ESRF Research Articles

The in crystallo optical spectroscopy toolbox

Over the past ten years, there has been a surge in the demand for in crystallo optical spectroscopy (icOS), since optical spectroscopy is one of the few biophysical characterization methods applicable to both protein solutions and crystals. Historically, icOS has been used to compare the state of proteins in crystals and in solution, and to assess their functionality by determining the redox state of metal ions, cofactors or chromophores. The recent rejuvenation of time-resolved crystallography experiments has sparked a renewed interest in optical spectroscopy as a bridge between kinetic studies in solution and in the crystalline state. The method of icOS can be defined as the ensemble of spectroscopic techniques in the UV–visible–infrared range that can be applied to crystals. It has also been instrumental in understanding specific X-ray radiation damage to redox-sensitive parts of proteins. Spectra recorded from crystals are affected by crystal orientation, shape or position due to various optical phenomena. Fortunately, these can be modelled and their effect can be corrected. The icOS laboratory at the European Synchrotron Radiation Facility (ESRF) specializes in recording UV–Vis absorption, fluorescence emission and Raman spectra from protein crystals. Here, we present a suite of utilities that streamline the analysis and correction of UV–Vis absorption icOS data, encased in a graphical interface. This was originally developed for the icOS laboratory at ESRF but is available as a standalone package, with the aim of making icOS more accessible.

Structural dependency of polymer dynamics by means of small-angle X-ray photon correlation spectroscopy and wide-angle X-ray scattering on the D2AM beamline.

X-ray photon correlation spectroscopy (XPCS) has become a pivotal technique for exploring nanoscale dynamic phenomena across various materials, facilitated by advancements in synchrotron radiation sources and beamline upgrades. The recent Extremely Brilliant Source (EBS) upgrade at the European Synchrotron Radiation Facility (ESRF) in Grenoble, France, has notably improved brilliance and coherence length, thereby enhancing the capabilities of XPCS and related techniques. Here, we present a dedicated setup on the D2AM beamline at the ESRF, enabling simultaneous XPCS and wide-angle X-ray scattering measurements. The setup developed and its performance are detailed in the first part. Then, the XPCS capabilities are evaluated by studying polymer-based materials, with particular attention to the effects of temperature, crystallinity and macromolecular orientation on polymer dynamics. The study on the influence of temperature revealed that XPCS in the case of entangled polymers is an efficient technique to probe the dynamics of the macromolecular network, complementary to classical spectroscopy techniques. In addition, in situ measurements during the polymer crystallization revealed that increased crystallinity slows down macromolecular dynamics. Conversely, studies on stretched samples indicate that macromolecular orientation accelerates these dynamics. This work represents a novel investigation into the effect of crystallinity on macromolecular dynamics using XPCS, opening new avenues for research in polymer science.

Analysis of microcracking processes in microconcrete under confined compression utilising synchrotron-based ultra-high speed X-ray phase contrast imaging

Analysis of microcracking processes in microconcrete under confined compression utilising synchrotron-based ultra-high speed X-ray phase contrast imaging

Electrothermal instabilities observed by x-ray radiography of underwater sub-microsecond electrical explosions of aluminum, silver, and molybdenum wires

We present measurements of the wavelength of electrothermal instabilities (ETI) formed during underwater electrical explosions of aluminum (Al), silver (Ag), and molybdenum (Mo) wires. Wires were exploded using a ∼450 ns rise time and ∼120 kA amplitude current pulse delivered by a pulse generator. Images of the exploding wires were captured by multi-frame synchrotron radiography at the ID19 beamline of the European Synchrotron Radiation Facility. Resolvable ETI was observed only in Al and Ag wires after the vaporization phase, whereas no such instabilities were detectable in Mo wires. Fourier analysis revealed that the ETI wavelengths in Al and Ag wires were comparable within the spatial resolution error, despite their different minimal instability wavelengths, which were predicted to develop during the melting phase. These minimal wavelengths were calculated using the linear ETI development theory and the simulated average wire temperature. The latter was calculated using one-dimensional hydrodynamic simulations, considering uniform current density across the wire cross-sectional area.

