Microscopy Beamline Research Articles

Placental Element Content Assessed via Synchrotron-Based X-ray Fluorescence Microscopy Identifies Low Molybdenum Concentrations in Foetal Growth Restriction, Postdate Delivery and Stillbirth.

Placental health and foetal development are dependent upon element homeostasis. Analytical techniques such as mass spectroscopy can provide quantitative data on element concentrations in placental tissue but do not show spatial distribution or co-localisation of elements that may affect placental function. The present study used synchrotron-based X-ray fluorescence microscopy to elucidate element content and distribution in healthy and pathological placental tissue. The X-ray fluorescence microscopy (XFM) beamline at the Australian Synchrotron was used to image trace metal content of 19 placental sections from healthy term (n = 5, 37-39 weeks), foetal growth-restricted (n = 3, <32 weeks, birth weight <3rd centile), postdate (n = 7, >41 completed weeks), and stillbirth-complicated pregnancies (n = 4, 37-40 weeks). Samples were cryo-sectioned and freeze-dried. The concentration and distribution of fourteen elements were detected in all samples: arsenic, bromine, calcium, chlorine, copper, iron, molybdenum, phosphorous, potassium, rubidium, selenium, strontium, sulphur, and zinc. The elements zinc, calcium, phosphorous, and strontium were significantly increased in stillbirth placental tissue in comparison to healthy-term controls. Strontium, zinc, and calcium were found to co-localise in stillbirth tissue samples, and calcium and strontium concentrations were correlated in all placental groups. Molybdenum was significantly decreased in stillbirth, foetal growth-restricted, and postdate placental tissue in comparison to healthy-term samples (p < 0.0001). Synchrotron-based XFM reveals elemental distribution within biological samples such as the placenta, allowing for the co-localisation of metal deposits that may have a pathological role. Our pilot study further indicates low concentrations of placental molybdenum in pregnancies complicated by foetal growth restriction, postdate delivery, and stillbirth.

Characterization of iLGADs using soft X-rays

Experiments at synchrotron radiation sources and X-ray Free-Electron Lasers in the soft X-ray energy range (250 eV–2 keV) stand to benefit from the adaptation of the hybrid silicon detector technology for low energy photons. Inverse Low Gain Avalanche Diode (iLGAD) sensors provide an internal gain, enhancing the signal-to-noise ratio and allowing single photon detection below 1 keV using hybrid detectors. In addition, an optimization of the entrance window of these sensors enhances their quantum efficiency (QE).In this work, the QE and the gain of a batch of different iLGAD diodes with optimized entrance windows were characterized using soft X-rays at the Surface/Interface:Microscopy beamline of the Swiss Light Source synchrotron. Above 250 eV, the QE is larger than 55% for all sensor variations, while the charge collection efficiency is close to 100%. The average gain depends on the gain layer design of the iLGADs and increases with photon energy. A fitting procedure is introduced to extract the multiplication factor as a function of the absorption depth of X-ray photons inside the sensors. In particular, the multiplication factors for electron- and hole-triggered avalanches are estimated, corresponding to photon absorption beyond or before the gain layer, respectively.

Advances in sparse dynamic scanning in spectromicroscopy through compressive sensing.

Scanning microscopies and spectroscopies like X-ray Fluorescence (XRF), Scanning Transmission X-ray Microscopy (STXM), and Ptychography are of very high scientific importance as they can be employed in several research fields. Methodology and technology advances aim at analysing larger samples at better resolutions, improved sensitivities and higher acquisition speeds. The frontiers of those advances are in detectors, radiation sources, motors, but also in acquisition and analysis software together with general methodology improvements. We have recently introduced and fully implemented an intelligent scanning methodology based on compressive sensing, on a soft X-ray microscopy beamline. This demonstrated sparse low energy XRF scanning of dynamically chosen regions of interest in combination with STXM, yielding spectroimaging data in the megapixel-range and in shorter timeframes than were previously not feasible. This research has been further developed and has been applied to scientific applications in biology. The developments are mostly in the dynamic triggering decisional mechanism in order to incorporate modern Machine Learning (ML) but also in the suitable integration of the method in the control system, making it available for other beamlines and imaging techniques. On the applications front, the method was previously successfully used on different samples, from lung and ovarian human tissues to plant root sections. This manuscript introduces the latest methodology advances and demonstrates their applications in life and environmental sciences. Lastly, it highlights the auxiliary development of a mobile application, designed to assist the user in the selection of specific regions of interest in an easy way.

