A correction procedure for secondary scattering contributions from windows in small-angle X-ray scattering and ultra-small-angle X-ray scattering.

This article describes a correction procedure for the removal of indirect background contributions to measured small-angle X-ray scattering patterns. The high scattering power of a sample in the ultra-small-angle region may serve as a secondary source for a window placed in front of the detector. The resulting secondary scattering appears as a sample-dependent background in the measured pattern that cannot be directly subtracted. This is an intricate problem in measurements at ultra-low angles, which can significantly reduce the useful dynamic range of detection. Two different procedures are presented to retrieve the real scattering profile of the sample.

Recent advances in synchrotron scattering methods for probing the structure and dynamics of colloids.

Recent progress in synchrotron based X-ray scattering methods applied to colloid science is reviewed. An important figure of merit of these techniques is that they enable in situ investigations of colloidal systems under the desired thermophysical and rheological conditions. An ensemble averaged simultaneous structural and dynamical information can be derived albeit in reciprocal space. Significant improvements in X-ray source brilliance and advances in detector technology have overcome some of the limitations in the past. Notably coherent X-ray scattering techniques have become more competitive and they provide complementary information to laboratory based real space methods. For a system with sufficient scattering contrast, size ranges from nm to several µm and time scales down to µs are now amenable to X-ray scattering investigations. A wide variety of sample environments can be combined with scattering experiments further enriching the science that could be pursued by means of advanced X-ray scattering instruments. Some of these recent progresses are illustrated via representative examples. To derive quantitative information from the scattering data, rigorous data analysis or modeling is required. Development of powerful computational tools including the use of artificial intelligence have become the emerging trend.

Orthotropic organization of a cellulose nanocrystal suspension realized via the combined action of frontal ultrafiltration and ultrasound as revealed by in situ SAXS.

HYPOTHESIS: Rodlike cellulose nanocrystals (CNCs) exhibit significant potential as building blocks for creating uniform, sustainable materials. However, a critical hurdle lies in the need to enhance existing or devise novel processing that provides improved control over the alignment and arrangement of CNCs across a wide spatial range. Specifically, the challenge is to achieve orthotropic organization in a single-step processing, which entails creating non-uniform CNC orientations to generate spatial variations in anisotropy. EXPERIMENTS: A novel processing method combining frontal ultrafiltration (FU) and ultrasound (US) has been developed. A dedicated channel-cell was designed to simultaneously generate (1) a vertical acoustic force thanks to a vibrating blade at the top and (2) a transmembrane pressure force at the bottom. Time-resolved in situ small-angle X-ray scattering permitted to probe the dynamical structural organization/orientation of CNCs during the processing. FINDINGS: For the first time, a typical three-layer orthotropic structure that resembles the articular cartilage organization was achieved in one step during the FU/US process: a first layer composed of CNCs having their director aligned parallel to the horizontal membrane surface, a second intermediate isotropic layer, and a third layer of CNCs with their director vertically oriented along the direction of US wave propagation direction.

Dynamics of anisotropic colloidal systems: What to choose, DLS, DDM or XPCS?

Investigation of the dynamics of colloids in bulk can be hindered by issues such as multiple scattering and sample opacity. These challenges are exacerbated when dealing with inorganic materials. In this study, we employed a model system of Akaganeite colloidal rods to assess three leading dynamics measurement techniques: 3D-(depolarized) dynamic light scattering (3D-(D)DLS), polarized-differential dynamic microscopy (P-DDM), and x-ray photon correlation spectroscopy (XPCS). Our analysis revealed that the translational and rotational diffusion coefficients captured by these methods show a remarkable alignment. Additionally, by examining the q-ranges and maximum volume fractions for each approach, we offer insights into the best technique for investigating the dynamics of anisotropic systems at the colloidal scale.

Improvement of ultra-small-angle XPCS with the Extremely Brilliant Source.

Recent technical developments and the performance of the X-ray photon correlation spectroscopy (XPCS) method over the ultra-small-angle range with the Extremely Brilliant Source (EBS) at the ESRF are described. With higher monochromatic coherent photon flux (∼1012 photons s-1) provided by the EBS and the availability of a fast pixel array detector (EIGER 500K detector operating at 23000 frames s-1), XPCS has become more competitive for probing faster dynamics in relatively dilute suspensions. One of the goals of the present development is to increase the user-friendliness of the method. This is achieved by means of a Python-based graphical user interface that enables online visualization and analysis of the processed data. The improved performance of XPCS on the Time-Resolved Ultra-Small-Angle X-ray Scattering instrument (ID02 beamline) is demonstrated using dilute model colloidal suspensions in several different applications.

