Measures for spectral quality in low-voltage X-ray microanalysis

Characteristic x-ray production with energetic electrons depends strongly on the overvoltage, the ratio of the incident beam energy to the critical excitation energy for the atomic species of interest. Low-voltage x-ray microanalysis (beam energy < or = 5 keV) is especially susceptible to artifacts due to sample charging because the overvoltage is low and even slight charging can strongly affect peak intensities. The Duane-Hunt bremsstrahlung limit is a good diagnostic to detect sample charging. Dynamic charging effects, however, can influence spectra despite an apparently satisfactory Duane-Hunt limit. Dynamic charging effects must be examined by time series experiments, or through use of dynamic energy windows continuously measuring count rates placed across the spectrum. When charging is a problem, conductive surface coatings can eliminate the effects. When pristine surfaces must be examined without coating, the use of a conductive grid can control charging so that useful x-ray spectra can be obtained.     

X-ray microanalysis of porous materials using Monte Carlo simulations

Quantitative X-ray microanalysis models, such as ZAF or φ(ρz) methods, are normally based on solid, flat-polished specimens. This limits their use in various domains where porous materials are studied, such as powder metallurgy, catalysts, foams, etc. Previous experimental studies have shown that an increase in porosity leads to a deficit in X-ray emission for various materials, such as graphite, Cr(2) O(3) , CuO, ZnS (Ichinokawa et al., '69), Al(2) O(3) , and Ag (Lakis et al., '92). However, the mechanisms responsible for this decrease are unclear. The porosity by itself does not explain the loss in intensity, other mechanisms have therefore been proposed, such as extra energy loss by the diffusion of electrons by surface plasmons generated at the pores-solid interfaces, surface roughness, extra charging at the pores-solid interface, or carbon diffusion in the pores. However, the exact mechanism is still unclear. In order to better understand the effects of porosity on quantitative microanalysis, a new approach using Monte Carlo simulations was developed by Gauvin (2005) using a constant pore size. In this new study, the X-ray emissions model was modified to include a random log normal distribution of pores size in the simulated materials. This article presents, after a literature review of the previous works performed about X-ray microanalysis of porous materials, some of the results obtained with Gauvin's modified model. They are then compared with experimental results.     

Charging compensation of alumina samples by using an oxygen microinjector in the environmental scanning electron microscope

A gas microinjector system was set up in an environmental scanning electron microscope (ESEM) to create an oxygen atmosphere around the alumina samples for the charging compensation under a pressure between 2 x 10(-5) Pa approximately 2 x 10(-2) Pa. At low pressures, the skirt effect of the electron scattering can be degraded, which results in improvement of the imaging contrast and increase of the signal/noise ratio. The sample current (I(SC)) and the Duane-Hunt limit were measured to evaluate the charging effect.     

X-ray microanalysis of freeze-dried and frozen-hydrated cryosections

The elemental composition and the ultrastructure of biological cells were studied by scanning transmission electron microscopy (STEM) combined with energy dispersive X-ray microanalysis. The preparation technique involves cryofixation, cryoultramicrotomy, cryotransfer, and freeze-drying of samples. Freeze-dried cryosections 100-nm thick appeared to be appropriate for measuring the distribution of diffusible elements and water in different compartments of the cells. The lateral analytical resolution was less than 50 nm, depending on ice crystal damage and section thickness. The detection limit was in the range of 10 mmol/kg dry weight for all elements with an atomic number higher than 12; for sodium and magnesium the detection limits were about 30 and 20 mmol/kg dry weight, respectively. The darkfield intensity in STEM is linearly related to the mass thickness. Thus, it becomes possible to measure the water content in intracellular compartments by using the darkfield signal of the dry mass remaining after freeze-drying. By combining the X-ray microanalytical data expressed as dry weight concentrations with the measurements of the water content, physiologically more meaningful wet weight concentrations of elements were determined. In comparison to freeze-dried cryosections frozen-hydrated sections showed poor contrast and were very sensitive against radiation damage, resulting in mass loss. The high electron exposure required for recording X-ray spectra made reproducible microanalysis of ultrathin (about 100-nm thick) frozen-hydrated sections impossible. The mass loss could be reduced by carbon coating; however, the improvement achieved thus far is still insufficient for applications in X-ray microanalysis. Therefore, at present only bulk specimens or at least 1-μm thick sections can be used for X-ray microanalysis of frozen-hydrated biological samples.     

Reduction of background due to backscattered electrons in energy-dispersive X-ray microanalysis

When energy-dispersive X-ray microanalysis is practised in a scanning electron microscope, most of the spectral background may come from electrons entering the detector. This background can be eliminated by deflecting magnets. Alternatively, the electrons can be blocked by a plastic film but only at the cost of suppression of the low-energy end of the X-ray spectrum.     