Size-Dependent Penetration of Nanoparticles in Tumor Spheroids: A Multidimensional and Quantitative Study of Transcellular and Paracellular Pathways.

Tumor penetration of nanoparticles is crucial in nanomedicine, but the mechanisms of tumor penetration are poorly understood. This work presents a multidimensional, quantitative approach to investigate the tissue penetration behavior of nanoparticles, with focuses on the particle size effect on penetration pathways, in an MDA-MB-231 tumor spheroid model using a combination of spectrometry, microscopy, and synchrotron beamline techniques. Quasi-spherical gold nanoparticles of different sizes are synthesized and incubated with 2D and 3D MDA-MB-231 cells and spheroids with or without an energy-dependent cell uptake inhibitor. The distribution and penetration pathways of nanoparticles in spheroids are visualized and quantified by inductively coupled plasma mass spectrometry, two-photon microscopy, and synchrotron X-ray fluorescence microscopy. The results reveal that 15nm nanoparticles penetrate spheroids mainly through an energy-independent transcellular pathway, while 60nm nanoparticles penetrate primarily through an energy-dependent transcellular pathway. Meanwhile, 22nm nanoparticles penetrate through both transcellular and paracellular pathways and they demonstrate the greatest penetration ability in comparison to other two sizes. The multidimensional analytical methodology developed through this work offers a generalizable approach to quantitatively study the tissue penetration of nanoparticles, and the results provide important insights into the designs of nanoparticles with high accumulation at a target site.

Shielding design aspects of SR in 3 GeV ILSF

Iranian Light Source Facility (ILSF) is an under-construction synchrotron radiation accelerator consisting of a 150 MeV linac, a booster synchrotron operating from 150 MeV to 3 GeV, and a 3 GeV storage ring that stores a maximum of 400 mA current of electrons. As the stored beam circulates, a fraction of the beam is lost due to interactions with gas molecules, interactions among beam particles, and orbital bending, which produce radiation. The bulk shielding calculation for the ILSF and the input parameters used for this analysis are discussed in this paper. The potential of skyshine neutrons to cause radiation hazards is investigated as well. Moreover, the design and shielding simulation using the FLUKA Monte Carlo code is presented for the linac beam stop and primary and scattered gas bremsstrahlung for the first optics enclosure of the ILSF spectro microscopy beamline. Our designed radiation shielding system guarantees that the annual dose in all areas around the ILSF machine does not exceed the dose limit of 1 mSv.

Scanning photoelectron spectromicroscopy: From static to operando studies of functional materials

The scanning photoelectron microscope (SPEM), developed more than 30 years ago, has undergone numerous technical developments, providing an incredibly vast kind of feasible sample environments, which span from the traditional high spatial resolution core level based chemical analysis to insitu and operando complex experiments, including also electrochemical setups and operational electronic devices at various temperatures. Another important step ahead is overcoming the so-called pressure gap for operando studies, recently extended to near ambient values by building special environmental cells. Using recent results of conventional and unconventional experiments, obtained with SPEM at the ESCA Microscopy beamline at Elettra Sincrotrone Trieste the present review demonstrates the current potential of this type of photoelectron spectromicroscopy to explore the interfacial properties of functional materials with high spatial resolution.

MAXPEEM: a spectromicroscopy beamline at MAX IV laboratory.

MAXPEEM, a dedicated photoemission electron microscopy beamline at MAX IV Laboratory, houses a state-of-the-art aberration-corrected spectroscopic photoemission and low-energy electron microscope (AC-SPELEEM). This powerful instrument offers a wide range of complementary techniques providing structural, chemical and magnetic sensitivities with a single-digit nanometre spatial resolution. The beamline can deliver a high photon flux of≥1015 photons s-1 (0.1% bandwidth)-1 in the range 30-1200 eV with full control of the polarization from an elliptically polarized undulator. The microscope has several features which make it unique from similar instruments. The X-rays from the synchrotron pass through the first beam separator and impinge the surface at normal incidence. The microscope is equipped with an energy analyzer and an aberration corrector which improves both the resolution and the transmission compared with standard microscopes. A new fiber-coupled CMOS camera features an improved modulation transfer function, dynamic range and signal-to-noise ratio compared with the traditional MCP-CCD detection system.