ULtimate MAGnetic characterization (ULMAG) at the ID12 beamline of ESRF: from element-specific properties to macroscopic functionalities.

We present a novel instrument designed for advanced magnetic study, installed at the ID12 beamline of the European Synchrotron Radiation Facility in Grenoble, France. This instrument offers the unique capability to simultaneously measure element-specific microscopic and macroscopic properties related to the magnetic, electronic and structural characteristics of materials. In addition to X-ray absorption, X-ray magnetic circular dichroism alongside X-ray diffraction patterns, the macroscopic magnetization, volume changes, caloric properties and electrical resistivity of magnetic materials could be measured strictly under the same experimental conditions as a function of both magnetic field (up to ±7 T) and temperature (ranging from 2.05 K to 325 K). To demonstrate the capability of this new instrument, we present two case studies highlighting its performance in investigating first-order magneto-structural phase transitions, namely in DyCo2 and FeRh alloys.

Automatisation developments on ESRF-EBS BM05 beamline

X-ray computed tomography and radiography imaging are currently the second most exploited technique at the European Synchrotron Radiation Facility (ESRF) for commercial services. To maximise the growth, the beamlines must be flexible enough to accommodate a wide range of materials and sample sizes, whilst conserving operational efficiency and ensuring reliable and high-quality imaging outcomes. In this paper, we present some of the latest developments on the BM05 tomography beamline from the ESRF in terms of automatisation, processes, and procedures that ensure an efficient and robust operation of the beamline. For all users, the automatisation developments are focused on being able to set-up the beamline in a pre-defined state as easily and quickly as possible, so that time consuming user interventions (and potential mistakes) are minimised. For academic users, a graphical user interface (GUI) called Daiquiri tomo has been developed as a simple tool for non-experts to be able to perform most of their scanning operations in a user-friendly and efficient fashion, with minimum support from beamline scientists. For commercial users, a robotic sample changer has been assembled to be able to scan series of up to 45 samples, and maximise usage of night shifts. Overall, those improvements are key in reaching operational excellence and X-ray computed tomography and radiography imaging are projected to rival macromolecular crystallography (MX) as the first technique for commercial services at ESRF by the end of 2025.

Three-dimensional structure of entire hydrated murine hearts at histological resolution

Imaging the entire cardiomyocyte network in entire small animal hearts at single cell resolution is a formidable challenge. Optical microscopy provides sufficient contrast and resolution in 2d, however fails to deliver non-destructive 3d reconstructions with isotropic resolution. It requires several invasive preparation steps, which introduce structural artefacts, namely dehydration, physical slicing and staining, or for the case of light sheet microscopy also clearing of the tissue. Our goal is to provide 3d reconstructions of the cardiomyocyte network in entire hydrated murine hearts, and to develop a methodology for quantitative analysis of heart pathologies based on X-ray phase contrast computed tomography (XPCT). We have used XPCT at two beamlines of the extremely brilliant source (EBS) at the European Synchrotron Radiation Facility (ESRF) to scan wild-type murine hearts at high resolution, as well as a series of murine hearts of different pathological models, at reduced resolution and higher throughput. All hearts were obtained from the small animal facility of the university medical center in Göttingen. The hearts were fixed in formalin, stored and measured non-destructively in phosphate buffer solution. The high resolution dataset allows to discern individual cardiomyocytes in the tissue. All datasets have been analyzed using semi-automated image segmentation of the ventricles, rotation into a common coordinate system, classification into different anatomical compartments, and finally the structure tensor approach. A 3d streamline representation of the cardiomyocyte orientation vector field is provided. The different cardiovascular disease models are analysed based on metrics derived from the 3d structure tensor. An entire hydrated murine heart has been covered at an isotropic voxel size of 1.6μm (distributed over several volumes). A binned and fused dataset of this heart is available at 3.2μm, and has been analyzed by the structure tensor approach to yield the ventricular cardiomyocyte network or mesh, i.e. the aggregation of the cardiomyocyte chains in particular in the ventricular wall. Semi-automatic determination of structural metrics is already achieved and the corresponding tools and resulting data are made publically available. XPCT using extremely brilliant undulator radiation is close to achieve single cell reconstruction in an entire small animal organ.

