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Laboratory Soft X-Ray Cryo Microscopy: Source, System and Bio Applications
KTH, School of Engineering Sciences (SCI), Applied Physics, Biomedical and X-ray Physics.ORCID iD: 0000-0001-7237-6590
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Soft x-ray microscopes routinely perform high-resolution 3D imaging of biological cells in their near-native environment with short exposure times at synchrotron radiation facilities. Some laboratory-sized microscopes are aiming to make this imaging technique more accessible to a wider scientific community. However, these systems have been hampered by source instabilities hindering routine imaging of biological samples with short exposure times.

This Thesis presents work performed on the Stockholm laboratory x-ray microscope. A novel heat control system has been implemented, improving the stability of the laser-produced plasma source. In combination with recent upgrades to the imaging system and an improved cryofixation method, the microscope now has the capability to routinely produce images with 10-second exposure time of cryofixed biological samples. This has allowed for tomographic imaging of cell autophagy and cell-cell interactions. Furthermore, a numerical 3D image formation model is presented as well as a novel reconstruction approach dealing with the limited depth of focus in x-ray microscopes.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2017. , p. 63
Series
TRITA-FYS, ISSN 0280-316X ; 2017:21
Keywords [en]
X-ray microscopy, X-ray optics, Instrumentation, Laser plasma, Cryofixation, Tomography, Tomographic reconstruction, Autophagy, Immune synapse, Physics
National Category
Physical Sciences
Research subject
Physics
Identifiers
URN: urn:nbn:se:kth:diva-206428ISBN: 978-91-7729-369-9 (print)OAI: oai:DiVA.org:kth-206428DiVA, id: diva2:1092920
Public defence
2017-06-02, FD5, Roslagstullsbacken 21, Stockholm, 13:00 (English)
Opponent
Supervisors
Funder
Swedish Research CouncilKnut and Alice Wallenberg Foundation
Note

QC 20170505

Available from: 2017-05-05 Created: 2017-05-04 Last updated: 2017-05-05Bibliographically approved
List of papers
1. High average brightness water window source for short-exposure cryomicroscopy
Open this publication in new window or tab >>High average brightness water window source for short-exposure cryomicroscopy
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2012 (English)In: Optics Letters, ISSN 0146-9592, E-ISSN 1539-4794, Vol. 37, no 21, p. 4425-4427Article in journal (Refereed) Published
Abstract [en]

Laboratory water window cryomicroscopy has recently demonstrated similar image quality as synchrotron-based microscopy but still with much longer exposure times, prohibiting the spread to a wider scientific community. Here we demonstrate high-resolution laboratory water window imaging of cryofrozen cells with 10 s range exposure times. The major improvement is the operation of a lambda = 2.48 nm, 2 kHz liquid nitrogen jet laser plasma source with high spatial and temporal stability at high average brightness >1.5 x 10(12) ph/(s x sr x mu m(2) x line), i.e., close to that of early synchrotrons. Thus, this source enables not only biological x-ray microscopy in the home laboratory but potentially other applications previously only accessible at synchrotron facilities.

Place, publisher, year, edition, pages
Optical Society of America, 2012
Keywords
X-Ray Microscopy
National Category
Other Physics Topics
Identifiers
urn:nbn:se:kth:diva-107087 (URN)10.1364/OL.37.004425 (DOI)000310577700024 ()2-s2.0-84868308552 (Scopus ID)
Funder
Swedish Research CouncilSwedish Foundation for Strategic Research
Note

QC 20121207

Available from: 2012-12-07 Created: 2012-12-06 Last updated: 2017-12-07Bibliographically approved
2. The Stockholm laboratory cryo x-ray microscope: towards cell-cell interaction studies
Open this publication in new window or tab >>The Stockholm laboratory cryo x-ray microscope: towards cell-cell interaction studies
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2013 (English)In: 11th International Conference On X-Ray Microscopy (XRM2012), Institute of Physics (IOP), 2013, p. 012054-Conference paper, Published paper (Refereed)
Abstract [en]

We describe recent improvements in the Stockholm laboratory x-ray microscope and the first experiments aiming towards studies of cell-cell interaction. The shorter exposure time due to a higher brightness laser-plasma source will become of large importance for tomography while the reproducible cryo preparation of few-cell samples is essential for the interaction studies.

Place, publisher, year, edition, pages
Institute of Physics (IOP), 2013
Series
Journal of Physics Conference Series, ISSN 1742-6588 ; 463
National Category
Other Physics Topics
Identifiers
urn:nbn:se:kth:diva-139243 (URN)10.1088/1742-6596/463/1/012054 (DOI)000327949000054 ()2-s2.0-84891288729 (Scopus ID)
Conference
11th International Conference on X-ray Microscopy (XRM), AUG 05-10, 2012, Shanghai, China
Note

QC 20140113

Available from: 2014-01-13 Created: 2014-01-08 Last updated: 2017-05-04Bibliographically approved
3. Stability of liquid-nitrogen-jet laser-plasma targets
Open this publication in new window or tab >>Stability of liquid-nitrogen-jet laser-plasma targets
2015 (English)In: Journal of Applied Physics, ISSN 0021-8979, E-ISSN 1089-7550, Vol. 118, no 17, article id 174902Article in journal (Refereed) Published
Abstract [en]