Development of 150 Mpixel lens-coupled X-ray imaging detectors equipped with diffusion-free transparent scintillators based on an analytical optimization approach

X-ray microscopy experiments at synchrotron radiation facilities generally require X-ray imaging detectors with high spatial resolution and a wide field of view. For such purposes, lens-coupled X-ray imaging detectors are widely used. The SPring-8 X-ray microscopy beamlines require detectors that satisfy a 200 nm spatial resolution and over 50 mm field of view. We have carried out a systematic optical design based on an analytical optimization approach and elucidated that a set of five detector systems is able to meet these requirements. Their objective lenses have magnifications of 20x, 7x, 5.2x, 3.5x, and 1x. All the systems are designed to be equipped with 150 Mpixel CMOS image sensors. To verify our optimization approach, we have built a detector system with a magnification of 3.5x. The detector has successfully resolved 1.2 µm line-and-space patterns for the entire field of view, which is close to the optical diffraction-limited resolving power of 1.0 µm line-and-space patterns.

Water-cooled magnetic coupling drive attenuator design for wiggler light source in the TPS 31A beamline.

An attenuator is generally used to decrease the power of an X-ray beam and prevent damage to detector sensors and other optical components. Therefore, attenuators are designed using foil or gas to absorb light source power. In this project, a large aperture and a water-cooling attenuator system are construed for the TPS 31A Projection X-ray Microscope and Transmission X-ray Microscope beamline. The source size of the wiggler is 300 µm × 7 µm on TPS 31A. The X-ray beam size at the sample position is 50 mm × 20 mm, located 49.5 m from the source. The light emission power is 1000 W in white-beam operation mode. The attenuator is needed to absorb energy for the light source and it has 12 foil carriers. The absorption foil size is 56 mm × 46 mm for the beam size across different beamline operation modes, and the cooling capacity is greater than 1000 W. This study applies a magnetic coupling-type attenuator system with foil carrier cooling carried out by the side chamber walls without the feedthrough having water enter the chamber to solve the thermal dissipation issue.

Near ambient pressure photoelectron spectro-microscopy: from gas–solid interface to operando devices

Near ambient pressure scanning photoelectron microscopy adds to the widely used photoemission spectroscopy and its chemically selective capability two key features: (1) the possibility to chemically analyse samples in a more realistic gas pressure condition and (2) the capability to investigate a system at the relevant spatial scale. To achieve these goals the approach developed at the ESCA Microscopy beamline at the Elettra Synchrotron facility combines the submicron lateral resolution of a Scanning Photoelectron Microscope with a custom designed Near Ambient Pressure Cell where a gas pressure up to 0.1 mbar can be achieved. In this manuscript a review of experiments performed with this unique setup will be presented to illustrate its potentiality in both fundamental and applicative research such as the oxidation reactivity and gas sensitivity of metal oxides and semiconductors. In particular, the capability to perform operando experiments with this setup opens the possibility to study operating devices and to properly address the real nature of the studied systems, because if microscopy and spectroscopy are simultaneously combined in a single technique it can yield to more conclusive results.

Zone plate-based full-field transmission X-ray microscopy beamline design at nearly diffraction-limited synchrotron radiation facility

With the development of full-field transmission X-ray microscopy (TXM) for research in biology, life science, material science and physics, zone plate-based full-field TXM is becoming a powerful tool for obtaining quantitative internal structural and chemical information at the nanoscale when combined with spectroscopic imaging. For the TXM beamline design, the best choice is to match the phase space needed by the zone plate from the source point. However, for the nearly diffraction-limited synchrotron radiation sources, it is not easy to fully match the needed phase space due to its low-emittance. Using redshift radiation from an undulator, under consideration of experimental requirements and the heat load effect, this paper describes the design of TXM beamline at the nearly diffraction-limited High Energy Photon Source (HEPS). In this design, not only sufficient illumination angle can be acquired, but also a nearly annular illumination can be obtained to match the illumination required for zone plate imaging. Furthermore, about two times photon flux can be obtained at source point and the photons can be fully used in this design using redshift undulator radiation. The presented design can also be informative for the same kind of beamline design at other nearly diffraction-limited synchrotron photon sources.

