Search

Here we present two robotic sample changers integrated into the experimental stations for the macromolecular crystallography (MX) beamlines AMX and FMX, and the biological small-angle scattering (bioSAXS) beamline LiX. They enable fully automated unattended data collection and remote access to the beamlines. The system designs incorporate high-throughput, versatility, high-capacity, resource sharing and robustness. All systems are centered around a six-axis industrial robotic arm coupled with a force torque sensor and in-house end effectors (grippers). They have the same software architecture and the facility standard EPICS-based BEAST alarm system. The MX system is compatible with SPINE bases and Unipucks. It comprises a liquid nitrogen dewar holding 384 samples (24 Unipucks) and a stay-cold gripper, and utilizes machine vision software to track the sample during operations and to calculate the final mount position on the goniometer. The bioSAXS system has an in-house engineered sample storage unit that can hold up to 360 samples (20 sample holders) which keeps samples at a user-set temperature (277 K to 300 K). The MX systems were deployed in early 2017 and the bioSAXS system in early 2019.

Two new macromolecular crystallography (MX) beamlines at the National Synchrotron Light Source II, FMX and AMX, opened for general user operation in February 2017 [Schneider et al. (2013). J. Phys. Conf. Ser. 425, 012003; Fuchs et al. (2014). J. Phys. Conf. Ser. 493, 012021; Fuchs et al. (2016). AIP Conf. Proc. SRI2015, 1741, 030006]. FMX, the micro-focusing Frontier MX beamline in sector 17-ID-2 at NSLS-II, covers a 5–30 keV photon energy range and delivers a flux of 4.0 × 1012 photons s−1 at 1 Å into a 1 µm × 1.5 µm to 10 µm × 10 µm (V × H) variable focus, expected to reach 5 × 1012 photons s−1 at final storage-ring current. This flux density surpasses most MX beamlines by nearly two orders of magnitude. The high brightness and microbeam capability of FMX are focused on solving difficult crystallographic challenges. The beamline's flexible design supports a wide range of structure determination methods – serial crystallography on micrometre-sized crystals, raster optimization of diffraction from inhomogeneous crystals, high-resolution data collection from large-unit-cell crystals, room-temperature data collection for crystals that are difficult to freeze and for studying conformational dynamics, and fully automated data collection for sample-screening and ligand-binding studies. FMX's high dose rate reduces data collection times for applications like serial crystallography to minutes rather than hours. With associated sample lifetimes as short as a few milliseconds, new rapid sample-delivery methods have been implemented, such as an ultra-high-speed high-precision piezo scanner goniometer [Gao et al. (2018). J. Synchrotron Rad. 25, 1362–1370], new microcrystal-optimized micromesh well sample holders [Guo et al. (2018). IUCrJ, 5, 238–246] and highly viscous media injectors [Weierstall et al. (2014). Nat. Commun. 5, 3309]. The new beamline pushes the frontier of synchrotron crystallography and enables users to determine structures from difficult-to-crystallize targets like membrane proteins, using previously intractable crystals of a few micrometres in size, and to obtain quality structures from irregular larger crystals.

The instrumentation and methods available for collecting almost simultaneous single-crystal electronic absorption correlated with X-ray diffraction data at NSLS beamline X26-C are reviewed, as well as a very brief outline of its Raman spectroscopy capability., The research philosophy and new capabilities installed at NSLS beamline X26-C to support electronic absorption and Raman spectroscopies coupled with X-ray diffraction are reviewed. This beamline is dedicated full time to multidisciplinary studies with goals that include revealing the relationship between the electronic and atomic structures in macromolecules. The beamline instrumentation has been fully integrated such that optical absorption spectra and X-ray diffraction images are interlaced. Therefore, optical changes induced by X-ray exposure can be correlated with X-ray diffraction data collection. The installation of Raman spectroscopy into the beamline is also briefly reviewed. Data are now routinely generated almost simultaneously from three complementary types of experiments from the same sample. The beamline is available now to the NSLS general user population.