Microscopic jets of cryogenic substances such as liquid nitrogen are important regenerative high-density targets for high-repetition rate, high-brightness laser-plasma soft x-ray sources. When operated in vacuum such liquid jets exhibit several non-classical instabilities that negatively influence the x-ray source's spatial and temporal stability, yield, and brightness, parameters that all are important for applications such as water-window microscopy. In the present paper, we investigate liquid-nitrogen jets with a flash-illumination imaging system that allows for a quantitative stability analysis with high spatial and temporal resolution. Direct and indirect consequences of evaporation are identified as the key reasons for the observed instabilities. Operating the jets in an approximately 100 mbar ambient atmosphere counteracts the effects of evaporation and produces highly stable liquid nitrogen jets. For operation in vacuum, which is necessary for the laser plasmas, we improve the stability by introducing an external radiative heating element. The method significantly extends the distance from the nozzle that can be used for liquid-jet laser plasmas, which is of importance for high-average-power applications. Finally, we show that laser-plasma operation with the heating-element-stabilized jet shows improved short-term and long-term temporal stability in its water-window x-ray emission.

Place, publisher, year, edition, pages
American Institute of Physics (AIP), 2015
Keywords
Electric heating elements, Electron sources, Evaporation, Fighter aircraft, Image resolution, Jets, Laser beams, Laser produced plasmas, Liquid nitrogen, Liquids, Luminance, Nitrogen, Nitrogen plasma, Plasma stability, Stability, Vacuum applications, X ray apparatus, X ray optics, Ambient atmosphere, High average power, High brightness lasers, High repetition rate, Quantitative stability analysis, Radiative heating, Spatial and temporal resolutions, Temporal stability, Plasma interactions
National Category
Physical Sciences
Identifiers
urn:nbn:se:kth:diva-181201 (URN)10.1063/1.4935143 (DOI)000364584200024 ()2-s2.0-84946811961 (Scopus ID)
Funder
Swedish Research Council
Note

QC 20160205

Available from: 2016-02-05 Created: 2016-01-29 Last updated: 2017-11-30Bibliographically approved
4. Laboratory cryo x-ray microscopy for 3D cell imaging
Open this publication in new window or tab >>Laboratory cryo x-ray microscopy for 3D cell imaging
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(English)Manuscript (preprint) (Other academic)
Keywords
Cryo, Tomography, Compact X-ray Microscopy, Autophagy, Immune synapse
National Category
Natural Sciences
Research subject
Physics
Identifiers
urn:nbn:se:kth:diva-206202 (URN)
Funder
Swedish Research Council
Note

QCR 20170503

Available from: 2017-05-03 Created: 2017-05-03 Last updated: 2017-05-04Bibliographically approved
5. 3D simulation of the image formation in soft x-ray microscopes
Open this publication in new window or tab >>3D simulation of the image formation in soft x-ray microscopes
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2014 (English)In: Optics Express, ISSN 1094-4087, E-ISSN 1094-4087, Vol. 22, no 25, p. 30756-30768Article in journal (Refereed) Published
Abstract [en]

In water-window soft x-ray microscopy the studied object is typically larger than the depth of focus and the sample illumination is often partially coherent. This blurs out-of-focus features and may introduce considerable fringing. Understanding the influence of these phenomena on the image formation is therefore important when interpreting experimental data. Here we present a wave-propagation model operating in 3D for simulating the image formation of thick objects in partially coherent soft x-ray microscopes. The model is compared with present simulation methods as well as with experiments. The results show that our model predicts the image formation of transmission soft x-ray microscopes more accurately than previous models.

Place, publisher, year, edition, pages
Optical Society of America, 2014
Keywords
Wave propagation, X ray microscopes, 3D simulations, Depth of focus, Out-of-focus, Partially coherent, Propagation modeling, Soft X-ray, Soft x-ray microscopy
National Category
Atom and Molecular Physics and Optics
Identifiers
urn:nbn:se:kth:diva-160068 (URN)10.1364/OE.22.030756 (DOI)000346368800068 ()2-s2.0-84919663855 (Scopus ID)
Funder
Swedish Research Council
Note

QC 20150227

QC 20160324

Available from: 2015-02-27 Created: 2015-02-13 Last updated: 2017-12-04Bibliographically approved
6. Tomographic reconstruction in soft x-ray microscopy using focus-stack back-projection
Open this publication in new window or tab >>Tomographic reconstruction in soft x-ray microscopy using focus-stack back-projection
2015 (English)In: Optics Letters, ISSN 0146-9592, E-ISSN 1539-4794, Vol. 40, no 10, p. 2201-2204Article in journal (Refereed) Published
Abstract [en]

Tomographic reconstruction in soft x-ray microscopy is a powerful technique for obtaining high-resolution 3D images of biological samples. However, the depth of focus of such zone-plate-based microscopes is typically shorter than the thickness of many relevant biological objects, challenging the validity of the projection assumption used in conventional reconstruction algorithms. In order to make full use of the soft x-ray microscopes' high resolution, the tomographic reconstruction needs to take the depth of focus into account. Here we present a method to achieve high resolution in the full sample when the depth of focus is short compared to the sample thickness. The method relies on the back-projection of focus-stacked image data from x-ray microscopy. We demonstrate the method on theoretical and experimental data.

Place, publisher, year, edition, pages
Optical Society of America, 2015
Keywords
Image-Formation, Beamline
National Category
Other Physics Topics
Identifiers
urn:nbn:se:kth:diva-169265 (URN)10.1364/OL.40.002201 (DOI)000354708300012 ()2-s2.0-84981357563 (Scopus ID)
Funder
Swedish Research Council
Note

QC 20150615

Available from: 2015-06-15 Created: 2015-06-12 Last updated: 2017-12-04Bibliographically approved

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