A sample preparation for quantitative determination of magnesium in individual lymphocytes by electron probe X-ray microanalysis

We present a sample preparation method for measuring magnesium in individual whole lymphocytes by electron probe X-ray microanalysis. We use Burkitt's lymphoma cells in culture as the test sample and compare X-ray microanalysis of individual cells with atomic absorption analysis of pooled cell populations. We determine the magnesium peak-to-local continuum X-ray intensity ratio by electron probe X-ray microanalysis and calculate a mean cell magnesium concentration of 39± 19 mmol/kg dry weight from analysis of 100 cells. We determine a mean cell magnesium concentration of 34 ±4 mmol/kg dry weight by atomic absorption analysis of pooled cells in three cell cultures. The mean cell magnesium concentrations determined by the two methods are not significantly different. We find a 10% coefficient of variation for both methods of analysis and a 30% coefficient of variation in magnesium concentration among individual cells by electron probe X-ray microanalysis. We wash cells in ammonium nitrate for microanalysis or in buffered saline glucose for atomic absorption analysis. We find cells washed in either solution have the same cell viability (85%), recovery (75%), cell volume (555 μm³) and cytology. We air dry cells on thin film supports and show by magnesium X-ray mapping that magnesium is within the cells. We conclude that: (a) our microanalysis cell preparation method preserves whole intact lymphocytes; (b) there is no systematic difference in results from the two methods of analysis; (c) electron probe X-ray microanalysis can determine the variation in magnesium concentration among individual cells.     

Charging effect in electron-irradiated ice

To maintain the original distribution pattern of diffusible elements in biological samples, electron probe microanalysis is carried out with frozen hydrated bulk specimens and cryosections, analysed at temperatures below 130 K. Ice has a very low intrinsic conductivity at this working temperature and surface- and space-charging appears, when uncoated specimens are irradiated with non-penetrating electrons. Although coating with a grounded conductor abolishes the surface potential, the build-up of an internal space-charge field is possible, depending on the sample thickness and beam voltage used. Consequently, the geometry of the X-ray source volume and the spectral distribution of the emitted continuous and characteristic X-rays are affected. To simulate the situation for microanalysis of frozen hydrated specimens the charging process in electron irradiated ice is studied by recording simultaneous specimen currents from the top and bottom of ice layer preparations. The external currents yield information on the build-up of internal space-charge fields which result from the balance of charge injection, storage, and transport. Irradiation of uncoated bulk specimens with a finely focused beam results in the build-up of a space-charge field close to the surface, which causes a reduction of the depth of microprobe analysis. In coated bulk specimens the induced conductivity renders possible a current flow to the front electrode, thereby limiting the space-charge field. Sections with an effective rear electrode will not charge appreciably if the electron range is larger than about half the section thickness.     

Mass determination of thin biological specimens using backscattered electrons. Application in quantitative X-ray microanalysis on an automated STEM system

The capabilities of modern computerized X-ray analysis systems can be expanded to the acquisition of various signals available in the electron probe microanalyzer, in parallel with the X-rays. These facilities allow the use of backscattered electrons for the measurement of the total specimen mass thickness, which can be used in mass fraction calculations, up to a (biological) specimen thickness of 10 micron. A mass measurement procedure based on the use of backscattered electrons may become an alternative for the X-ray continuum normalization method, often used in electron probe X-ray microanalysis. A mass measurement procedure using backscattered electrons is described, and preliminary results are given.     

An experimental model of beam broadening in the variable pressure scanning electron microscope

In the variable pressure scanning electron microscope (VP-SEM) the incident electrons pass through a gaseous environment and the beam is scattered by these interactions. We show here that the experimental intensity profile of the scattered beam can be described as Gaussian in form to a high level of accuracy. This provides a rapid means of accounting for the effects of beam scatter in imaging and microanalysis because the standard deviation of the Gaussian is a simple function of parameters such as working distance, beam energy, gas type and pressure.     

Effect of the sample annealing temperature and sample crystallographic orientation on the charge kinetics of MgO single crystals subjected to keV electron irradiation

This paper focuses on the effect of sample annealing temperature and crystallographic orientation on the secondary electron yield of MgO during charging by a defocused electron beam irradiation. The experimental results show that there are two regimes during the charging process that are better identified by plotting the logarithm of the secondary electron emission yield, lnσ , as function of the total trapped charge in the material Q_T. The impact of the annealing temperature and crystallographic orientation on the evolution of lnσ is presented here. The slope of the asymptotic regime of the curve lnσ as function of Q_T, expressed in cm² per trapped charge, is probably linked to the elementary cross section of electron–hole recombination, σ_hole, which controls the trapping evolution in the reach of the stationary flow regime.     