Synchrotron X-ray fluorescence microscopy-enabled elemental mapping illuminates the 'battle for nutrients' between plant and pathogen.

Metal homeostasis is integral to normal plant growth and development. During plant–pathogen interactions, the host and pathogen compete for the same nutrients, potentially impacting nutritional homeostasis. Our knowledge of outcome of the interaction in terms of metal homeostasis is still limited. Here, we employed the X-ray fluorescence microscopy (XFM) beamline at the Australian Synchrotron to visualize and analyse the fate of nutrients in wheat leaves infected with Pyrenophora tritici-repentis, a necrotrophic fungal pathogen. We sought to (i) evaluate the utility of XFM for sub-micron mapping of essential mineral nutrients and (ii) examine the spatiotemporal impact of a pathogen on nutrient distribution in leaves. XFM maps of K, Ca, Fe, Cu, Mn, and Zn revealed substantial hyperaccumulation within, and depletion around, the infected region relative to uninfected control samples. Fungal mycelia were visualized as thread-like structures in the Cu and Zn maps. The hyperaccumulation of Mn in the lesion and localized depletion in asymptomatic tissue surrounding the lesion was unexpected. Similarly, Ca accumulated at the periphery of the symptomatic region and as microaccumulations aligning with fungal mycelia. Collectively, our results highlight that XFM imaging provides the capability for high-resolution mapping of elements to probe nutrient distribution in hydrated diseased leaves in situ.

The XFM beamline at the Australian Synchrotron.

The X-ray fluorescence microscopy (XFM) beamline is an in-vacuum undulator-based X-ray fluorescence (XRF) microprobe beamline at the 3 GeV Australian Synchrotron. The beamline delivers hard X-rays in the 4-27 keV energy range, permitting K emission to Cd and L and M emission for all other heavier elements. With a practical low-energy detection cut-off of approximately 1.5 keV, low-Z detection is constrained to Si, with Al detectable under favourable circumstances. The beamline has two scanning stations: a Kirkpatrick-Baez mirror microprobe, which produces a focal spot of 2 µm × 2 µm FWHM, and a large-area scanning `milliprobe', which has the beam size defined by slits. Energy-dispersive detector systems include the Maia 384, Vortex-EM and Vortex-ME3 for XRF measurement, and the EIGER2 X 1 Mpixel array detector for scanning X-ray diffraction microscopy measurements. The beamline uses event-mode data acquisition that eliminates detector system time overheads, and motion control overheads are significantly reduced through the application of an efficient raster scanning algorithm. The minimal overheads, in conjunction with short dwell times per pixel, have allowed XFM to establish techniques such as full spectroscopic XANES fluorescence imaging, XRF tomography, fly scanning ptychography and high-definition XRF imaging over large areas. XFM provides diverse analysis capabilities in the fields of medicine, biology, geology, materials science and cultural heritage. This paper discusses the beamline status, scientific showcases and future upgrades.

Impact of lossy compression of X-ray projections onto reconstructed tomographic slices

Modern detectors used at synchrotron tomographic microscopy beamlines typically have sensors with more than 4-5 mega-pixels and are capable of acquiring 100-1000 frames per second at full frame. As a consequence, a data rate of a few TB per day can easily be exceeded, reaching peaks of a few tens of TB per day for time-resolved tomographic experiments. This data needs to be post-processed, analysed, stored and possibly transferred, imposing a significant burden onto the IT infrastructure. Compression of tomographic data, as routinely done for diffraction experiments, is therefore highly desirable. This study considers a set of representative datasets and investigates the effect of lossy compression of the original X-ray projections onto the final tomographic reconstructions. It demonstrates that a compression factor of at least three to four times does not generally impact the reconstruction quality. Potentially, compression with this factor could therefore be used in a transparent way to the user community, for instance, prior to data archiving. Higher factors (six to eight times) can be achieved for tomographic volumes with a high signal-to-noise ratio as it is the case for phase-retrieved datasets. Although a relationship between the dataset signal-to-noise ratio and a safe compression factor exists, this is not simple and, even considering additional dataset characteristics such as image entropy and high-frequency content variation, the automatic optimization of the compression factor for each single dataset, beyond the conservative factor of three to four, is not straightforward.

