Nano-Di(cid:27)raction . . . . 71
3.1.4 Results: Cellular Nano-Di(cid:27)raction . . . . . . . . . . . . . . 72
3.1.5 Details and Theory: Ptychographic Imaging . . . . . . . . . 79
ii Contents
3.1.6 Details: Tomographic Reconstruction . . . . . . . . . . . . 82
3.1.7 Results: PtychographicImagingoftheResolutionChartand
Wave-Field Analysis . . . . . . . . . . . . . . . . . . . . . . 83
3.1.8 Resolution vs. Coherence . . . . . . . . . . . . . . . . . . . 94
3.1.9 Results: Ptychographic Imaging Towards a 3D Representa-
tion of D. radiodurans Cells . . . . . . . . . . . . . . . . . . 97
3.1.10 Results: Tomo-Ptychographic Imaging of D. radiodurans cells100
3.1.11 Results: TEM Analysis of D. radiodurans . . . . . . . . . . 103
3.1.12 Discussion & Conclusion . . . . . . . . . . . . . . . . . . . 107
3.2 Scanning Hard X-ray Microscopy Imaging Modalities for Geobio-
logical Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 110
3.2.1 Sample Origin & Sample Preparations . . . . . . . . . . . . 111
3.2.2 Details: Scanning X-Ray Fluorescence and Scanning
Transmission X-Ray Experiments . . . . . . . . . . . . . . . 111
3.2.3 Details: ScanningTransmissionX-RayandX-RayPtychog-
raphy Experiment . . . . . . . . . . . . . . . . . . . . . . . 113
3.2.4 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119
3.2.5 Details & Results: SEM-EDX Analysis. . . . . . . . . . . . 126
3.2.6 Discussion & Conclusion . . . . . . . . . . . . . . . . . . . . 129
3.3 STCS in High-Resolution X-Ray Ptychography Using KB Focusing 132
3.3.1 Experimental Set-up . . . . . . . . . . . . . . . . . . . . . . 133
3.3.2 Details: Analysis . . . . . . . . . . . . . . . . . . . . . . . . 136
3.3.3 Results & Discussion . . . . . . . . . . . . . . . . . . . . . . 138
3.3.4 Summary & Conclusion . . . . . . . . . . . . . . . . . . . . 142
3.4 An STCS Application in High-Resolution X-Ray Ptychography . . 144
3.4.1 Experimental Set-up . . . . . . . . . . . . . . . . . . . . . . 145
3.4.2 Details: STCS Preparations . . . . . . . . . . . . . . . . . . 147
3.4.3 Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147
3.4.4 Details: Data Recording & Data Treatment . . . . . . . . . 148
3.4.5 Results: Ptychographic Reconstruction of KB Wave-Fields 149
3.4.6 Results: Ptychographic Imaging of Nanowires . . . . . . . . 153
3.4.7 Discussion, Conclusion & Outlook . . . . . . . . . . . . . . 155
3.5 Waveguide-Based Imaging of B. thuringiensis and B. subtilis . . . 161
3.5.1 Experimental Set-up . . . . . . . . . . . . . . . . . . . . . . 161
3.5.2 Sample Preparation . . . . . . . . . . . . . . . . . . . . . . 162
3.5.3 Details: Theory, Data Recording & Data Treatment . . . . 164
3.5.4 Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 172
3.5.5 Conclusion, Discussion & Outlook . . . . . . . . . . . . . . 185
4 Summary, Conclusion & Outlook 187
A Fourier Transform Terminology 191
B Culture Media 192