Ionic composition of rat airway surface liquid determined by X-ray microanalysis

The thin layer of liquid that lines the conducting airway epithelium, the airway surface liquid (ASL), is important for mucociliary clearance. Altered ionic composition and/ or volume of the ASL play a major role in the pathology of airway diseases such as cystic fibrosis. Since the ASL is a thin layer, it has been difficult to exactly determine its composition. The present paper describes two techniques that have been developed and used to study ASL composition: X-ray microanalysis of frozen hydrated rat trachea, and an ion-exchange (dextran) bead method, where dextran beads were placed on the airway epithelium to equilibrate with the ASL; the beads were then collected under silicone oil, dried and analyzed by X-ray microanalysis. The results from both frozen-hydrated specimens and from the dextran beads showed that ASL from rat trachea is hypotonic. Concentrations of Na, P, S, and K were higher in the frozen-hydrated ASL, in which mainly the mucus layer is analyzed, compared with the dextran bead method, in which mainly the periciliary liquid is sampled. Also the composition of rat nasal fluid was investigated by the dextran bead method. This fluid was somewhat hypertonic because of a high K concentration. The ionic composition of the nasal and tracheal fluid can be manipulated by cholinergic or alpha- or beta-adrenergic stimulation. Collecting ASL with dextran beads did not disturb the integrity of the airway epithelium. The ionic composition of the collected beads remained stable for several days during storage in silicone oil. It is concluded that X-ray microanalysis is a suitable method to determine the ionic composition of ASL.     

Mistakes encountered during automatic peak identification in low beam energy X-ray microanalysis

Automated peak identification in electron beam excited X-ray microanalysis with energy dispersive X-ray spectrometry (EDS) is subject to occasional mistakes even on well-separated, high-intensity peaks arising from major constituents. The problem is exacerbated when analysis conditions are restricted to operation in the "low beam energy scanning electron microscopy" (i.e. "low voltage scanning electron microscopy" or LVSEM) regime where the incident beam energy is 5 keV or less. These low beam energy microanalysis conditions force the analyst to use low fluorescence yield L-shell and M-shell peaks rather than higher yield K-shell and L-shell peaks typically selected for elements of intermediate and high atomic number under conventional high beam energy (>10 keV) conditions. Misidentifications can arise in automated peak identification procedures when only a single energy channel is used to characterize an EDS peak. The effect of the EDS measurement process is to convolve the closely spaced Lalpha-Lbeta and Malpha-Mbeta peaks into a single peak with a peak channel shift of 20 eV or more from the Lalpha or Malpha value, which is typically sought in an X-ray database. An extensive list of problem situations encountered in low beam energy microanalysis is presented based upon observed peak identification mistakes as well as likely troublesome situations based upon proximity in peak energy. Robust automatic peak identification requires implementation of peak fitting that utilizes the full peak shape.     

The preparation and X-ray microanalysis of bulk frozen hydrated vacuolate plant tissue

A series of modifications have been devised which allow the peak to background ratio X-ray analytical method to be used more effectively to measure elemental concentrations in large vacuolate plant cells. Planar, frozen-hydrated fracture faces of bulk plant tissue are coated with a thin film of evaporated chromium, which prevents surface charging. Provided the film is sufficiently thin, c. 5–10 nm, there is no attenuation of the electron beam and only a small absorption of soft X-rays. The chromium makes a small but measurable contribution to the spectral background and suitable corrections may be made to the quantitative results. An improved back-scattered imaging system is described, which helps to overcome the problem of spurious X-ray signals from rough surfaces. The microscope column has been modified to permit a continuous readout of beam current, sensu stricta, during X-ray microanalysis and to allow rapid exchange of the electron gun assembly during low temperature operation. Calculations are given relating the size of the X-ray interactive volume to electron penetration and X-ray emission in both frozen hydrated and frozen dried cells. The problems of X-ray microanalysis are discussed in relation to the highly vacuolate cells found in most mature plant tissues and an example given of the distribution of four major cations in tobacco leaves.     

Experimental data and model simulations of beam spread in the environmental scanning electron microscope

This work describes the comparison of experimental measurements of electron beam spread in the environmental scanning electron microscope with model predictions. Beam spreading is the result of primary electrons being scattered out of the focused beam by interaction with gas molecules in the low-vacuum specimen chamber. The scattered electrons form a skirt of electrons around the central probe. The intensity of the skirt depends on gas pressure in the chamber, beam-gas path length, beam energy, and gas composition. A model has been independently developed that, under a given set of conditions, predicts the radial intensity distribution of the scattered electrons. Experimental measurements of the intensity of the beam skirt were made under controlled conditions for comparison with model predictions of beam skirting. The model predicts the trends observed in the experimentally determined scattering intensities; however, there does appear to be a systematic deviation from the experimental measurements.     

