A New Method for Dead Time Calibration and a New Expression for Correction of WDS Intensities for Microanalysis.

Observed photon count rates must be corrected for detector dead time effects for accurate quantification, especially at high count rates. We present the "constant k-ratio" method, a new approach for calibrating dead time for wavelength dispersive spectrometers by measuring k-ratios as a function of beam current. The method is based on the observation that for a given emission line at a specific take-off angle and electron beam energy, the intensity ratio from two materials containing the element should remain constant as a function of beam current, if the dead time calibration is accurate. The method has the advantage that it does not rely on the linearity of the beam current picoammeter, yet also allows the analyst to evaluate the picoammeter linearity, another critical parameter in EPMA calibration. By simultaneously comparing k-ratios for all spectrometers, one can also ascertain k-ratio consensus, essential for inter-laboratory comparisons. We also introduce improved dead time expressions and provide best practices on how to perform these instrument calibrations using this new "constant k-ratio" method. These improvements enable quantitative analysis of major and minor elements with high accuracy at high beam currents, simultaneously with trace elements with high sensitivity, for point analyses and X-ray mapping.

Assessing the Accuracy of Mass Attenuation Coefficients for Soft X-ray EPMA.

The use of soft X-rays in electron probe microanalysis (EPMA) has gained renewed interest over the past decades due to the advent of new detector technologies. Because X-ray absorption is the dominant correction for soft X-rays, a reliable set of mass attenuation coefficients (MACs) is needed for accurate composition determination. Although several MAC tabulations cover the soft X-ray range, the accuracy of such tabulations below 1 keV is not firmly established. In this study, we assess the accuracy of MAC tabulations in the soft X-ray region by comparing tabulated values for Be, B, C, N, O, and F Kα X-rays with experimental data available in the literature. We find that the 1993 semi-empirical MAC compilation of Henke et al. [(1993). Low-energy X-ray interaction coefficients: Photoabsorption, scattering, transmission and reflection at E=50-30000 eV, Z=1-92. Atom Data Nucl Data Tables54, 181-342] and the more recent theoretical MAC calculations of Sabbatucci and Salvat [(2016). Theory and calculation of the atomic photoeffect. Rad Phys Chem121, 122-140] perform slightly better than the rest of the considered tabulations. The Sabbatucci-Salvat dataset also provides the best agreement with the few existing experimental measurements for Al L2,3M X-rays.

Electron Probe Microanalysis of Transition Metals using L lines: The Effect of Self-absorption.

Electron microprobe-based quantitative compositional measurement of first-row transition metals using their L$\alpha$ X-ray lines is hampered by, among other effects, self-absorption. This effect, which occurs when a broad X-ray line is located close to a broad absorption edge, is not accounted for by matrix corrections. To assess the error due to neglecting self-absorption, we calculate the L$\alpha$ X-ray intensity emitted from metallic Fe, Ni, Cu, and Zn targets, assuming a Lorentzian profile for the X-ray line and taking into account the energy dependence of the mass absorption coefficient near the absorption edge. We find that calculated X-ray intensities depart increasingly, for increasing electron beam energy, from those obtained assuming a narrow X-ray line and a single fixed absorption coefficient (conventional approach), with a maximum deviation of $\sim$15% for Ni and of $\sim$10% for Fe. In contrast, X-ray intensities calculated for metallic Zn and Cu do not differ significantly from those obtained using the conventional approach. The implications of these results for the analysis of transition-metal compounds by electron probe microanalysis as well as strategies to account for self-absorption effects are discussed.

ϕ(ρz) Distributions in Bulk and Thin Film Samples for EPMA. Part 1: A Modified ϕ(ρz) Distribution for Bulk Materials, Including Characteristic and Bremsstrahlung Fluorescence.