Exploiting fourth-generation synchrotron radiation for enzyme and photoreceptor characterization.

The upgrade of the European Synchrotron Radiation Facility (ESRF) in Grenoble, France to an Extremely Brilliant Source (EBS) is expected to enable time-resolved synchrotron serial crystallography (SSX) experiments with sub-millisecond time resolution. ID29 is a new beamline dedicated to SSX experiments at ESRF-EBS. Here, we report experiments emerging from the initial phase of user operation at ID29. We first used microcrystals of photoactive yellow protein as a model system to exploit the potential of microsecond pulses for SSX. Subsequently, we investigated microcrystals of cytochrome c nitrite reductase (ccNiR) with microsecond X-ray pulses. CcNiR is a decaheme protein that is ideal for the investigation of radiation damage at the various heme-iron sites. Finally, we performed a proof-of-concept subsecond time-resolved SSX experiment by photoactivating microcrystals of a myxobacterial phytochrome.

Improving Catalytic Activity of Solid Oxide Electrolysers for the Electrosplitting of Water Vapor and CO2 by the Addition of Affordable and Eco-Friendly Promoter Metals

Recently, Solid Oxide Electrolysis Cells (SOECs) can be considered a solution for the elimination of atmospheric CO2 and limiting its negative environmental impact. Thanks to high operation temperatures, full-solid construction and high Faradaic efficiency it is a promising method for a production of great amounts of relatively cheap and pure hydrogen. Furthermore, these ceramic cells can be set to perform CO2 electroreduction along with water vapor. Thanks to that, it is possible to simultaneously supply various chemical processes with H2-CO feedstock using one, relatively simple setup. In that sense, the SOECs can provide ‘green’ hydrogen and reduce the CO2 amount by delivering the valuables such as syngas or other chemicals. Recently, high temperature H2O/CO2 coelectrolysis in SOECs became reasonable alternative to other technologies of syngas production such as steam reforming of biogas or coal gasification, which both release huge amount of the greenhouse gases.The main constituent of water-hydrogen electrode is bifunctional composite of NiO and Zr0.84Y0.16O2-δ, which upon the reduction forms metallic network of Ni within the scaffold of ionic conductor. This type of electrode is quite well established for working with water vapor in electrolysis mode but struggles under the atmosphere containing CO and/or CO2. Ni grains are considered active, but not the most efficient towards RWGS as well as electrochemical splitting of H2O/CO2. Over the years other transition metals - Cu, Fe, or Co - as well as their oxides - MnO, ZnO - were found to be more prominent and used in catalytic reactors. Further increase of the activity and the efficiency can be achieved thanks to promoter metals and their oxides e.g., K2O, Na2O, Rb2O, MgO, CaO. The addition of the transition metals can be promising alternative to the usage of the noble metals considering mostly their much lower price. On the other hand, the addition of alkaline metal oxides can increase the adsorption rate of CO2 and prolong the retention time in the proximity of the electrode. As the SOECs are generally non-pressurized systems, the efficiency of the electrode surface catalysts should be pushed to maximum in order to provide valuable yield of the outlet gases.A series of modified SOECs was prepared by the addition of 5 mol.% of the transition metal (Co, Cu, Mn, Fe) or alkaline metal oxide (Na, K, Ca, Mg, Rb, Li, Sr) nanoparticles. The samples have been fabricated by simple wet impregnation method. The SOECs were tested for the coelectrolysis of CO2/H2O mixture with an addition of H2 for electrode protection against the reoxidation. The electrical tests were considering the utilization factor of water vapor and electrochemical behavior of the cells under various CO2:H2:H2O ratios to better understand the mechanisms of syngas formation.It was found out that depending on the reducibility of the metals they got dissolved into Ni grains (e.g., Co, Fe) or formed the secondary phases on the top of the grains (e.g. MnO). It was observed as an accumulation of the metal ions inside Ni-rich grains or dissipation of the ions all over the imaging area as seen in STXM images. Alkaline metal oxides formed a mixture of the phases all over Ni and YSZ grains. A series of in-situ XAS measurements at the ESRF (France) was performed to observe the formation of various mixed compounds and the response of electrode's components to CO2-rich atmosphere at high temperature. The surficial changes of the oxidation states were determined using in-situ XPS imaging of modified electrode structure using SPEM at the Elettra (Italy). The addition of secondary metal alters the energy levels on Ni surface and increases the tendency to form carbonate-like compounds, what prolongs the retention time of CO2. The addition of guest metal highly increased CO2 conversion and selectivity towards CO production as well as the electrical efficiency due to the alterations of basic-acid sites and introduction of active redox couples. Acknowledgements This work was supported by a project funded by National Science Centre Poland, based on decision UMO-2021/43/B/ST8/01831 (OPUS 22). The XAS and XPS experiments were supported by CERIC-ERIC consortium and performed at the European Synchrotron Radiation Facility (ESRF) and the Elettra Sincrotrone Trieste, respectively. The access to ESRF was financed by the Polish Ministry of Education and Science – decision no. 2021/WK/11.