Orientational ordering and assembly of silica-nickel Janus particles in a magnetic field.

The orientation ordering and assembly behavior of silica-nickel Janus particles in a static external magnetic field were probed by ultra small-angle X-ray scattering (USAXS). Even in a weak applied field, the net magnetic moments of the individual particles aligned in the direction of the field, as indicated by the anisotropy in the recorded USAXS patterns. X-ray photon correlation spectroscopy (XPCS) measurements on these suspensions revealed that the corresponding particle dynamics are primarily Brownian diffusion [Zinn, Sharpnack & Narayanan (2023). Soft Matter, 19, 2311-2318]. At higher fields, the magnetic forces led to chain-like configurations of particles, as indicated by an additional feature in the USAXS pattern. A theoretical framework is provided for the quantitative interpretation of the observed anisotropic scattering diagrams and the corresponding degree of orientation. No anisotropy was detected when the magnetic field was applied along the beam direction, which is also replicated by the model. The method presented here could be useful for the interpretation of oriented scattering patterns from a wide variety of particulate systems. The combination of USAXS and XPCS is a powerful approach for investigating asymmetric colloidal particles in external fields.

Dependence of myosin filament structure on intracellular calcium concentration in skeletal muscle.

Contraction of skeletal muscle is triggered by an increase in intracellular calcium concentration that relieves the structural block on actin-binding sites in resting muscle, potentially allowing myosin motors to bind and generate force. However, most myosin motors are not available for actin binding because they are stabilized in folded helical tracks on the surface of myosin-containing thick filaments. High-force contraction depends on the release of the folded motors, which can be triggered by stress in the thick filament backbone, but additional mechanisms may link the activation of the thick filaments to that of the thin filaments or to intracellular calcium concentration. Here, we used x-ray diffraction in combination with temperature-jump activation to determine the steady-state calcium dependence of thick filament structure and myosin motor conformation in near-physiological conditions. We found that x-ray signals associated with the perpendicular motors characteristic of isometric force generation had almost the same calcium sensitivity as force, but x-ray signals associated with perturbations in the folded myosin helix had a much higher calcium sensitivity. Moreover, a new population of myosin motors with a longer axial periodicity became prominent at low levels of calcium activation and may represent an intermediate regulatory state of the myosin motors in the physiological pathway of filament activation.

Fate of Magnetic Nanoparticles during Stimulated Healing of Thermoplastic Elastomers.

We have investigated the heating mechanism in industrially relevant, multi-block copolymers filled with Fe nanoparticles and subjected to an oscillatory magnetic field that enables polymer healing in a contactless manner. While this procedure aims to extend the lifetime of a wide range of thermoplastic polymers, repeated or prolonged stimulus healing is likely to modify their structure, mechanics, and ability to heat, which must therefore be characterized in depth. In particular, our work sheds light on the physical origin of the secondary heating mechanism detected in soft systems subjected to magnetic hyperthermia and triggered by copolymer chain dissociation. In spite of earlier observations, the origin of this additional heating remained unclear. By using both static and dynamic X-ray scattering methods (small-angle X-ray scattering and X-ray photon correlation spectroscopy, respectively), we demonstrate that beyond magnetic hysteresis losses, the enormous drop of viscosity at the polymer melting temperature enables motion of nanoparticles that generates additional heat through friction. Additionally, we show that applying induction heating for a few minutes is found to magnetize the nanoparticles, which causes them to align in dipolar chains and leads to nonmonotonic translational dynamics. By extrapolating these observations to rotational dynamics and the corresponding amount of heat generated through friction, we not only clarify the origin of the secondary heating mechanism but also rationalize the presence of a possible temperature maximum observed during induction heating.

Small-angle X-ray scattering in the era of fourth-generation light sources.

