High-pressure X-ray photon correlation spectroscopy at fourth-generation synchrotron sources.

A new experimental setup combining X-ray photon correlation spectroscopy (XPCS) in the hard X-ray regime and a high-pressure sample environment has been developed to monitor the pressure dependence of the internal motion of complex systems down to the atomic scale in the multi-gigapascal range, from room temperature to 600 K. The high flux of coherent high-energy X-rays at fourth-generation synchrotron sources solves the problems caused by the absorption of diamond anvil cells used to generate high pressure, enabling the measurement of the intermediate scattering function over six orders of magnitude in time, from 10-3 s to 103 s. The constraints posed by the high-pressure generation such as the preservation of X-ray coherence, as well as the sample, pressure and temperature stability, are discussed, and the feasibility of high-pressure XPCS is demonstrated through results obtained on metallic glasses.

Lipid vesicle pools studied by passive X-ray microrheology.

Vesicle pools can form by attractive interaction in a solution, mediated by proteins or divalent ions such as calcium. The pools, which are alternatively also denoted as vesicle clusters, form by liquid-liquid phase separation (LLPS) from an initially homogeneous solution. Due to the short range liquid-like order of vesicles in the pool or cluster, the vesicle-rich phase can also be regarded as a condensate, and one would like to better understand not only the structure of these systems, but also their dynamics. The diffusion of vesicles, in particular, is expected to change when vesicles are arrested in a pool. Here we investigate whether passive microrheology based on X-ray photon correlation spectroscopy (XPCS) is a suitable tool to study model systems of artificial lipid vesicles exhibiting LLPS, and more generally also other heterogeneous biomolecular fluids. We show that by adding highly scattering tracer particles to the solution, valuable information on the single vesicle as well as collective dynamics can be inferred. While the correlation functions reveal freely diffusing tracer particles in solutions at low CaCl[Formula: see text] concentrations, the relaxation rate [Formula: see text] shows a nonlinear dependence on [Formula: see text] at a higher concentration of around 8 mM CaCl[Formula: see text], characterised by two linear regimes with a broad cross-over. We explain this finding based on arrested diffusion in percolating vesicle clusters.

Pressure-induced nonmonotonic cross-over of steady relaxation dynamics in a metallic glass.

Relaxation dynamics, as a key to understand glass formation and glassy properties, remains an elusive and challenging issue in condensed matter physics. In this work, in situ high-pressure synchrotron high-energy X-ray photon correlation spectroscopy has been developed to probe the atomic-scale relaxation dynamics of a cerium-based metallic glass during compression. Although the sample density continuously increases, the collective atomic motion initially slows down as generally expected and then counterintuitively accelerates with further compression (density increase), showing an unusual nonmonotonic pressure-induced steady relaxation dynamics cross-over at ~3 GPa. Furthermore, by combining in situ high-pressure synchrotron X-ray diffraction, the relaxation dynamics anomaly is evidenced to closely correlate with the dramatic changes in local atomic structures during compression, rather than monotonically scaling with either sample density or overall stress level. These findings could provide insight into relaxation dynamics and their relationship with local atomic structures of glasses.

The Extremely Brilliant Source storage ring of the European Synchrotron Radiation Facility.

The Extremely Brilliant Source (EBS) is the experimental implementation of the novel Hybrid Multi Bend Achromat (HMBA) storage ring magnetic lattice concept, which has been realised at European Synchrotron Radiation Facility. We present its successful commissioning and first operation. We highlight the strengths of the HMBA design and compare them to the previous designs, on which most operational synchrotron X-ray sources are based. We report on the EBS storage ring's significantly improved horizontal electron beam emittance and other key beam parameters. EBS extends the reach of synchrotron X-ray science confirming the HMBA concept for future facility upgrades and new constructions.

Probing Cage Relaxation in Concentrated Protein Solutions by X-Ray Photon Correlation Spectroscopy.

Diffusion of proteins on length scales of their size is crucial for understanding the machinery of living cells. X-ray photon correlation spectroscopy (XPCS) is currently the only way to access long-time collective diffusion on these length scales, but radiation damage so far limits the use in biological systems. We apply a new approach to use XPCS to measure cage relaxation in crowded α-crystallin solutions. This allows us to correct for radiation effects, obtain missing information on long time diffusion, and support the fundamental analogy between protein and colloid dynamical arrest.

