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Coherent Diffractive Imaging with X-ray Lasers
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Molecular biophysics.ORCID iD: 0000-0003-1251-0465
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The newly emerging technology of X-ray free-electron lasers (XFELs) has the potential to revolutionise molecular imaging. XFELs generate very intense X-ray pulses and predictions suggest that they may be used for structure determination to atomic resolution even for single molecules. XFELs produce femtosecond pulses that outrun processes of radiation damage and permit the study of structures at room temperature and of structural dynamics.

While the first demonstrations of flash X-ray diffractive imaging (FXI) on biological particles were encouraging, they also revealed technical challenges. In this work we demonstrated how some of these challenges can be overcome. We exemplified, with heterogeneous cell organelles, how tens of thousands of FXI diffraction patterns can be collected, sorted, and analysed in an automatic data processing pipeline. We improved  image resolution and reduced problems with missing data. We validated, described, and deposited the experimental data in the Coherent X-ray Imaging Data Bank.

We demonstrated that aerosol injection can be used to collect FXI data at high hit ratios and with low background. We reduced problems with non-volatile sample contaminants by decreasing aerosol droplet sizes from ~1000 nm to ~150 nm. We achieved this by adapting an electrospray aerosoliser to the Uppsala sample injector. Mie scattering imaging was used as a diagnostic tool to measure positions, sizes, and velocities of individual injected particles.

XFEL experiments generate large amounts of data at high rates. Preparation, execution, and data analysis of these experiments benefits from specialised software. In this work we present new open-source software tools that facilitates prediction, online-monitoring, display, and pre-processing of XFEL diffraction data.

We hope that this work is a valuable contribution in the quest of transitioning FXI from its first experimental demonstration into a technique that fulfills its potentials.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. , 84 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1451
Keyword [en]
coherent diffractive X-ray imaging, lensless imaging, coherent X-ray diffractive imaging, flash diffractive imaging, single particle imaging, aerosol injection, electrospray injection, substrate-free sample delivery, carboxysome, phase retrieval, X-ray diffraction software, X-ray free-electron laser, XFEL, FEL, CXI, CDI, CXDI, FXI
National Category
Biophysics Atom and Molecular Physics and Optics Structural Biology
Identifiers
URN: urn:nbn:se:uu:diva-306609ISBN: 978-91-554-9748-4OAI: oai:DiVA.org:uu-306609DiVA: diva2:1043706
Public defence
2016-12-19, E10:1307-E10:1309, Biomedical Centre, Husargatan 3, Uppsala, 09:30 (English)
Opponent
Supervisors
Available from: 2016-11-29 Created: 2016-10-30 Last updated: 2016-11-29
List of papers
1. High-throughput imaging of heterogeneous cell organelles with an X-ray laser
Open this publication in new window or tab >>High-throughput imaging of heterogeneous cell organelles with an X-ray laser
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2014 (English)In: Nature Photonics, ISSN 1749-4885, E-ISSN 1749-4893, Vol. 8, no 12, 943-949 p.Article in journal (Refereed) Published
Abstract [en]

We overcome two of the most daunting challenges in single-particle diffractive imaging: collecting many high-quality diffraction patterns on a small amount of sample and separating components from mixed samples. We demonstrate this on carboxysomes, which are polyhedral cell organelles that vary in size and facilitate up to 40% of Earth's carbon fixation. A new aerosol sample-injector allowed us to record 70,000 low-noise diffraction patterns in 12 min with the Linac Coherent Light Source running at 120 Hz. We separate different structures directly from the diffraction data and show that the size distribution is preserved during sample delivery. We automate phase retrieval and avoid reconstruction artefacts caused by missing modes. We attain the highest-resolution reconstructions on the smallest single biological objects imaged with an X-ray laser to date. These advances lay the foundations for accurate, high-throughput structure determination by flash-diffractive imaging and offer a means to study structure and structural heterogeneity in biology and elsewhere.

National Category
Structural Biology
Identifiers
urn:nbn:se:uu:diva-237619 (URN)10.1038/nphoton.2014.270 (DOI)000345818600014 ()
Available from: 2014-12-03 Created: 2014-12-03 Last updated: 2016-10-31Bibliographically approved
2. A data set from flash X-ray imaging of carboxysomes
Open this publication in new window or tab >>A data set from flash X-ray imaging of carboxysomes
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2016 (English)In: Scientific Data, ISSN 1012-0602, E-ISSN 2052-4463, Vol. 3, 160061Article in journal (Refereed) Published
Abstract [en]