Recently, fourth-generation synchrotron sources with several orders of magnitude higher brightness and higher degree of coherence compared with third-generation sources have come into operation. These new X-ray sources offer exciting opportunities for the investigation of soft matter and biological specimens by small-angle X-ray scattering (SAXS) and related scattering methods. The improved beam properties together with the advanced pixel array detectors readily enhance the angular resolution of SAXS and ultra-small-angle X-ray scattering in the pinhole collimation. The high degree of coherence is a major boost for the X-ray photon correlation spectroscopy (XPCS) technique, enabling the equilibrium dynamics to be probed over broader time and length scales. This article presents some representative examples illustrating the performance of SAXS and XPCS with the Extremely Brilliant Source at the European Synchrotron Radiation Facility. The rapid onset of radiation damage is a significant challenge with the vast majority of samples, and appropriate protocols need to be adopted for circumventing this problem.

High unimer concentration and asymmetric bilayer observed in small unilamellar vesicles of mixed surfactants TDMAO/LiPFO.

The mixed surfactant system of tetradecyldimethylamine oxide (TDMAO) and lithium perfluorooctanoate (LiPFO) is known to spontaneously self-assemble into well-defined small unilamellar vesicles. For a quantitative analysis of small-angle x-ray scattering on this model system, we complemented the measurements with densitometry, conductimetry, and contrast-variation small-angle neutron scattering. The analysis points to two main findings: first, the vesicles formed to contain a much higher mole fraction (0.61-0.64) of TDMAO than the bulk sample (0.43) and predicted by Regular Solution Theory (RST, 0.46). In consequence, the unimer concentration of LiPFO is more than 5 times higher than predicted by RST. Second, the vesicle bilayer is asymmetric with a higher fraction of LiPFO on the outside. These findings on a model system should be of broader relevance for the understanding of similar mixed surfactant vesicle systems and thereby also be of importance for their use in a number of applications.

Titin activates myosin filaments in skeletal muscle by switching from an extensible spring to a mechanical rectifier.

Titin is a molecular spring in parallel with myosin motors in each muscle half-sarcomere, responsible for passive force development at sarcomere length (SL) above the physiological range (>2.7 µm). The role of titin at physiological SL is unclear and is investigated here in single intact muscle cells of the frog (Rana esculenta), by combining half-sarcomere mechanics and synchrotron X-ray diffraction in the presence of 20 µM para-nitro-blebbistatin, which abolishes the activity of myosin motors and maintains them in the resting state even during activation of the cell by electrical stimulation. We show that, during cell activation at physiological SL, titin in the I-band switches from an SL-dependent extensible spring (OFF-state) to an SL-independent rectifier (ON-state) that allows free shortening while resisting stretch with an effective stiffness of ~3 pN nm-1 per half-thick filament. In this way, I-band titin efficiently transmits any load increase to the myosin filament in the A-band. Small-angle X-ray diffraction signals reveal that, with I-band titin ON, the periodic interactions of A-band titin with myosin motors alter their resting disposition in a load-dependent manner, biasing the azimuthal orientation of the motors toward actin. This work sets the stage for future investigations on scaffold and mechanosensing-based signaling functions of titin in health and disease.

Dynamics of magnetic Janus colloids studied by ultra small-angle X-ray photon correlation spectroscopy.

The orientation behavior and the translational dynamics of spherical magnetic silica-nickel Janus colloids in an external magnetic field have been studied by small-angle X-ray scattering and X-ray photon correlation spectroscopy at ultra small-angles. For weak applied fields and at low volume fractions, the particle dynamics is dominated by Brownian motion even though the net magnetic moments of the individual particles are aligned in the direction of the field as indicated by the anisotropy in the small-angle scattering patterns. For higher fields the magnetic forces result in more complex structural changes with nickel caps of Janus particles pointing predominantly along the applied magnetic field. The alignment ultimately leads to chain-like configurations and the intensity-intensity autocorrelation functions, g2(q,t), show a second slower decay which becomes more pronounced at higher volume fractions. A direction dependent analysis of g2(q,t) revealed a faster than exponential decay perpendicular to the field which is related to the sedimentation of magnetically ordered domains. The corresponding velocity fluctuations could be decoupled from the diffusion of particles by decomposing g2(q,t) into advective and diffusive contributions. Finally, the particle dynamics becomes anisotropic at higher volume fractions and strong magnetic fields. The derived translational diffusion coefficients indicate slower particle dynamics perpendicular to the field as compared to the parallel direction.

Specific analysis of highly absorbing nanoporous powder by small-angle X-ray scattering.