Real-time reconstruction and visualisation towards dynamic feedback control during time-resolved tomography experiments at TOMCAT

Tomographic X-ray microscopy beamlines at synchrotron light sources worldwide have pushed the achievable time-resolution for dynamic 3-dimensional structural investigations down to a fraction of a second, allowing the study of quickly evolving systems. The large data rates involved impose heavy demands on computational resources, making it difficult to readily process and interrogate the resulting volumes. The data acquisition is thus performed essentially blindly. Such a sequential process makes it hard to notice problems with the measurement protocol or sample conditions, potentially rendering the acquired data unusable, and it keeps the user from optimizing the experimental parameters of the imaging task at hand. We present an efficient approach to address this issue based on the real-time reconstruction, visualisation and on-the-fly analysis of a small number of arbitrarily oriented slices. This solution, requiring only a single additional computing workstation, has been implemented at the TOMCAT beamline of the Swiss Light Source. The system is able to process multiple sets of slices per second, thus pushing the reconstruction throughput on the same level as the data acquisition. This enables the monitoring of dynamic processes as they occur and represents the next crucial step towards adaptive feedback control of time-resolved in situ tomographic experiments.

An optimized 19F (p, αγ)16O elemental mapping facility

A new target stage that allows a close positioning of a large 5in × 5in NaI(Tl) detector was implemented at the 45° beam-line end-station at the Centre for Ion Beam Applications (CIBA). Here we present preliminary results demonstrating the improvement in the F detection limit available in the new geometry. The reaction 19F(p, αγ)16O was used to map fluorine in untreated and fluorine/laser treated areas on human tooth samples.Proton Induced Gamma-Ray Emission (PIGE) was used in combination with Proton Induced X-Ray Emission (PIXE) and Rutherford Backscattering Spectrometry (RBS) for quantitative elemental mapping of fluorine. A 2 MeV proton beam focused to 5~10 μm spot size with a beam current of 0.5–1 nA was used along with a new in-vacuum piezo-driven three-axis target-stage at the nuclear microscopy beamline end-station. The new stage, which replaces the manual target positioning system, optimizes the solid angle subtended by factor of 13.

Full-field X-ray Imaging, a Workhorse Microscopy Beamline at NSLS II for Material Science Researches

Full-field X-ray Imaging, a Workhorse Microscopy Beamline at NSLS II for Material Science Researches

(Invited) A New Transmission X-Ray Microscope Beamline at the NSLS-II

The Transmission X-ray Microscope (TXM) has proven to be a very useful tool for energy storage systems research. It has been used to study, in 2D and 3D, the morphological and chemical state changes in energy storage systems with several tens of nanometer spatial resolution. Up until recently, however, the typical data acquisition time for a single energy 3D tomography data set is usually between 20-240 minutes at a synchrotron. In this talk, we present a new TXM instrument at the Full-field X-ray Imaging (FXI) beamline at the National Synchrotron Light Source II at Brookhaven National Laboratory. At FXI, a full 3D tomography data set can be collected in one minute. This capability opens up new modes of measurements which previously were time-prohibitive, such as 3D XANES or EXAFS, and the ability to measure a large number of samples for statistical significance. The technical details of the beamline and the instrument, as well as some early measurement examples will be discussed.

Proposal for a photoelectron spectroscopy and microscopy beamline (0.5-11 keV) at the High Energy Photon Source.

An optical design study of a beamline proposed for the new 6 GeV synchrotron, the High Energy Photon Source (HEPS), to be built in Beijing, China, is described. The beamline is designed to cover an energy range from 0.5 to 11 keV with two experimental stations, one for X-ray photoelectron spectroscopy (PES) experiments and the other for photoelectron emission microscopy (XPEEM) experiments. A 5 m APPLE II-type undulator with a relatively long magnetic period (55 mm) is used as the only radiation source. To optimize the optical efficiency for the full energy range, the beamline is split into a soft X-ray branch that is based on a variable-line-spacing plane-grating monochromator and a tender X-ray branch that uses a four-bounce monochromator with three Si channel-cut pairs. To allow both PES and XPEEM to be performed over the entire energy range, two toroidal mirrors and a bendable KB mirror pair are employed to deliver the soft and tender beams, respectively, to either of two experimental stations.

Soft/MAX - A new Soft X-ray Microscopy and Coherent Imaging Beamline at the MAX IV Facility.

Soft/MAX - A new Soft X-ray Microscopy and Coherent Imaging Beamline at the MAX IV Facility.