Self-transformation of solid CaCO3 microspheres into core-shell and hollow hierarchical structures revealed by coherent X-ray diffraction imaging.

The self-transformation of solid microspheres into complex core-shell and hollow architectures cannot be explained by classical Ostwald ripening alone. Here, coherent X-ray diffraction imaging and 3D X-ray fluorescence were used to visualize in 3D the formation of hollow microparticles of calcium carbonate in the presence of polystyrene sulfonate (PSS). During the dissolution of the core made from 10-25 nm crystals, the shell developed a global spheroidal shape composed of an innermost layer of 30 nm particles containing high PSS content on which oriented vaterite crystals grew with their c axis mainly oriented along the meridians. The stabilizing role of PSS and the minimization of the intercrystal dipolar energy can explain in combination with Ostwald ripening the formation of these sophisticated structures as encountered in many systems such as ZnO, TiO2, Fe2O3, Co3O4, MnO2, Cu2O, ZnS, CaCO3 and Ca8H2(PO4)6·5H2O.

Resolving molecular diffusion and aggregation of antibody proteins with megahertz X-ray free-electron laser pulses.

X-ray free-electron lasers (XFELs) with megahertz repetition rate can provide novel insights into structural dynamics of biological macromolecule solutions. However, very high dose rates can lead to beam-induced dynamics and structural changes due to radiation damage. Here, we probe the dynamics of dense antibody protein (Ig-PEG) solutions using megahertz X-ray photon correlation spectroscopy (MHz-XPCS) at the European XFEL. By varying the total dose and dose rate, we identify a regime for measuring the motion of proteins in their first coordination shell, quantify XFEL-induced effects such as driven motion, and map out the extent of agglomeration dynamics. The results indicate that for average dose rates below 1.06 kGy µs-1 in a time window up to 10 µs, it is possible to capture the protein dynamics before the onset of beam induced aggregation. We refer to this approach as correlation before aggregation and demonstrate that MHz-XPCS bridges an important spatio-temporal gap in measurement techniques for biological samples.

Nanoscale imaging of shale fragments with coherent X-ray diffraction.

Despite the abundance of shales in the Earth's crust and their industrial and environmental importance, their microscale physical properties are poorly understood, owing to the presence of many structurally related mineral phases and a porous network structure spanning several length scales. Here, the use of coherent X-ray diffraction imaging (CXDI) to study the internal structure of microscopic shale fragments is demonstrated. Simultaneous wide-angle X-ray diffraction (WAXD) measurement facilitated the study of the mineralogy of the shale microparticles. It was possible to identify pyrite nanocrystals as inclusions in the quartz-clay matrix and the volume of closed unconnected pores was estimated. The combined CXDI-WAXD analysis enabled the establishment of a correlation between sample morphology and crystallite shape and size. The results highlight the potential of the combined CXDI-WAXD approach as an upcoming imaging modality for 3D nanoscale studies of shales and other geological formations via serial measurements of microscopic fragments.

Ion-Mediated Cross-linking of Biopolymers Confined at Liquid/Liquid Interfaces Probed by In Situ High-Energy Grazing Incidence X-ray Photon Correlation Spectroscopy.

As manifested in biological cell membranes, the confinement of chemical reactions at the 2D interfaces significantly improves the reaction efficacy. The interface between two liquid phases is used in various key processes in industries, such as in food emulsification and floatation. However, monitoring the changes in the mechanics and dynamics of molecules confined at the liquid/liquid interfaces still remains a scientific challenge because it is nontrivial to access the interface buried under a liquid phase. Herein, we report the in situ monitoring of the cross-linking of polyalginate mediated by Ca2+ ions at the oil/water interface by grazing incidence X-ray photon correlation spectroscopy (GIXPCS). We first optimized the reaction conditions with the aid of interfacial shear rheology and then performed GIXPCS using a high-energy synchrotron X-ray beam (22 keV) that guarantees sufficiently high transmittance through the oil phase. The intensity autocorrelation functions implied that the formation of a percolated network of polyalginate is accompanied by increasing relaxation time. Moreover, the relaxation rate scales linearly with the momentum transfer parallel to the interface, suggesting that the process is driven by hyperdiffusive propagation but not by Brownian diffusion. Our data indicated that high-energy GIXPCS has potential for in situ monitoring of changes in the dynamics of polymers confined between two liquid phases.