Electron probe microanalysis is a nondestructive technique widely used to determine the elemental composition of bulk samples. This was extended to layered specimens, with the development of appropriate software. The traditional quantification method requires the use of matrix correction procedures based upon models of the ionization depth distribution, the so-called ϕ(ρz) distribution. Most of these models have led to commercial quantification programs but only few of them allow the quantification of layered specimens. Therefore, we developed BadgerFilm, a free open-source thin film program available to the general public. This program implements a documented ϕ(ρz) model as well as algorithms to calculate fluorescence in bulk and thin film samples. Part 1 of the present work aims at describing the operation of the implemented ϕ(ρz) distribution model and validating its implementation against experimental measurements and Monte Carlo simulations on bulk samples. The program has the ability to predict absolute X-ray intensities that can be directly compared to Monte Carlo simulations. We demonstrate that the implemented model works very well for bulk materials. And as will be shown in Part 2, BadgerFilm predictions for thin film specimens are also shown to be in good agreements with experimental and Monte Carlo results.

ϕ(ρz) Distributions in Bulk and Thin-Film Samples for EPMA. Part 2: BadgerFilm: A New Thin-Film Analysis Program.

Since the 1960s, thicknesses and compositions of thin-film specimens have been determined by using the nondestructive technique of electron probe microanalysis. This approach, refined in the 1990s, is based upon models of the ionization depth distribution, the so-called ϕ(ρz) distribution, and is capable of analyzing layered specimens. Most of these quantification models have led to commercial programs. However, these programs may have possible limitations: some may not be directly compatible with modern computers, they often are "black boxes" making it difficult to assess the reliability of the results, and they are sometimes expensive enough to restrain many labs from purchasing them. We present a user-friendly, free, open-source program, BadgerFilm, implementing a documented ϕ(ρz) model and algorithms allowing the quantification of stratified samples. The program has the ability to calculate absolute X-ray intensities that can be directly compared with Monte Carlo simulations. We give a detailed explanation for the operation of the employed ϕ(ρz) model in thin films. A wide range of detailed Monte Carlo simulations and experimental data have been used to evaluate and validate the accuracy of the implemented algorithms. BadgerFilm demonstrated excellent quantification results for the films and in many cases for the substrates.

Biomineral armor in leaf-cutter ants.

Although calcareous anatomical structures have evolved in diverse animal groups, such structures have been unknown in insects. Here, we report the discovery of high-magnesium calcite [CaMg(CO3)2] armor overlaying the exoskeletons of major workers of the leaf-cutter ant Acromyrmex echinatior. Live-rearing and in vitro synthesis experiments indicate that the biomineral layer accumulates rapidly as ant workers mature, that the layer is continuously distributed, covering nearly the entire integument, and that the ant epicuticle catalyzes biomineral nucleation and growth. In situ nanoindentation demonstrates that the biomineral layer significantly hardens the exoskeleton. Increased survival of ant workers with biomineralized exoskeletons during aggressive encounters with other ants and reduced infection by entomopathogenic fungi demonstrate the protective role of the biomineral layer. The discovery of biogenic high-magnesium calcite in the relatively well-studied leaf-cutting ants suggests that calcareous biominerals enriched in magnesium may be more common in metazoans than previously recognized.

Magnesium isotope analysis of olivine and pyroxene by SIMS: Evaluation of matrix effects.