Deep learning for 3D vascular segmentation in hierarchical phase contrast tomography: a case study on kidney

Automated blood vessel segmentation is critical for biomedical image analysis, as vessel morphology changes are associated with numerous pathologies. Still, precise segmentation is difficult due to the complexity of vascular structures, anatomical variations across patients, the scarcity of annotated public datasets, and the quality of images. Our goal is to provide a foundation on the topic and identify a robust baseline model for application to vascular segmentation using a new imaging modality, Hierarchical Phase-Contrast Tomography (HiP-CT). We begin with an extensive review of current machine-learning approaches for vascular segmentation across various organs. Our work introduces a meticulously curated training dataset, verified by double annotators, consisting of vascular data from three kidneys imaged using HiP-CT as part of the Human Organ Atlas Project. HiP-CT pioneered at the European Synchrotron Radiation Facility in 2020, revolutionizes 3D organ imaging by offering a resolution of around 20 μm/voxel and enabling highly detailed localised zooms up to 1–2 μm/voxel without physical sectioning. We leverage the nnU-Net framework to evaluate model performance on this high-resolution dataset, using both known and novel samples, and implementing metrics tailored for vascular structures. Our comprehensive review and empirical analysis on HiP-CT data sets a new standard for evaluating machine learning models in high-resolution organ imaging. Our three experiments yielded Dice similarity coefficient (DSC) scores of 0.9523, 0.9410, and 0.8585, respectively. Nevertheless, DSC primarily assesses voxel-to-voxel concordance, overlooking several crucial characteristics of the vessels and should not be the sole metric for deciding the performance of vascular segmentation. Our results show that while segmentations yielded reasonably high scores-such as centerline DSC ranging from 0.82 to 0.88, certain errors persisted. Specifically, large vessels that collapsed due to the lack of hydrostatic pressure (HiP-CT is an ex vivo technique) were segmented poorly. Moreover, decreased connectivity in finer vessels and higher segmentation errors at vessel boundaries were observed. Such errors, particularly in significant vessels, obstruct the understanding of the structures by interrupting vascular tree connectivity. Our study establishes the benchmark across various evaluation metrics, for vascular segmentation of HiP-CT imaging data, an imaging technology that has the potential to substantively shift our understanding of human vascular networks.

Study of a gel dosimeter based on Ag nanoparticles for applications in radiation therapy with synchrotron X-rays at ultrahigh dose rate compared to 60Co γ-rays

Study of a gel dosimeter based on Ag nanoparticles for applications in radiation therapy with synchrotron X-rays at ultrahigh dose rate compared to 60Co γ-rays

A miniature X-ray diffraction setup on ID20 at the European Synchrotron Radiation Facility.

We describe an ultra-compact setup for in situ X-ray diffraction on the inelastic X-ray scattering beamline ID20 at the European Synchrotron Radiation Facility. The main motivation for the design and construction of this setup is the increasing demand for on-the-fly sample characterization, as well as ease of navigation through a sample's phase diagram, for example subjected to high-pressure and/or high-temperature conditions. We provide technical details and demonstrate the performance of the setup.

