Heterogeneous and self-organizing mineralization of bone matrix promoted by hydroxyapatite nanoparticles

The mineralization process is crucial to the load-bearing characteristics of the bone extracellular matrix. In this work, we have studied the spatiotemporal dynamics of mineral deposition by human bone marrow mesenchymal stem cells differentiating toward osteoblasts promoted by the presence of exogenous hydroxyapatite nanoparticles. At molecular level, the added nanoparticles positively modulated the expression of bone-specific markers and enhanced calcified matrix deposition during osteogenic differentiation. The nucleation, growth and spatial arrangement of newly deposited hydroxyapatite nanocrystals have been evaluated using Scanning Micro X-Ray Diffraction and Scanning Micro X-Ray Fluorescence. As leading results, we have found the emergence of a complex scenario where the spatial organization and temporal evolution of the process exhibit a heterogeneous and self-organizing dynamics. At the same time the possibility to control the differentiation kinetic through the addition of synthetic nanoparticles, paves the way to empower the generation of more structured bone scaffolds in tissue engineering and to design new drugs in regenerative medicine

In Situ Ptychography of Heterogeneous Catalysts using Hard X-Rays: High Resolution Imaging at Ambient Pressure and Elevated Temperature

A new closed cell is presented for in situ X-ray ptychography which allows studies under gas flow and at elevated temperature. In order to gain complementary information by transmission and scanning electron microscopy, the cell makes use of a Protochips E-chipTM which contains a small, thin electron transparent window and allows heating. Two gold-based systems, 50 nm gold particles and nanoporous gold as a relevant catalyst sample, were used for studying the feasibility of the cell. Measurements showing a resolution around 40 nm have been achieved under a flow of synthetic air and during heating up to temperatures of 933 K. An elevated temperature exhibited little influence on image quality and resolution. With this study, the potential of in situ hard X-ray ptychography for investigating annealing processes of real catalyst samples is demonstrated. Furthermore, the possibility to use the same sample holder for ex situ electron microscopy before and after the in situ study underlines the unique possibilities available with this combination of electron microscopy and X-ray microscopy on the same sample

Megahertz pulse trains enable multi-hit serial femtosecond crystallography experiments at X-ray free electron lasers

The European X-ray Free Electron Laser (XFEL) and Linac Coherent Light Source (LCLS) II are extremely intense sources of X-rays capable of generating Serial Femtosecond Crystallography (SFX) data at megahertz (MHz) repetition rates. Previous work has shown that it is possible to use consecutive X-ray pulses to collect diffraction patterns from individual crystals. Here, we exploit the MHz pulse structure of the European XFEL to obtain two complete datasets from the same lysozyme crystal, first hit and the second hit, before it exits the beam. The two datasets, separated by <1 µs, yield up to 2.1 Å resolution structures. Comparisons between the two structures reveal no indications of radiation damage or significant changes within the active site, consistent with the calculated dose estimates. This demonstrates MHz SFX can be used as a tool for tracking sub-microsecond structural changes in individual single crystals, a technique we refer to as multi-hit SFX

Microfluidic devices for X-ray studies on hydrated cells

X-ray studies of biological cells in microfluidic devices provide a method to probe cellular structures or structural changes at the molecular level in a precisely controlled environment. However, the device design and the used materials must be compatible with X-ray scattering techniques as well as the cell culture in the devices. For this purpose, we develop new types of X-ray compatible microfluidic devices, which are based on a UV-curable adhesive as a moldable material, and thin Kapton films and silicon nitride membrane windows as a growth substrate for cells and as a window material for X-rays. Using these devices, we perform scanning X-ray diffraction experiments with a nano-focused beam on fixed cells in buffer solution. In principle, these microfluidic devices also allow for X-ray studies on living cells

Linear focus solar simulator test bench for non-destructive optical efficiency testing of parabolic trough receivers

This paper describes a new 2nd generation linear focus solar simulator test bench. The test bench is used for the non-destructive calorimetric measurement of relative optical efficiency of parabolic trough receivers. Compared to the predecessor, the test bench has been optimized for faster measurement, better long term stability and easier operation. The test bench optics is an elliptical shaped glass mirror trough with flat end mirrors. The receiver and six solar simulator lamps, metal halide lamps with 2.5 kWe each are located in their respective focal lines. Absorbed power is measured via temperature increase of the water flowing through the receiver and the mass flow rate. By comparison to the absorbed power of a reference receiver, the optical efficiency of a sample can be determined relative to the optical efficiency of the reference receiver

Membrane fusion intermediates and the effect of cholesterol: An in-house X-ray scattering study

We have developed an X-ray scattering setup which allows to study membrane fusion intermediates or other nonlamellar lipid mesophases by laboratory-scale X-ray sources alone, thus taking advantage of unrestricted beamtime compared to synchrotron sources. We report results of a study of pure lipid bilayers and phospholipid/cholesterol binary mixtures. Stalks, putative intermediate structures occurring during the membrane fusion process, can clearly be identified from reconstructed electron density maps. Phase diagrams of the lyotropic phase behavior of DOPC/cholesterol and DPhPC/cholesterol samples are presented. If cholesterol is present in moderate concentrations, it can substantially promote the formation of stalks at higher degree of hydration. In addition, a possibly new phase in DOPC/cholesterol is found at high cholesterol content in the low humidity range

Anisotropic x-ray scattering and orientation fields in cardiac tissue cells

X-ray diffraction from biomolecular assemblies is a powerful technique which can provide structural information about complex architectures such as the locomotor systems underlying muscle contraction. However, in its conventional form, macromolecular diffraction averages over large ensembles. Progress in x-ray optics has now enabled to probe structures on sub-cellular scales, with the beam confined to a distinct organelle. Here, we use scanning small angle x-ray scattering (scanning SAXS) to probe the diffraction from cytoskeleton networks in cardiac tissue cells. In particular, we focus on actin-myosin composites, which we identify as the dominating contribution to the anisotropic diffraction patterns, by correlation with optical fluorescence microscopy. To this end, we use a principal component analysis approach to quantify direction, degree of orientation, nematic order, and the second moment of the scattering distribution in each scan point. We compare the fiber orientation from micrographs of fluorescently labeled actin fibers to the structure orientation of the x-ray dataset and thus correlate signals of two different measurements: the native electron density distribution of the local probing area versus specifically labeled constituents of the sample. Further, we develop a robust and automated fitting approach based on a power law expansion, in order to describe the local structure factor in each scan point over a broad range of the momentum transfer ${q}_{{\rm{r}}}$. Finally, we demonstrate how the methodology shown for freeze dried cells in the first part of the paper can be translated to alive cell recordings

Influence of cholesterol on the collective dynamics of the phospholipid acyl chains in model membranes

We have studied the packing and collective dynamics of the phospholipid acyl chains in a model membrane composed of 1,2 dimyristoyl sn glycero 3 phosphatidylcholine DMPC and cholesterol in varied phase state. After a structural characterization of this two component model bilayer using X ray reflectivity, we have carried out coherent inelastic neutron scattering to investigate the chain dynamics. Both DMPC cholesterol membranes exhibited much sharper and more pronounced low energy inelastic excitations than a pure DMPC membrane. In the high energy regime above 10meV, the insertion of cholesterol into the membrane was found to shift the position of the inelastic excitation towards values otherwise found in the pure lipids gel phase. Thus, the dissipative collective short range dynamics of the acyl chains is strongly influenced by the presence of cholestero