Deciphering the intrinsic dynamics from the beam-induced atomic motions in oxide glasses.

Probing the microscopic slow structural relaxation in oxide glasses by X-ray photon correlation spectroscopy (XPCS) revealed faster than expected dynamics induced by the X-ray illumination. The fast beam-induced dynamics mask true slow structural relaxation in glasses and challenges application of XPCS to probe the atomic dynamics in oxide glasses. Here an approach that allows estimation of the true relaxation time of the sample in the presence of beam-induced dynamics is presented. The method requires two measurements either with different X-ray beam intensities or at different temperatures. Using numerical simulations it is shown that the slowest estimated true relaxation time is limited by the accuracy of the measured relaxation times of the sample. By analyzing the reported microscopic dynamics in SiO2, GeO2 and B2O3 glasses, it is concluded that the beam-induced dynamics show rich behavior depending on the sample.

Free log-likelihood as an unbiased metric for coherent diffraction imaging.

Coherent Diffraction Imaging (CDI), a technique where an object is reconstructed from a single (2D or 3D) diffraction pattern, recovers the lost diffraction phases without a priori knowledge of the extent (support) of the object. The uncertainty of the object support can lead to over-fitting and prevents an unambiguous metric evaluation of solutions. We propose to use a 'free' log-likelihood indicator, where a small percentage of points are masked from the reconstruction algorithms, as an unbiased metric to evaluate the validity of computed solutions, independent of the sample studied. We also show how a set of solutions can be analysed through an eigen-decomposition to yield a better estimate of the real object. Example analysis on experimental data is presented both for a test pattern dataset, and the diffraction pattern from a live cyanobacteria cell. The method allows the validation of reconstructions on a wide range of materials (hard condensed or biological), and should be particularly relevant for 4th generation synchrotrons and X-ray free electron lasers, where large, high-throughput datasets require a method for unsupervised data evaluation.

Anisotropic and heterogeneous dynamics in stretched elastomer nanocomposites.

We use X-ray photon correlation spectroscopy (XPCS) to investigate the dynamics of a stretched elastomer by means of probe particles. The particles dispersed in the elastomer were carbon black or silica aggregates classically used for elastomer reinforcement but their volume fraction is very low (φ < 10-2). We show that their dynamics is slower in the direction of the tensile strain than in the perpendicular one. For hydroxylated silica which is poorly wetted by the elastomer, there is no anisotropy. Two-time correlation functions confirm anisotropic dynamics and suggest dynamical heterogeneity already expected from the q-1 behavior of the relaxation times. The height χ* of the peak of the dynamical susceptibility, determined by the normalized variance of the instantaneous correlation function, is larger in the direction parallel to the strain than in the perpendicular one. It also appears that its q dependence changes with the morphology of the probe particle. Therefore, the heterogeneous dynamic probed by the particles is not related only to that of the strained elastomer matrix. In fact, it results from modification of the dynamics of the polymer chains near the surface of the particles and within the aggregate porosity (bound polymer). It is concluded that XPCS is a powerful method for investigating the dynamics, at a given strain, of the bound polymer-particle units which are responsible, at large volume fractions, for the reinforcement.

Mechanical Competence and Bone Quality Develop During Skeletal Growth.

Bone fracture risk is influenced by bone quality, which encompasses bone's composition as well as its multiscale organization and architecture. Aging and disease deteriorate bone quality, leading to reduced mechanical properties and higher fracture incidence. Largely unexplored is how bone quality and mechanical competence progress during longitudinal bone growth. Human femoral cortical bone was acquired from fetal (n = 1), infantile (n = 3), and 2- to 14-year-old cases (n = 4) at the mid-diaphysis. Bone quality was assessed in terms of bone structure, osteocyte characteristics, mineralization, and collagen orientation. The mechanical properties were investigated by measuring tensile deformation at multiple length scales via synchrotron X-ray diffraction. We find dramatic differences in mechanical resistance with age. Specifically, cortical bone in 2- to 14-year-old cases exhibits a 160% greater stiffness and 83% higher strength than fetal/infantile cases. The higher mechanical resistance of the 2- to 14-year-old cases is associated with advantageous bone quality, specifically higher bone volume fraction, better micronscale organization (woven versus lamellar), and higher mean mineralization compared with fetal/infantile cases. Our study reveals that bone quality is superior after remodeling/modeling processes convert the primary woven bone structure to lamellar bone. In this cohort of female children, the microstructural differences at the femoral diaphysis were apparent between the 1- to 2-year-old cases. Indeed, the lamellar bone in 2- to 14-year-old cases had a superior structural organization (collagen and osteocyte characteristics) and composition for resisting deformation and fracture than fetal/infantile bone. Mechanistically, the changes in bone quality during longitudinal bone growth lead to higher fracture resistance because collagen fibrils are better aligned to resist tensile forces, while elevated mean mineralization reinforces the collagen scaffold. Thus, our results reveal inherent weaknesses of the fetal/infantile skeleton signifying its inferior bone quality. These results have implications for pediatric fracture risk, as bone produced at ossification centers during children's longitudinal bone growth could display similarly weak points. © 2019 American Society for Bone and Mineral Research.