Hard x-ray magnetic circular dichroism under high magnetic field

We describe high-field X-ray magnetic circular dichroism (XMCD) setup developed at the European Synchrotron Radiation Facility beamline ID12, which is dedicated to polarization dependent X-ray absorption spectroscopy in the soft and hard X-ray range from 2 to 15 keV. The static magnetic field up to 17 T is generated by a superconducting solenoid. Performance of the setup is illustrated with three representative studies: metamagnetic transitions in PrCo2Ge2 single crystal, Pauli paramagnetism in uranium monocarbide, and induced magnetism on gold atoms in different environment: MnAu4, UAu4 and a pure gold metal.

Phase diagram of ruthenium characterized In Situ by synchrotron X-ray diffraction and Ab Initio simulations

In this work, an in situ characterization of the high-pressure and high-temperature phase diagram of ruthenium was carried out. This experiment was performed using a combination of laser-heated diamond anvil cell (LH-DAC) and X-ray diffraction (XRD) techniques at the beamline ID27 of the European Synchrotron Radiation Facility (ESRF). XRD patterns were collected in the range of 15 GPa to 110 GPa and from ambient temperature up to 6600 K. While the hcp-fcc transition, predicted to occur at elevated temperatures was not observed, this study has produced the first experimental observation of a solid–liquid phase transition of Ru at HP conditions. A P-V-T equation of state valid up to 100 GPa and 3000 K is reported.

Bimetallic Ir-Ru 25:75 Catalyst for PEM-WE: Uncovering the Origin of Its Activity by Operando Wide Angle X-Ray Scattering

Proton Exchange Membrane Water Electrolyzers (PEM-WEs) are entering the phase of commercial mass production. However, the issue of an iridium catalyst for the anode remains. Our previous work presented an iridium-ruthenium-based catalyst (25% Ir = 158 µg cm-2, 75% Ru) prepared as a thin film on the surface-enhanced PEM-WE-anode via magnetron sputtering. This catalyst demonstrated excellent activity and stability in single-cell PEM-WE (1 A cm-2 at 1.606 V, 80 °C, degradation rate – 1.3 µV h-1 at 1 A cm-2 for 500 hours). The origin of its outstanding electrochemical performance was, however, not known. We present a complete operando Wide Angle X-Ray Scattering (WAXS) study done at the European Synchrotron Radiation Facility (ESRF) using a dedicated PEM-WE cell. The structure of each sample is observed for several hours under PEM-WE conditions with increasing current density from 100 mA cm-2 to 600 mA cm-2. Consequently, we show the connection of the electrochemical performance of the Ir-Ru 25:75 catalyst to its structural properties and uncover the interplay between the face-centered-cubic iridium and hexagonal-close-packed ruthenium.

Deciphering the Impact of Current, Composition, and Potential on the Lithiation Behavior of Graphite in Silicon-Graphite Anodes