The characterization of nanoporous powders of highly absorbing compounds by small-angle X-ray scattering (SAXS) involves overcoming several difficulties before quantitative information related to the porous texture, such as the specific surface and the porous volume, can be derived. In this article, first, the contribution of the grain facet reflectivity and scattering from the bulk of a grain with the density of ThO2, a highly absorbing material, were calculated. Microporous ThO2 powder having micrometric grain size was characterized, in which the scattering signal is predominant. A high-resolution synchrotron instrument was used in order to cover a wider q range and minimize the absorption effect, and the results were compared with those obtained using a laboratory X-ray source. Concerning the absorption problem existing with a laboratory X-ray source, a new and robust experimental method was proposed to correctly determine the scattering intensity of the highly absorbing granular samples on an absolute scale. This method allows one to calculate accurately the porous volume and the specific surface via Porod's law and the invariant using a laboratory SAXS instrument. This last result opens new perspectives for the characterization of the volume and the specific surface of highly absorbing actinide oxide powders.

Performance of the time-resolved ultra-small-angle X-ray scattering beamline with the Extremely Brilliant Source.

The new technical features and enhanced performance of the ID02 beamline with the Extremely Brilliant Source (EBS) at the ESRF are described. The beamline enables static and kinetic investigations of a broad range of systems from ångström to micrometre size scales and down to the sub-millisecond time range by combining different small-angle X-ray scattering techniques in a single instrument. In addition, a nearly coherent beam obtained in the high-resolution mode allows multispeckle X-ray photon correlation spectroscopy measurements down to the microsecond range over the ultra-small- and small-angle regions. While the scattering vector (of magnitude q) range covered is the same as before, 0.001 ≤ q ≤ 50 nm-1 for an X-ray wavelength of 1 Å, the EBS permits relaxation of the collimation conditions, thereby obtaining a higher flux throughput and lower background. In particular, a coherent photon flux in excess of 1012 photons s-1 can be routinely obtained, allowing dynamic studies of relatively dilute samples. The enhanced beam properties are complemented by advanced pixel-array detectors and high-throughput data reduction pipelines. All these developments together open new opportunities for structural, dynamic and kinetic investigations of out-of-equilibrium soft matter and biophysical systems.

Cooling intact and demembranated trabeculae from rat heart releases myosin motors from their inhibited conformation.

Myosin filament-based regulation supplements actin filament-based regulation to control the strength and speed of contraction in heart muscle. In diastole, myosin motors form a folded helical array that inhibits actin interaction; during contraction, they are released from that array. A similar structural transition has been observed in mammalian skeletal muscle, in which cooling below physiological temperature has been shown to reproduce some of the structural features of the activation of myosin filaments during active contraction. Here, we used small-angle x-ray diffraction to characterize the structural changes in the myosin filaments associated with cooling of resting and relaxed trabeculae from the right ventricle of rat hearts from 39°C to 7°C. In intact quiescent trabeculae, cooling disrupted the folded helical conformation of the myosin motors and induced extension of the filament backbone, as observed in the transition from diastole to peak systolic force at 27°C. Demembranation of trabeculae in relaxing conditions induced expansion of the filament lattice, but the structure of the myosin filaments was mostly preserved at 39°C. Cooling of relaxed demembranated trabeculae induced changes in motor conformation and filament structure similar to those observed in intact quiescent trabeculae. Osmotic compression of the filament lattice to restore its spacing to that of intact trabeculae at 39°C stabilized the helical folded state against disruption by cooling. The myosin filament structure and motor conformation of intact trabeculae at 39°C were largely preserved in demembranated trabeculae at 27°C or above in the presence of Dextran, allowing the physiological mechanisms of myosin filament-based regulation to be studied in those conditions.

High-resolution structural elucidation of extremely swollen lyotropic phases.

Self-assembled lyotropic phases are important in a variety of applications, in particular microemulsions are essential for formulation science. A spectacular situation arises when microemulsions are made to swell by systematically increasing the bending modulus of the surfactant film separating the oil and water regions. In an attempt to realize such extremely swollen microemulsion phases, Peter et al. [Phys. Rev. Lett., 76 (1996) 3866] found a variety of lyotropic phases including a long-range ordered three-dimensional cubic phase over a narrow section of the complex phase diagram of a pseudo-quaternary system composed of decane, brine, octanol, and sodium dodecyl sulfate. In this work, the same region of the phase diagram was reinvestigated using high-resolution small-angle X-ray scattering (HR-SAXS) and rheo-SAXS, which is an important technical aspect for homogenizing the sample and orienting the structural units. Whilst the formation of a swollen two-dimensional hexagonal phase was observed, the structural features of a cubic phase were not detected. The long correlation lengths noted prior were also seen here, over 2000 nm for the hexagonal phase, taken from rheo-SAXS measurements. Based on the measurements covering more than three orders of magnitude in scattering vector, the structure appeared to be an organization of elongated swollen emulsion droplets, which could form an interconnected structure, dense liquid-like order, or further order into a hexagonal morphology with unusually large lattice spacings for a surfactant system.