Micro-SRIXE (synchrotron-radiation-induced X-ray emission) and micro-XAS (X-ray absorption spectroscopy) were used to probe the uptake of exogenous metals by cells. The high flux and the sub-micron resolution of the hard X-ray microprobe, offer the experimenter the ability to obtain highly sensitive spatial and structural information of cellular elements. In this work the uptake of carcinogenic Cr(VI) was compared with that of a relatively non-toxic Cr(III) complex by micro-SRIXE mapping of whole cells. High intracellular Cr concentrations were observed in Cr(VI)-treated cells, while no significant Cr uptake was observed for Cr(III)-treated cells, as is consistent with uptake studies performed by other techniques. Micro-XANES analysis of Cr(V)- and Cr(VI)-treated cells showed that the predominant oxidation product following cellular metabolism was Cr(III). As shown by X-ray microscopic analysis of thin-sectioned cells, however, the reduction of Cr(VI) to Cr(III) did not occur at a fast enough rate to exclude Cr entry into the cell nucleus.

Effects of sample treatment must be well understood to avoid artifacts during analysis. The effect of drying and storage was examined on nine medium- to high-Mn aerobic soils using extractable Mn(II), the Cr Oxidation Test, estimated soluble C, and XANES spectroscopy. Long-term storage (430 d) at 3°C had little effect on the Cr test. Air drying at room temperature (25°C ± 3°C) caused a drop in the Cr test within 24 h, with a further decline to as low as 45% of the original after 72 h, and less than 2% after 264 d. Extractability of Mn with pH 4.8 NH 4 OAc increased nearly linearly over the same time period from 0.2 mmol kg -1 to as high as 2.3 mmol kg -1 , Increases in the absorbance of the extract at 360 nm, an estimate of soluble C, were well correlated within each soil with the increase in Mn(II). Pretreatments to remove soluble organics did not cause any increases in the Cr test of dried samples. Therefore, the loss of Cr oxidizing ability appears to be due to reduction of the oxides, not because of increased reduction of the Cr(VI) formed. No changes in XANES spectra were found after short-term air drying at room temperature, but in the three samples examined after 428 d of drying, the main-edge position had a downward shift of about 1.5 eV, indicating reduction. These results confirm previous findings that studies on the reactivity of soil Mn oxides need to avoid sample drying.

The examination of soils by x-ray absorption near-edge structure (XANES) spectroscopy can provide valuable dues for determining the structure and composition of Mn oxides. XANES experiments can be conducted at ambient temperatures using field-moist samples. This feature is attractive in the study of soil Mn because of the well-documented effects of sample drying on reactivity. The present study was undertaken to determine whether the analysis of field-moist soils by XANES spectroscopy has an effect on the oxidation state of the Mn. Repeated scans were taken on the same spot using seven medium-to high-Mn soils. In all 10 experiments with untreated soils, the main absorption edge dropped in relative energy in successive scans, indicating reduction of Mn. The change in eV was as high as 1.7 eV after 150 min of exposure time. After a series of scans showing reduction, small movement of the sample in the x-ray beam and reexamination showed no reduction, indicating that the effect was limited to the x-ray spot. Modeling the change with Mn(II) was successful and showed increases in reduced Mn between 6 and 20% after three to five repeated scans. The slopes of eV change vs. exposure time were remarkably similar in most experiments, even with large differences in the density of photons between runs. Comparisons of initial moist scans with those of recently air-dried samples found little difference, suggesting minimal reduction in a single XANES scan under our experimental conditions.