Graphite is the most used anode material for current-generation lithium-ion batteries due to its beneficial cycle life, availability and reasonable rate capability. Still, the modest capacity of 372 mAhg-1 represents a bottleneck in the pursuit of high-energy density anodes. Enabling much higher theoretical capacities of 3600 mAhg-1 [1], silicon represents a promising candidate as anode material for next generation batteries. However, the alloying-based lithiation mechanism of silicon is accompanied by a large volumetric change of up to 300% upon cycling, leading to fast degradation due to electrode pulverization, continuous SEI formation and extensive electrolyte consumption. Even though shallow cycling of silicon anodes has been demonstrated in laboratory scale cells [2], the mixing of silicon with graphite is considered the most viable approach to lift energy density while maintaining appropriate cycle life.Due to the fundamentally different kinetics and potentials of Li insertion and extraction of both materials, the development of silicon-graphite composite electrodes is not straightforward. One of the key questions for optimization is how the incorporation of silicon impacts the (de)lithiation behavior of graphite as a function of the silicon:graphite mass ratio, operating current and electrode potential. While lithiation of graphite in composite electrodes of 15 wt% silicon has been shown to occur in a similar manner as pure graphite, higher fractions of silicon are expected to exert more stress on the graphite and represent kinetic barriers for Li diffusion. This study presents results from in-situ X-ray diffraction experiments (XRD) of silicon-graphite/Li half cells conducted at the European Synchrotron Radiation Facility (ESRF). Serving as a reference, the current-dependent (de)lithiation behavior of a pure graphite/Li half cell has been thoroughly investigated and systematically compared to the (de)lithiation behavior of two selected silicon-graphite composite anodes (silicon:graphite ratio of 30:70 and 70:30, respectively) exposed to the same formation cycle and C-rate protocol. An optimized and novel measurement setup was used, based on a perforated current collector of the anode, providing a complete picture of the graphite in-plane and interplanar structural changes as well as the evolution of semicrystalline silicon peaks which would usually be obscured by the current collector signal.By correlating the electrochemical features (voltage curve and differential capacity plot) with the in-situ diffraction data, it was possible to identify and assign the occurrence and absence of dilute-stage and ordered graphite intercalation compounds (GICs) for the respective electrodes, yielding an in-depth insight into the graphite state of charge upon (de)lithiation and how it is influenced by the silicon content and applied C-rate.As expected, the silicon is (de)lithiated within the entire potential range, whereas graphite is most electrochemically active at potentials lower than 260 mV. In both composite electrodes, graphite attains a lower lithiation degree compared to pure graphite. This is because the high theoretical capacity of silicon results in high specific currents, ultimately challenging the rate capability of graphite. Moreover, the high delithiation overpotential encountered in the composite electrodes results in a highly asymmetrical lithiation/delithiation behavior of graphite. Also, graphite lithiation is less uniform in the composite anodes, indicated by the coexistence of dilute GICs upon (de)lithiation, compared to pure graphite where dilute phases are fully consumed (formed) as (de)lithiation progresses. Surprisingly, the in-situ XRD studies did not reveal signs of structural degradation and strain of graphite as a result of mechanical interaction with silicon. Instead, the volume change of graphite is well correlated with the amorphization of crystalline, unreacted silicon and the volume evolution of silicon crystallites upon charge-discharge.To mitigate the observed effects, the work suggests nanostructuring and advanced electrode architectures towards higher utilization and more homogeneous lithiation of graphite. Overall, the study provides a demonstration of a suitable operando cell for studies of anodes for Li-based systems, and aids the rational design of silicon-graphite composite anodes.

Response of the first POLAR-2 prototype to polarized beams

POLAR-2 is a dedicated gamma-ray polarimeter currently foreseen to be launched towards the China Space Station around 2027. The design of the detector is based on the legacy of its predecessor mission POLAR which was launched in 2016. POLAR-2 aims to measure the polarization of the Gamma-ray Burst prompt emission within the 30–800 keV energy range. Thanks to its high sensitivity to gamma-ray polarization, as well as its large effective area, POLAR-2 will provide the most precise measurements of this type to date. Such measurements are key to improve our understanding of the astrophysical processes responsible for Gamma-Ray Bursts. The detector consists of a segmented array of plastic scintillator bars, each one of which is read out by a Silicon PhotoMultiplier channel. The flight model of POLAR-2 will contain a total of 6400 scintillators. These are divided into 100 groups of 64 bars each, in so-called polarimeter modules. In recent years, the collaboration has designed and produced the first prototypes of these polarimeter modules and subjected these to space qualification tests. In addition, in April 2023, the first of these modules were calibrated using fully polarized gamma-ray beams at the European Synchrotron Radiation Facility (ESRF) in France. In this work, we will present the results of this calibration campaign and compare these to the simulated performance of the POLAR-2 modules. Potential improvements to the design are also discussed. Finally, the measurements are used, in combination with the verified simulation framework, to estimate the scientific performance of the full POLAR-2 detector and compare it to its predecessor.

Deep Learning for 3D Vascular Segmentation in Phase Contrast Tomography.