In situ coherent X-ray diffraction imaging of radiation-induced mass loss in metal-polymer composite spheres.

A major limitation to the use of coherent X-ray diffraction imaging (CXDI) for imaging soft materials like polymers and biological tissue is that the radiation can cause extensive damage to the sample under investigation. In this study, CXDI has been used to monitor radiation-induced structural changes in metal-coated poly(methyl methacrylate) microspheres. Using a coherent undulator X-ray beam with 8.10 keV photon energy, 14 tomograms at a resolution of ∼30 nm were measured consecutively, which resulted in an accumulated dose of 30 GGy. The three-dimensional images confirmed that the polymer core was strongly affected by the absorbed dose, giving pronounced mass loss. Specifically, as the metal-polymer composite was exposed to the X-ray beam, a bubble-like region of reduced density grew within the composite, almost filling the entire volume within the thin metallic shell in the last tomogram. The bubble seemed to have its initiation point at a hole in the metal coating, emphasizing that the free polymer surface plays an important role in the degradation process. The irradiation of an uncoated polystyrene microsphere gave further evidence for mass loss at the free surface as the radius decreased with increased dose. The CXDI study was complemented by X-ray photon correlation spectroscopy, which proved efficient in establishing exposure dose limits. Our results demonstrate that radiation-induced structural changes at the tens of nanometer scale in soft materials can be followed as a function of dose, which is important for the further development of soft-matter technology.

High-resolution coherent x-ray diffraction imaging of metal-coated polymer microspheres.

Coherent x-ray diffraction imaging (CXDI) is becoming an important 3D quantitative microscopy technique, allowing structural investigation of a wide range of delicate mesoscale samples that cannot be imaged by other techniques like electron microscopy. Here we report high-resolution 3D CXDI performed on spherical microcomposites consisting of a polymer core coated with a triple layer of nickel-gold-silica. These composites are of high interest to the microelectronics industry, where they are applied in conducting adhesives as fine-pitch electrical contacts-which requires an exceptional degree of uniformity and reproducibility. Experimental techniques that can assess the state of the composites non-destructively, preferably also while embedded in electronic chips, are thus in high demand. We demonstrate that using CXDI, all four different material components of the composite could be identified, with radii matching well to the nominal specifications of the manufacturer. Moreover, CXDI provided detailed maps of layer thicknesses, roughnesses, and defects such as holes, thus also facilitating cross-layer correlations. The side length of the voxels in the reconstruction, given by the experimental geometry, was 16 nm. The effective resolution enabled resolving even the thinnest coating layer of ∼20 nm nominal width. We discuss critically the influence of the weak phase approximation and the projection approximation on the reconstructed electron density estimates, demonstrating that the latter has to be employed. We conclude that CXDI has excellent potential as a metrology tool for microscale composites.

Lensless Tomographic Imaging of Near Surface Structures of Frozen Hydrated Malaria-Infected Human Erythrocytes by Coherent X-Ray Diffraction Microscopy.