Ultra-intense femtosecond X-ray pulses from X-ray lasers permit structural studies on single particles and biomolecules without crystals. We present a large data set on inherently heterogeneous, polyhedral carboxysome particles. Carboxysomes are cell organelles that vary in size and facilitate up to 40% of Earth’s carbon fixation by cyanobacteria and certain proteobacteria. Variation in size hinders crystallization. Carboxysomes appear icosahedral in the electron microscope. A protein shell encapsulates a large number of Rubisco molecules in paracrystalline arrays inside the organelle. We used carboxysomes with a mean diameter of 115±26 nm from Halothiobacillus neapolitanus. A new aerosol sample-injector allowed us to record 70,000 low-noise diffraction patterns in 12 min. Every diffraction pattern is a unique structure measurement and high-throughput imaging allows sampling the space of structural variability. The different structures can be separated and phased directly from the diffraction data and open a way for accurate, high-throughput studies on structures and structural heterogeneity in biology and elsewhere.

National Category
Biophysics
Identifiers
urn:nbn:se:uu:diva-300202 (URN)10.1038/sdata.2016.61 (DOI)
External cooperation:
Note

Data Descriptor

Available from: 2016-08-05 Created: 2016-08-05 Last updated: 2016-10-31Bibliographically approved
3. Condor: a simulation tool for flash X-ray imaging
Open this publication in new window or tab >>Condor: a simulation tool for flash X-ray imaging
2016 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 49, 1356-1362 p.Article, review/survey (Refereed) Published
Abstract [en]

Flash X-ray imaging has the potential to determine structures down to molecular resolution without the need for crystallization. The ability to accurately predict the diffraction signal and to identify the optimal experimental configuration within the limits of the instrument is important for successful data collection. This article introduces Condor, an open-source simulation tool to predict X-ray far-field scattering amplitudes of isolated particles for customized experimental designs and samples, which the user defines by an atomic or a refractive index model. The software enables researchers to test whether their envisaged imaging experiment is feasible, and to optimize critical parameters for reaching the best possible result. It also aims to support researchers who intend to create or advance reconstruction algorithms by simulating realistic test data. Condor is designed to be easy to use and can be either installed as a Python package or used from its web interface (http://lmb.icm.uu.se/condor). X-ray free-electron lasers have high running costs and beam time at these facilities is precious. Data quality can be substantially improved by using simulations to guide the experimental design and simplify data analysis.

Keyword
femtosecond coherent diffractive imaging, X-ray free-electron lasers, simulation, single-particle imaging, computer programs
National Category
Biophysics
Identifiers
urn:nbn:se:uu:diva-300211 (URN)10.1107/S1600576716009213 (DOI)000382755900027 ()
Available from: 2016-08-05 Created: 2016-08-05 Last updated: 2016-10-31Bibliographically approved
4. Hummingbird: monitoring and analyzing flash X-ray imaging experiments in real time
Open this publication in new window or tab >>Hummingbird: monitoring and analyzing flash X-ray imaging experiments in real time
2016 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 49, 1042-1047 p.Article in journal (Refereed) Published
National Category
Biophysics Software Engineering
Identifiers
urn:nbn:se:uu:diva-287197 (URN)10.1107/S1600576716005926 (DOI)000377020600036 ()
Projects
eSSENCE
Available from: 2016-04-18 Created: 2016-04-22 Last updated: 2016-10-31Bibliographically approved
5. Cheetah: software for high-throughput reduction and analysis of serial femtosecond X-ray diffraction data
Open this publication in new window or tab >>Cheetah: software for high-throughput reduction and analysis of serial femtosecond X-ray diffraction data
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2014 (English)In: Journal of applied crystallography, ISSN 0021-8898, E-ISSN 1600-5767, Vol. 47, 1118-1131 p.Article in journal (Refereed) Published
Abstract [en]

The emerging technique of serial X-ray diffraction, in which diffraction data are collected from samples flowing across a pulsed X-ray source at repetition rates of 100 Hz or higher, has necessitated the development of new software in order to handle the large data volumes produced. Sorting of data according to different criteria and rapid filtering of events to retain only diffraction patterns of interest results in significant reductions in data volume, thereby simplifying subsequent data analysis and management tasks. Meanwhile the generation of reduced data in the form of virtual powder patterns, radial stacks, histograms and other meta data creates data set summaries for analysis and overall experiment evaluation. Rapid data reduction early in the analysis pipeline is proving to be an essential first step in serial imaging experiments, prompting the authors to make the tool described in this article available to the general community. Originally developed for experiments at X-ray free-electron lasers, the software is based on a modular facility-independent library to promote portability between different experiments and is available under version 3 or later of the GNU General Public License.

National Category
Natural Sciences
Identifiers
urn:nbn:se:uu:diva-228011 (URN)10.1107/S1600576714007626 (DOI)000336738500032 ()
Available from: 2014-07-03 Created: 2014-07-02 Last updated: 2016-10-31Bibliographically approved

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