Automated blood vessel segmentation is critical for biomedical image analysis, as vessel morphology changes are associated with numerous pathologies. Still, precise segmentation is difficult due to the complexity of vascular structures, anatomical variations across patients, the scarcity of annotated public datasets, and the quality of images. Our goal is to provide a foundation on the topic and identify a robust baseline model for application to vascular segmentation using a new imaging modality, Hierarchical Phase-Contrast Tomography (HiP-CT). We begin with an extensive review of current machine learning approaches for vascular segmentation across various organs. Our work introduces a meticulously curated training dataset, verified by double annotators, consisting of vascular data from three kidneys imaged using Hierarchical Phase-Contrast Tomography (HiP-CT) as part of the Human Organ Atlas Project. HiP-CT, pioneered at the European Synchrotron Radiation Facility in 2020, revolutionizes 3D organ imaging by offering resolution around 20μm/voxel, and enabling highly detailed localized zooms up to 1μm/voxel without physical sectioning. We leverage the nnU-Net framework to evaluate model performance on this high-resolution dataset, using both known and novel samples, and implementing metrics tailored for vascular structures. Our comprehensive review and empirical analysis on HiP-CT data sets a new standard for evaluating machine learning models in high-resolution organ imaging. Our three experiments yielded Dice scores of 0.9523 and 0.9410, and 0.8585, respectively. Nevertheless, DSC primarily assesses voxel-to-voxel concordance, overlooking several crucial characteristics of the vessels and should not be the sole metric for deciding the performance of vascular segmentation. Our results show that while segmentations yielded reasonably high scores-such as centerline Dice values ranging from 0.82 to 0.88, certain errors persisted. Specifically, large vessels that collapsed due to the lack of hydro-static pressure (HiP-CT is an ex vivo technique) were segmented poorly. Moreover, decreased connectivity in finer vessels and higher segmentation errors at vessel boundaries were observed. Such errors, particularly in significant vessels, obstruct the understanding of the structures by interrupting vascular tree connectivity. Through our review and outputs, we aim to set a benchmark for subsequent model evaluations using various modalities, especially with the HiP-CT imaging database.

In situ serial crystallography facilitates 96-well plate structural analysis at low symmetry.

The advent of serial crystallography has rejuvenated and popularized room-temperature X-ray crystal structure determination. Structures determined at physiological temperature reveal protein flexibility and dynamics. In addition, challenging samples (e.g. large complexes, membrane proteins and viruses) form fragile crystals that are often difficult to harvest for cryo-crystallography. Moreover, a typical serial crystallography experiment requires a large number of microcrystals, mainly achievable through batch crystallization. Many medically relevant samples are expressed in mammalian cell lines, producing a meager quantity of protein that is incompatible with batch crystallization. This can limit the scope of serial crystallography approaches. Direct in situ data collection from a 96-well crystallization plate enables not only the identification of the best diffracting crystallization condition but also the possibility for structure determination under ambient conditions. Here, we describe an in situ serial crystallography (iSX) approach, facilitating direct measurement from crystallization plates mounted on a rapidly exchangeable universal plate holder deployed at a microfocus beamline, ID23-2, at the European Synchrotron Radiation Facility. We applied our iSX approach on a challenging project, autotaxin, a therapeutic target expressed in a stable human cell line, to determine the structure in the lowest-symmetry P1 space group at 3.0 Å resolution. Our in situ data collection strategy provided a complete dataset for structure determination while screening various crystallization conditions. Our data analysis reveals that the iSX approach is highly efficient at a microfocus beamline, improving throughput and demonstrating how crystallization plates can be routinely used as an alternative method of presenting samples for serial crystallography experiments at synchrotrons.

EBS status of the large-volume press at beamline ID06-LVP

ABSTRACT The European Synchrotron Radiation Facility (ESRF)'s large-volume press (LVP) instrument, at beamline ID06-LVP, is in constant evolution and made a significant step-change at the Extremely Brilliant Source upgrade; with improved beam characteristics (stability and focussing) and detection capabilities, with the installation of a custom-built Dectris 900 kW CdTe device. Over the same time-frame, higher pressure regimes have been reached through use of improved 6/6 compression, with the incorporation of a Drickamer device, and through the use of harder carbide grades in 6/8. The combined use of in-situ X-ray diffraction, imaging and pulse-echo ultrasonic high-pressure experiments has been enhanced. Temperature generation and measurement methods have been upgraded. User interaction is improved with live data inspection and treatment possible. Data calibration and integration have been thoroughly revised. Here, we describe the current status and highlight recent upgrades at ID06-LVP, with examples drawn from user measurements and in-house testing.