The performance of multi-collector secondary ion mass spectrometry (MC-SIMS) for Mg isotope ratio analysis was evaluated using 17 olivine and 5 pyroxene reference materials (RMs). The Mg isotope composition of these RMs was accurately and precisely determined by multi-collector inductively coupled plasma mass spectrometry (MC-ICP-MS), and these measured isotope ratios were used to evaluate SIMS instrumental mass bias as a function of the forsterite (Fo) content of olivine. The magnitude of the Mg isotope matrix effects were ~3‰ in δ25Mg, and are a complex function of olivine Fo content, that ranged from Fo59.3 to Fo100. In addition to these Mg isotope matrix effects, Si+ ion yields and Mg+/Si+ ion ratios varied as a complex function of the Fo content of the olivine RMs. For example, Si+ ion yields varied by ~33%. Based on the observations, we propose instrumental bias correction procedures for SIMS Mg isotope analysis of olivine using a combination of Mg+/Si+ ratios and Fo content of olivine. Using this correction method, the accuracy of δ25Mg analyses is 0.3‰, except for analysis of olivine with Fo86-88 where instrumental biases and Mg+/Si+ ratios change dramatically with Fo content, making it more difficult to assess the accuracy of Mg isotope ratio measurements by SIMS over this narrow range of Fo content. Five pyroxene RMs (3 orthopyroxenes and 2 clinopyroxenes) show smaller ranges of instrumental bias (~1.4‰ in δ25Mg) as compared to the olivine RMs. The instrumental bias for the 3 orthopyroxene RMs do not define a linear relationship with respect to enstatite (En) content, that ranged from En85.5 -96.3. The clinopyroxene RMs have similar En and wollastonite (Wo) contents but have δ25Mg values that differ by 0.5‰ relative to their δ25Mg values determined by MC-ICP-MS. These results indicate that additional factors (e.g., minor element abundances) likely contribute to SIMS instrumental mass fractionation. In order to better correct for these SIMS matrix effects, additional pyroxene RMs with various chemical compositions and known Mg isotope ratios are needed.

Quantitative Measurement of Iron-Silicides by EPMA Using the Fe Lα and Lß X-ray Lines: A New Twist to an Old Approach.

The recent availability of Schottky-type field emission electron microprobes provides incentive to consider analyzing micrometer-sized features. Yet, to quantify sub-micrometer-sized features, the electron interaction volume must be reduced by decreasing accelerating voltage. However, the K lines of the transition elements (e.g., Fe) then cannot be excited, and the L lines must be used. The Fe Lα1,2 line is the most intense of the L series but bonding effects change its atomic parameters because it involves a valence band electron transition. For successful traditional electron probe microanalysis, the mass absorption coefficient (MAC) must be accurately known, but the MAC of Fe Lα1,2 radiation by Fe atoms varies from one Fe-compound to another and is not well known. We show that the conventional method of measuring the MAC by an electron probe cannot be used in close proximity to absorption edges, making its accurate determination impossible. Fortunately, we demonstrate, using a set of Fe-silicide compounds, that it is possible to derive an accurate calibration curve, for a given accelerating voltage and takeoff angle, which can be used to quantify Fe in Fe-silicide compounds. The calibration curve can be applied to any spectrometer without calibration and gives accurate quantification results.

Searching for the Great Oxidation Event in North America: A Reappraisal of the Huronian Supergroup by SIMS Sulfur Four-Isotope Analysis.

Sedimentological observations from the Paleoproterozoic Huronian Supergroup are suggested to mark the rise in atmospheric oxygen at that time, which is commonly known as the Great Oxidation Event (GOE) and typically coupled with a transition from mass-independent fractionation (MIF) to mass-dependent fractionation (MDF) of sulfur isotopes. An early in situ study of S three-isotopes across the Huronian Supergroup by Papineau et al. ( 2007 ) identified a weak MIF-MDF transition. However, the interpretation and stratigraphic placement of this transition is ambiguous. In this study, all four S isotopes were analyzed for the first time in two Huronian drill cores by secondary ion mass spectrometer (SIMS), and both Δ33S and Δ36S were calculated. Based on improved precision and detailed petrography, we reinterpret the dominance of pyrrhotite in the studied sections, which was previously proposed as "early authigenic" in origin, as resulting from regional metamorphism. Small but analytically resolvable nonzero values of Δ33S (from -0.07‰ to +0.38‰) and Δ36S (from -4.1‰ to +1.0‰) persist throughout the lower Huronian Supergroup. Neither pronounced MIF-S signals nor a MIF-MDF transition are seen in this study. Four scenarios are proposed for the genesis of small nonzero Δ33S and Δ36S values in the Huronian: homogenization by regional metamorphism, recycling from older pyrite, dilution by magmatic fluids, and the occurrence of MDF. We argue that the precise location of the MIF-MDF transition in the Huronian remains unsolved. This putative transition may have been erased by postdepositional processes in the lower Huronian Supergroup, or may be located in the upper Huronian Supergroup. Our study highlights the importance of integrated scanning electron microscopy and secondary ion mass spectrometry techniques in deep-time studies and suggests that different analytical methods (bulk vs. SIMS) and diagenetic history (primary vs. metamorphic) among different basins may have caused inconsistent interpretations of S isotope profiles of the GOE successions at a global scale. Key Words: Great Oxidation Event (GOE)-Secondary ion mass spectrometer (SIMS)-Paleoproterozoic-Sulfur isotopes-Mass independent fractionation (MIF). Astrobiology 18, 519-538.