Observation of the ion-mirror effect during microscopy of insulating materials

Utilizing the ion beam of a focused ion beam (FIB)/scanning electron microscope (SEM) microscope to investigate non‐conductive samples, we observe a mirror image very much similar to the one that is commonly obtained with the electron beam and the same samples. To our knowledge this is the first observation of what can be called ‘Ion‐Mirror Effect’. This effect is produced by a positive charging of the sample obtained by rastering with high‐energy ions (30 kV) and a subsequent imaging with low energy ones (5 kV). The proposed explanation is that first a positive charge is trapped within the sample and eventually the lower energy ions are deflected back by the latter, and hit the surface of the microscope chamber very similar to what happens in the ‘Electron‐Mirror Effect’. The mirror image is produced after detection of the electrons produced by the interaction between ions and the chamber materials.     

Charging phenomena in PEEM imaging and spectroscopy

Spectromicroscopy with the imaging technique of X-ray photoelectron emission microscopy (X-PEEM) is a microchemical analytical tool installed in many synchrotron radiation laboratories, and which is finding application in diverse fields of research. The method of sample analysis, X-ray absorption spectroscopy, does not encounter the same problems as X-ray photoemission spectroscopy when sample charging occurs, hence even good insulators may often be analyzed without any apparent artifacts in images or spectra. We show, however, that charging effects cannot be neglected. We model the effect of surface charge formation on the secondary electron yield from uniform samples to demonstrate that surface charge primarily reduces the yield of electrons which may contribute to the detected signal. We illustrate that on non-uniform insulating samples, localized centers of charge may substantially affect microscope imaging and resolution as the electrostatic field close to the surface is distorted. Finally, in certain circumstances non-uniform surface charge may lead to unexpected lineshapes in X-ray absorption spectra causing, in some extreme cases, negative spectra. These negative spectra are explained, and several strategies are reviewed to minimize the impact of sample charging when analyzing poorly conducting samples of any nature.     

Vacuum electron acceleration and bunch compression by a flat-top laser beam

The field intensity distribution and phase velocity characteristics of a flat-top laser beam are analyzed and discussed. The dynamics of electron acceleration in this kind of beam are investigated using three-dimensional test particle simulations. Compared with the standard (i.e., TEM(00) mode) Gaussian beam, a flat-top laser beam has a stronger longitudinal electric field and a larger diffraction angle. These characteristics make it easier for electrons to be trapped and accelerated by the beam. With a flat-top shape, the laser beam is also applicable to the acceleration of low energy electron and bunch compression.     

The application of analytical electron microscopy to the study of diseased biological tissue

A routine method is described for X-ray microanalysis of thin specimens of biological tissue containing mineral particles in cancerous growths. Such a method allows information to be obtained that relates pathological history to histology, electron microscopy and X-ray microanalysis. Classification of minerals is possible in a way that is not provided by bulk analysis. The technique provides baselines of elemental data of minerals from various sources that may be used to classify particles found present in certain tumour growths.     

基于钛酸锂电池的短编组有轨电车车载储能方案仿真分析

适用于中小城市的短编组车载储能式低地板有轨电车由于全线仅需在车站设充电站，在正线区间不需要架设接触网，对城市景观及限高几乎无影响而在近年得到快速发展，但因短编组有轨电车车长较短，车顶安装空间受限，因此其车载储能装置容量非常有限，导致其在线路沿线设置的充电站数量较多，成本较高。为提高有轨电车车载储能装置容量，采用钛酸锂电池代替有轨电车常用的超级电容并在保山有轨电车T1线实际线路上进行了仿真分析，仿真结果表明，采用钛酸锂电池代替超级电容后，在相同体积下车载储能装置容量可提高近1倍，区间充电站数量约为原数量的1/2，降低了工程实施费用及后期充电站检修维护成本。     

Monte Carlo simulation of charging effects on linewidth metrology

Charging effects of scanning electron microscopes on the linewidth metrology of polymethylmethacrylate (PMMA) insulatorpatterns are investigated using Monte Carlo simulation. It is first revealed in detail how the nonunity yield of electron generation in the PMMA target leads to local charge accumulation and affects the image profile of secondary electrons as charging develops. Then the measurement offset due to charging effects is identified for various target patterns of isolated and array types. Finally, it is concluded that the measurement uncertainty caused by the measurement offset exceeds the error budget limit that will be allowed in the linewidth metrology of the next generation of semiconductors.