Entropic Comparison of Atomic-Resolution Electron Tomography of Crystals and Amorphous Materials.

Electron tomography bears promise for widespread determination of the three-dimensional arrangement of atoms in solids. However, it remains unclear whether methods successful for crystals are optimal for amorphous solids. Here, we explore the relative difficulty encountered in atomic-resolution tomography of crystalline and amorphous nanoparticles. We define an informational entropy to reveal the inherent importance of low-entropy zone-axis projections in the reconstruction of crystals. In turn, we propose considerations for optimal sampling for tomography of ordered and disordered materials.

Electron-Beam Mapping of Vibrational Modes with Nanometer Spatial Resolution.

We demonstrate that a focused beam of high-energy electrons can be used to map the vibrational modes of a material with a spatial resolution of the order of one nanometer. Our demonstration is performed on boron nitride, a polar dielectric which gives rise to both localized and delocalized electron-vibrational scattering, either of which can be selected in our off-axial experimental geometry. Our experimental results are well supported by our calculations, and should reconcile current controversy regarding the spatial resolution achievable in vibrational mapping with focused electron beams.

Observation of color center peaks in calcium fluoride.

Alkali halides such as calcium fluoride all have color center defects that absorb light in the visible region. Using a moncochromator equipped, aberration corrected, scanning transmission electron microscope (STEM) we recorded spectra showing the time evolution of the generation of F and H centers in calcium fluoride. The final stage of electron beam irradiation is the formation of metallic calcium nanoparticles. High resolution low loss spectra for the Vacuum Ultraviolet region were also recorded.

Fast calculation of the infrared spectra of large biomolecules.

Vibrational spectra of proteins potentially give insight into biologically significant molecular motion and the proportions of different types of secondary structure. Vibrational spectra can be calculated either from normal modes obtained by diagonalizing the mass-weighted Hessian or from the time autocorrelation function derived from molecular dynamics trajectories. The Hessian matrix is calculated from force fields because it is not practical to calculate the Hessian from quantum mechanics for large molecules. As an alternative to molecular dynamics the spectral response can be calculated from a time autocorrelation derived from numerical solution of the harmonic equations of motion, resulting in calculations at least 4 times faster. Because the calculation also scales linearly with number of atoms, N, it is faster than normal-mode calculations that scale as N (3) for proteins with more then 4,700 atoms. Using this method it is practical to perform all-atom calculations for large biological systems, for example viral capsids, with the order of 10(5) atoms.

Partial rootzone drying increases water-use efficiency of lemon Fino 49 trees independently of root-to-shoot ABA signalling.

To determine whether irrigation strategy altered the sensitivity of Citrus leaf gas exchange to soil, plant and atmospheric variables, mature (16-year-old) Fino 49 lemon trees (Citrus limon (L.) Burm. fil. grafted on Citrus macrophylla Wester) were exposed to three irrigation treatments: control (irrigated with 100% of crop potential evapotranspiration, ETc), deficit irrigation (DI) and partial rootzone drying (PRD) treatments,which received 75% ETc during the period of highest evaporative demand and 50% ETc otherwise. Furthermore, to assess the physiological significance of root-to-shoot ABA signalling, the seasonal dynamics of leaf xylem ABA concentration ([X-ABA]leaf) were evaluated over two soil wetting-drying cycles during a 2-week period in summer. Although stomatal conductance (gs) declined with increased leaf-to-air vapour pressure deficit (LAVPD), lower leaf water potential and soil water availability, [X-ABA]leaf was only related to stomatal closure in well irrigated trees under moderate (<2.5kPa) atmospheric vapour pressure deficit (VPD). Differences in [X-ABA]leaf were not detected between treatments either before or immediately after (<12h) rewatering the dry side of PRD trees. Leaf water potential was higher in control trees, but decreased similarly in all irrigation treatments as daily LAVPD increased. In contrast, DI and PRD trees showed lower stomatal sensitivity to LAVPD than control trees. Although DI and PRD decreased stomatal conductance and photosynthesis, these treatments did not significantly decrease yield, but PRD increased crop water use efficiency (WUE) by 83% compared with control trees. Thus PRD-induced enhancement of crop WUE in a semiarid environment seems to involve physiological mechanisms other than increased [X-ABA]leaf.

Dose, exposure time and resolution in serial X-ray crystallography.