Tephrostratigraphy of the Waki-Mille area of the Woranso-Mille paleoanthropological research project, Afar, Ethiopia.

Tephra geochemistry and (40)Ar/(39)Ar geochronology are reported for the Waki-Mille area in the northwestern part of the Woranso-Mille paleoanthropological project area in the west central Afar region of Ethiopia. Previous studies documented dentognathic fossils that are morphologically intermediate between Australopithecus anamensis and Australopithecus afarensis and some that are attributed to Australopithecus afarensis. Additional dentognathic remains from the study area were assigned to the newly identified species Australopithecus deyiremeda. These fossil hominin taxa were recovered from volcanic and sedimentary strata containing tuffs ranging in age from more than 3.77 million years ago (Ma) to less than 3.469 Ma. One of the tuffs was correlated based on geochemistry, feldspar mineralogy, and age to the Lokochot Tuff of the Omo-Turkana Basin of southern Ethiopia and Kenya. Variations in major and minor element abundances in volcanic glass demarcate ten geochemically distinct tuffs and tuff sequences, including three that are geochemically similar to widespread regional tuffs, specifically the Lomogol, Lokochot, and ß- Tulu Bor/Sidi Hakoma tuffs. A new (40)Ar/(39)Ar age for the Waki Tuff, which is geochemically similar to the Lomogol Tuff, is 3.664 ± 0.016 Ma. Other tuffs in the Waki-Mille area are geochemically dissimilar to regional tuffs documented to date. Identification of tuffs based on character, stratigraphic position, and geochemistry refines local stratigraphic correlations and delineates the geographic distributions of precisely dated fossiliferous levels within the Waki-Mille area.

Non-Sequential Spectral Acquisitions and Data Reconstruction to Remove Time-Dependent Effects from X-Ray Spectra.

X-ray spectroscopy techniques may require prolonged exposure of a sample to an electron beam to generate X-rays. With typical spectroscopic methods the measured signal is acquired while varying the independent parameter in a systematic way, for example, stepping a photon detector in a series of energy steps from one end of the range of interest to the other incrementally or varying the energy of the incoming excitation incrementally. This can be a time-consuming process when signal strength is low, and if the sample is affected by prolonged beam exposure it could potentially change the shape and position of the obtained X-ray spectrum. Hydrocarbon contamination is of particular concern in electron beam instruments because of the interaction between the hydrocarbons and the electron beam at the point of impact on the sample surface. The authors suggest that hydrocarbon contamination does affect the generated spectra and present methods for removing this (and other) time-dependent artifacts for low-voltage X-rays. The software is freely available as a compiled Windows executable and as source code (http://geoscience.wisc.edu/geoscience/?p=16417).

Low-voltage electron-probe microanalysis of Fe-Si compounds using soft X-rays.