X-ray fluorescence spectroscopy is demonstrated here as a novel, element-specific detector for capillary electrophoresis. Monochromatic 10 keV X-rays from a synchrotron light source are used to excite core electrons, causing emission of characteristic Kalpha X-ray fluorescence (XRF) lines. Using this technique, XRF energies provide elemental identification, while XRF intensities can be used to quantitate the metal composition of each eluent. An X-ray transparent polymer coupling is used to create a window for the on-line, X-ray detection. This coupling contributes no measurable extra-column variance, and electrophoretic mobilities for the metal complexes used as model solutes are highly reproducible. The combination of XRF detection with capillary electrophoresis (CE-XRF) creates the first on-line detection system that is element-specific, nondestructive, and directly applicable to a broad range of applications including nonelectroactive species. CE-XRF is successfully demonstrated here for high binding-constant complexes of Fe(III), Co(II), Cu(II), and Zn(II). Within a single injection, electropherograms are obtained for each element of interest, with the element identity obtained directly from the emission energy. In contrast with ICPMS, this detection technique is directly on-line and does not require volatilization of the eluent. As a result, element-specific detection is not limited by the sample or the buffer volatility or atomization efficiency. Simultaneous XRF and UV absorbance detection can be used to provide an on-line determination of metal/chelate ratios. Although XRF detection limits are presently only in the 0.1 mM (0.5 ng) range, both collection geometry and incident intensity have yet to be optimized. Further optimization is expected to enhance this detection limit by another 2-3 orders of magnitude. As a result, the advent of XRF detection combined with the separating power of CE presents new possibilities for on-line, element-specific analysis.

Mechanistic understanding of trace element cycling between surface waters and sediments can require highly spatially resolved information at water−sediment boundaries not obtainable by conventional sampling and analytical methods. This study demonstrates the application of micro-X-ray absorption near-edge structure (micro-XANES) spectroscopy for obtaining such direct, in-situ, spatially and temporally resolved information for selenium. The experiments determine the fate of soluble Se(VI) ponded over water-saturated oxidizing versus reducing sediments. Scanning micro-XANES analyses showed Se(VI) removal from ponded waters by diffusion into sediments and by conversion to Se(0) in the reducing sediment. Zones of accumulation of insoluble Se(0) were heterogeneously distributed. These results show that the scale over which volume averaging of chemical species is performed at sedi ment−water boundaries must be selected carefully and that the spatial resolution offered by micro-XANES is well-suited for these typ...

A high-flux insertion device and beamline for macromolecular crystallography has been built at the National Synchrotron Light Source (NSLS) that employs a mini-gap undulator source developed by the NSLS. The mini-gap undulator at beamline X29 is a hybrid-magnet device of period 12.5 mm operating at proven gaps of 3.3-10 mm. The beamline provides hard X-rays for macromolecular crystallography experiments from the second and third harmonics over an energy range of 5-15 keV. The X-ray optics is designed to deliver intense and highly collimated X-rays. Horizontal focusing is achieved by a cryogenically cooled sagittally focusing double-crystal monochromator with approximately 4.1:1 demagnification. A vertical focusing mirror downstream of the monochromator is used for harmonic rejection and vertical focusing. The experimental station hosts an Area Detector Systems Quantum 315 CCD detector with 2.2 s readout time between exposures and Crystal Logic goniostat for crystal rotation and detector positioning. An auto-mounter crystal changer has been installed to facilitate the high-throughput data collection required by the major users, which includes structural genomics projects and the Macromolecular Crystallography Research Resource mail-in program. X29 is 10(3) times brighter than any existing bending-magnet beamline at NSLS with an actual flux of 2.5 x 10(11) photons s(-1) through a 0.12 mm square aperture at 11.271 keV.

The oxidation state of intracellular chromium has been determined directly in mammalian lung cells exposed to mutagenic and carcinogenic chromium compounds. Microprobe X-ray absorption spectroscopy (XAS) experiments on single V79 Chinese hamster lung cells showed that Cr(VI) and Cr(V) complexes were reduced completely (>90%) to Cr(III) within 4 h of exposure of the cells. This result provides direct evidence for the hypothesis that these genotoxic oxidants react rapidly with intracellular reductants.