The resolution of X-ray diffraction microscopy is limited by the maximum dose that can be delivered prior to sample damage. In the proposed serial crystallography method, the damage problem is addressed by distributing the total dose over many identical hydrated macromolecules running continuously in a single-file train across a continuous X-ray beam, and resolution is then limited only by the available molecular and X-ray fluxes and molecular alignment. Orientation of the diffracting molecules is achieved by laser alignment. The incident X-ray fluence (energy/area) is evaluated that is required to obtain a given resolution from (i) an analytical model, giving the count rate at the maximum scattering angle for a model protein, (ii) explicit simulation of diffraction patterns for a GroEL-GroES protein complex, and (iii) the spatial frequency cut-off of the transfer function following iterative solution of the phase problem, and reconstruction of an electron density map in the projection approximation. These calculations include counting shot noise and multiple starts of the phasing algorithm. The results indicate counting time and the number of proteins needed within the beam at any instant for a given resolution and X-ray flux. An inverse fourth-power dependence of exposure time on resolution is confirmed, with important implications for all coherent X-ray imaging. It is found that multiple single-file protein beams will be needed for sub-nanometer resolution on current third-generation synchrotrons, but not on fourth-generation designs, where reconstruction of secondary protein structure at a resolution of 7 A should be possible with relatively short exposures.

Morphology of crystals in calcium oxalate monohydrate kidney stones.

Both scanning electron microscopy and atomic force microscopy (AFM) have shown that calcium oxalate monohydrate kidney stones are made up from arrangements of sub micron crystals. The purpose of this investigation was to determine the morphology of these crystals which was obscured by the presence of organic matrix in our earlier study. Sections of stones were treated to remove the protein component of the matrix and then imaged using AFM. Images obtained after proteolysis show that the crystals are in the form of plates stacked on (100) surfaces. These results were confirmed by scanning electron microscopy observations from selected regions of calcium oxalate kidney stone surfaces. The observed crystal sizes are consistent with both the known matrix mass fraction and crystallite growth in the passage through the collecting duct.

Can high-angle annular dark field scattering be represented by a local operator?

High-angle annular dark field imaging has become an invaluable technique for recording atomic resolution STEM images. Many analyses of high-angle annular dark field images assume that the signal is the result of a local scattering operator and can be represented as a simple convolution of a probe function with a set of atomic columns. The apparent simplicity of the technique and the straightforward increase in signal with atomic number have lead to the belief that it is possible to quantify impurity concentrations at atomic column resolution. The limitations in these assumptions are examined on the basis of approximations starting from a complete theory for high-angle scattering based on multi phonon excitations. Not surprisingly, the accuracy of the local scattering operator approximation improves as the inner cut-off angle is increased.

Schemes to determine the crystal potential under dynamical conditions using voltage variation.

Charge densities and crystal structures can be determined routinely from X-ray diffraction as X-ray scattering is relatively weak and single scattering can be assumed. The strong dynamical diffraction of high-energy electrons has prevented electron diffraction from being used in the same way. Dynamical diffraction describes both the propagation of the Bragg diffracted wave in the crystal and the scattering by the crystal potential. The balance between these two processes changes as a function of voltage due to relativistic effects. The difference in diffracted intensities recorded at two voltages is shown to be directly proportional to the crystal potential. This is confirmed by calculations using first-order perturbation theory which show negligible differences compared to exact calculation. It should therefore be possible to use differences in intensity measured as a function of voltage to determine the crystal potential directly. If the full complex wave function is available, then there is a particularly simple procedure to recover the potential, even under dynamical conditions.

Evidence for aggregation in oxalate stone formation: atomic force and low voltage scanning electron microscopy.

PURPOSE: The aim of this investigation was to differentiate between aggregation and crystal growth by studying the structure of oxalate stones at high spatial resolution using recently developed microscopy techniques. MATERIALS AND METHODS: Sections from 6 complete human oxalate stones and 4 stone fragments were prepared by ultramicrotomy and examined by both low voltage scanning electron microscopy and atomic force microscopy. RESULTS: The scanning electron microscopy showed lamellar structures up to 10 microns. in size, consistent with previous results, and provided evidence that these structures were composed of smaller particles. The atomic force microscopy clearly showed arrays of the small particles, whose size varied between 500A and 2800A. CONCLUSION: Our images suggest that an ordered aggregation of small crystallites is responsible for oxalate stone formation.

The thickness determination of organic crystals under low dose conditions using electron energy loss spectroscopy.

In the 3-dimensional (3-D) reconstruction of protein crystals with variable thicknesses the electron images and diffraction patterns can only be merged if the crystal thickness is known. Measurement of the thickness using the ratio of the number of inelastically scattered electrons to the number of electrons in the zero loss peak can be accomplished with parallel electron energy loss spectrometry (PEELS). A theoretical analysis of the accuracy of the technique on paraffin crystals of different thicknesses is presented. Our experimental studies with paraffin crystals show the feasibility of measuring a single layer of 47A with good accuracy under low dose and low temperature conditions. A simple experimental apparatus is proposed to obtain thicknesses from small regions of unstained protein crystals prior to collecting the 3-D data sets from the unexposed area of the same crystal.