Conventional electron-probe microanalysis has an X-ray analytical spatial resolution on the order of 1-4 µm width/depth. Many of the naturally occurring Fe-Si compounds analyzed in this study are smaller than 1 µm in size, requiring the use of lower accelerating potentials and nonstandard X-ray lines for analysis. Problems with the use of low-energy X-ray lines (soft X-rays) of iron for quantitative analyses are discussed and a review is given of the alternative X-ray lines that may be used for iron at or below 5 keV (i.e., accelerating voltage that allows analysis of areas of interest <1 µm). Problems include increased sensitivity to surface effects for soft X-rays, peak shifts (induced by chemical bonding, differential self-absorption, and/or buildup of carbon contamination), uncertainties in the mass attenuation coefficient for X-ray lines near absorption edges, and issues with spectral resolution and count rates from the available Bragg diffractors. In addition to the results from the traditionally used Fe Lα line, alternative approaches, utilizing Fe Lß, and Fe Ll-η lines, are discussed.

Vanadate and acetate biostimulation of contaminated sediments decreases diversity, selects for specific taxa, and decreases aqueous V5+ concentration.

Vanadium is a commercially important metal that is released into the environment by fossil fuel combustion and mining. Despite its prevalence as a contaminant, the potential for vanadium bioremediation has not been widely studied. Injection of acetate (as a carbon source) directly into an aquifer to biostimulate contaminated sediments in Colorado, United States, resulted in prolonged removal of aqueous vanadium for a period of at least two years. To further investigate this process, we simultaneously added acetate and vanadate (V(5+)) to columns that were packed with aquifer sediment and inserted into groundwater wells installed on the Colorado River floodplain. This allowed evaluation of the microbial response to amendments in columns that received an influx of natural groundwater. Our results demonstrate the removal of up to 99% of the added V(5+)(aq) and suggest microbial mediation. Most probable number measurements demonstrate up to a 50-fold increase in numbers of V(5+)-reducing cells in vanadium-amended columns compared to controls. 16S rRNA gene sequencing indicates decreased diversity and selection for specific taxa in columns that received vanadate compared to those that did not. Overall, our results demonstrate that acetate amendment can be an effective strategy for V removal, and that V bioremediation may be a viable technology.

Quantifying heavy metals sequestration by sulfate-reducing bacteria in an Acid mine drainage-contaminated natural wetland.

Bioremediation strategies that depend on bacterial sulfate reduction for heavy metals remediation harness the reactivity of these metals with biogenic aqueous sulfide. Quantitative knowledge of the degree to which specific toxic metals are partitioned into various sulfide, oxide, or other phases is important for predicting the long-term mobility of these metals under environmental conditions. Here we report the quantitative partitioning into sedimentary biogenic sulfides of a suite of metals and metalloids associated with acid mine drainage contamination of a natural estuarine wetland for over a century.

Effects of pseudowollastonite (CaSiO3) bioceramic on in vitro activity of human mesenchymal stem cells.

We report the effects of two pseudowollastonite (beta-CaSiO(3)) substrates on the attachment, viability, proliferation and osteogenic differentiation of human mesenchymal stem cells (hMSCs), and provide detailed mechanistic links of surface texture, soluble factors and culture media to cell activities. Cell attachment and viability were lower for psWf (fine-grained, roughness 0.74 microm) than for psWc (coarse-grained, roughness 1.25 microm) surface, and were ascribed to the greater specific area of the finer psWf particles resulting in higher release rate of Si, which is cytotoxic at high levels. Interestingly, proliferation was greater on psWf. Osteogenic differentiation occurred on both surfaces, indicated by calcium phosphate bone nodule formation and by osteocalcin, osteopontin and core-binding factor alpha-1 gene expression. Gene levels were lower on psWf than on psWc at day 8 in growth medium, explained by differences in Ca and/or Si concentrations between the two surfaces. Similar gene expression on both surfaces at day 16 in both growth and osteogenic induction media was attributed to pro-osteogenic effects of Ca and P at specific concentrations and complementary Ca and P levels on the two surfaces. In summary, soluble factors from substrates may be more important for osteogenic differentiation in growth medium than small surface roughness variations within a factor of 2. Optimum concentration ranges exist for individual soluble factors to balance cell toxicity/growth versus osteogenic differentiation, and soluble factors together have complex, cooperative or opposing, effects on a given cell activity.