SAXS/WAXS Investigation of Amyloid-ß(16-22) Peptide Nanotubes.

This brief report presents an X-ray scattering investigation of self-assembled nanotubes formed by a short peptide. X-ray scattering methods enable multiscale structural elucidation of these nanotubes in solution under the same conditions involved in the self-assembly process. In particular, the dimensions of nanotubes and the crystalline organization within their walls can be determined quantitatively. This is illustrated in the case of amyloid-ß(16-22) peptide nanotubes.

Multiscale Structural Elucidation of Peptide Nanotubes by X-Ray Scattering Methods.

This mini-review presents the structural investigations of the self-assembled peptide nanotubes using X-ray scattering techniques. As compared to electron microscopy, scattering methods enable studies of nanotubes in solution under the appropriate physicochemical conditions and probe their formation mechanism. In addition, a combination of X-ray scattering methods allow the elucidation of structural organization from the molecular scale to the dimension of nanotubes.

Breakdown and buildup mechanisms of cellulose nanocrystal suspensions under shear and upon relaxation probed by SAXS and SALS.

The breakdown and buildup mechanisms in concentrated cellulose nanocrystal (CNC) suspensions under shear and during relaxation upon cessation of shear were accessed by small-angle X-ray and light scattering combined with rheometry. The dynamic structural changes over nanometer to micrometer lengthscales were related to the well-known three-regime rheological behavior. In the shear-thinning regime I, the large liquid crystalline domains were progressively fragmented into micrometer-sized tactoids, with their cholesteric axis aligned perpendicular to the flow direction. The viscosity plateau of regime II was associated to a further disruption into submicrometer-sized elongated tactoids oriented along the velocity direction. At high shear rate, regime III corresponded to the parallel flow of individual CNCs along the velocity direction. Upon cessation of flow, the relaxation process occurred through a three-step buildup mechanisms: i) a fast reassembling of the individual CNCs into a nematic-like organization established up to micrometer lengthscales, ii) a slower formation of oriented large cholesteric domains, and iii) their isotropic redistribution.

Dependence of thick filament structure in relaxed mammalian skeletal muscle on temperature and interfilament spacing.

Contraction of skeletal muscle is regulated by structural changes in both actin-containing thin filaments and myosin-containing thick filaments, but myosin-based regulation is unlikely to be preserved after thick filament isolation, and its structural basis remains poorly characterized. Here, we describe the periodic features of the thick filament structure in situ by high-resolution small-angle x-ray diffraction and interference. We used both relaxed demembranated fibers and resting intact muscle preparations to assess whether thick filament regulation is preserved in demembranated fibers, which have been widely used for previous studies. We show that the thick filaments in both preparations exhibit two closely spaced axial periodicities, 43.1 nm and 45.5 nm, at near-physiological temperature. The shorter periodicity matches that of the myosin helix, and x-ray interference between the two arrays of myosin in the bipolar filament shows that all zones of the filament follow this periodicity. The 45.5-nm repeat has no helical component and originates from myosin layers closer to the filament midpoint associated with the titin super-repeat in that region. Cooling relaxed or resting muscle, which partially mimics the effects of calcium activation on thick filament structure, disrupts the helical order of the myosin motors, and they move out from the filament backbone. Compression of the filament lattice of demembranated fibers by 5% Dextran, which restores interfilament spacing to that in intact muscle, stabilizes the higher-temperature structure. The axial periodicity of the filament backbone increases on cooling, but in lattice-compressed fibers the periodicity of the myosin heads does not follow the extension of the backbone. Thick filament structure in lattice-compressed demembranated fibers at near-physiological temperature is similar to that in intact resting muscle, suggesting that the native structure of the thick filament is largely preserved after demembranation in these conditions, although not in the conditions used for most previous studies with this preparation.