Lensless, coherent X-ray diffraction microscopy has been drawing considerable attentions for tomographic imaging of whole human cells. In this study, we performed cryogenic coherent X-ray diffraction imaging of human erythrocytes with and without malaria infection. To shed light on structural features near the surface, "ghost cells" were prepared by the removal of cytoplasm. From two-dimensional images, we found that the surface of erythrocytes after 32 h of infection became much rougher compared to that of healthy, uninfected erythrocytes. The Gaussian roughness of an infected erythrocyte surface (69 nm) is about two times larger than that of an uninfected one (31 nm), reflecting the formation of protein knobs on infected erythrocyte surfaces. Three-dimensional tomography further enables to obtain images of the whole cells with no remarkable radiation damage, whose accuracy was estimated using phase retrieval transfer functions to be as good as 64 nm for uninfected and 80 nm for infected erythrocytes, respectively. Future improvements in phase retrieval algorithm, increase in degree of coherence, and higher flux in combination with complementary X-ray fluorescence are necessary to gain both structural and chemical details of mesoscopic architectures, such as cytoskeletons, membraneous structures, and protein complexes, in frozen hydrated human cells, especially under diseased states.

Anisotropic hydrodynamic function of dense confined colloids.

Dense colloidal dispersions exhibit complex wave-vector-dependent diffusion, which is controlled by both direct particle interactions and indirect nonadditive hydrodynamic interactions mediated by the solvent. In bulk the hydrodynamic interactions are probed routinely, but in confined geometries their studies have been hitherto hindered by additional complications due to confining walls. Here we solve this issue by combining high-energy x-ray photon correlation spectroscopy and small-angle x-ray-scattering experiments on colloid-filled microfluidic channels to yield the confined fluid's hydrodynamic function in the short-time limit. Most importantly, we find the confined fluid to exhibit a strongly anisotropic hydrodynamic function, similar to its anisotropic structure factor. This observation is important in order to guide future theoretical research.

Wavefront metrology for coherent hard X-rays by scanning a microsphere.

Characterization of the wavefront of an X-ray beam is of primary importance for all applications where coherence plays a major role. Imaging techniques based on numerically retrieving the phase from interference patterns are often relying on an a-priori assumption of the wavefront shape. In Coherent X-ray Diffraction Imaging (CXDI) a planar incoming wave field is often assumed for the inversion of the measured diffraction pattern, which allows retrieving the real space image via simple Fourier transformation. It is therefore important to know how reliable the plane wave approximation is to describe the real wavefront. Here, we demonstrate that the quantitative wavefront shape and flux distribution of an X-ray beam used for CXDI can be measured by using a micrometer size metal-coated polymer sphere serving in a similar way as the hole array in a Hartmann wavefront sensor. The method relies on monitoring the shape and center of the scattered intensity distribution in the far field using a 2D area detector while raster-scanning the microsphere with respect to the incoming beam. The reconstructed X-ray wavefront was found to have a well-defined central region of approximately 16 µm diameter and a weaker, asymmetric, intensity distribution extending 30 µm from the beam center. The phase front distortion was primarily spherical with an effective radius of 0.55 m which matches the distance to the last upstream beam-defining slit, and could be accurately represented by Zernike polynomials.

Multi-speckle X-ray photon correlation spectroscopy in the ultra-small-angle X-ray scattering range.

Multi-speckle X-ray photon correlation spectroscopy (XPCS) measurements in the ultra-small-angle range are performed using a long pinhole collimation instrument in combination with two-dimensional photon-counting and high-sensitivity imaging detectors. The feasibility of the presented setup to measure dynamics on different time and length scales pertinent to colloidal systems is shown. This setup offers new research opportunities, such as for example in the investigation of non-equilibrium dynamics in optically opaque, complex systems over length scales from tens of nanometres to several micrometres. In addition, due to the short duration of the X-ray exposure involved in the ultra-small-angle range, possible radiation-induced effects are alleviated. Furthermore, the performance of two different detectors, a photon-counting Pilatus 300K and an integrating FReLoN CCD, are compared, and their applicability for accurate XPCS measurements is demonstrated.

Anisotropic de Gennes Narrowing in Confined Fluids.

The collective diffusion of dense fluids in spatial confinement is studied by combining high-energy (21 keV) x-ray photon correlation spectroscopy and small-angle x-ray scattering from colloid-filled microfluidic channels. We find the structural relaxation in confinement to be slower compared to the bulk. The collective dynamics is wave vector dependent, akin to the de Gennes narrowing typically observed in bulk fluids. However, in stark contrast to the bulk, the structure factor and de Gennes narrowing in confinement are anisotropic. These experimental observations are essential in order to develop a microscopic theoretical description of collective diffusion of dense fluids in confined geometries.