The Environmental Scanning Electron Microscope, or ESEM, is the only class of SEM that can image in a gaseous environment that will maintain a sample in a fully wet state. The use of the patented Gaseous Secondary Electron Detector, or GSED, which amplifies the secondary electron signal with the gas, has allowed the ESEM to image a multitude of samples with true secondary contrast. Recently, several new modes of imaging in a gas have been developed and will allow further expansion of the capabilities of the ESEM.

To maintain pressures in the ESEM up to 20 Torr (27 mbar), the use of multiple, differentially pumped apertures, is required. This can place a restriction on the low magnification range. In the large field detection mode, all magnification restrictions are removed. Magnifications as low as lOx may be achieved. This is similar to many conventional SEMs.

Charge Contrast Imaging (CCI), is a recently reported technique allowing the imaging of the internal microstructure of non and poorly conductive materials. In previous work, the relationship between controlled growth events in batch-precipitated gibbsite, Al(OH)3 and the appearance of contrasting bands in CCIs has been empirically correlated. The data presented was spatial, based on the number and dimensions of growth rings, however no interpretation of the contrast variation was made. Suggestions that the contrast may be related to charge trapping, which in turn is related to conductivity pathways and impurity concentrations indicates that the technique may provide insight into crystallisation processes. The current work examines the spatial distribution of impurity elements and their effect on the contrast observed within growth layers in charge contrast images of gibbsite.

Analyses for Ca, Fe and Si, the main impurities in the solutions before and after precipitation indicate a preferential partitioning of Ca and to a lesser extent Si and Fe into gibbsite.

Understanding the various interactions between electrons and the specimen chamber gas is essential to using environmental and low-vacuum SEM’s to their full potential. The problem can be divided into two main topics: distortion of the primary beam on its way to the sample surface, and amplification of the various electron signals leaving the sample. These issues are important for all low-vacuum SEM’s, regardless of the system design or signal detection method. All instruments must contend with scattering of the primary beam in the gas, and the subsequent effects on resolution, background signals,.and x-ray generation. Similarly, while some instrument designs rely more heavily than others on signal amplification in the gas, all benefit from the concomitant charge neutralisation.

As the electron probe travels through the gas on its way to the specimen, elastic collisions occur with molecules in the gas.[1] Fortunately, for gas pressures of a few torr, the mean free path is on the order of tens of millimetres, and the vast majority of electrons reach the specimen unscattered.

Uncoated, non-conductive samples can be imaged and analyzed in the environmental scanning electron microscope (ESEM) due to effective charge neutralization at the sample surface by ionized gas molecules. Under some gas pressure and electron dose conditions, ESEM images of uncoated, poorly conductive samples often contain contrast not present in secondary or backscattered electron images of the (coated) samples obtained in conventional SEMs. It has been proposed that the contrast is related to charge trapping at defects and impurities. It has also been suggested that UV cathodoluminescence (CL) may contribute to contrast in the ESEM. In this paper, we present experimental evidence of contrast formation in the ESEM due to charge trapping in Dy doped zircon, electron trapping at oxygen vacancies in sapphire and the absence of signal generation by 360nm UV CL.

The specimens used in this study were (i) cross-sectioned Titanium in-diffusion doped sapphire single crystal, (ii) Dy doped synthetic Zircon7 and (iii) 43 μm epitaxial GaN grown on c-pane sapphire by hydride vapor phase epitaxy.

When a poorly conducting specimen is irradiated with an electron beam in a variable pressure electron microscope, the excess charge on the surface of the specimen can be neutralized by incident gas ions to prevent deflection and retarding of the electron beam. A small fraction (<10∼6) of the incident electrons are trapped at irradiation induced or pre-existing defects within the irradiated micro-volume of specimen. The trapped charge induces an electric field, which may result in the electro-migration and micro-segregation of charged mobile defect species within the irradiated volume of specimen. These charge induced effects are dependent on the density of trapping centers and their capture cross sections. In particular, evidence of these micro-diffusion processes can be directly observed in electron beam irradiated ultra pure silicon dioxide (SiO2) polymorphs using Cathodoluminescence (CL) microanalysis (spectroscopy and imaging). CL microanalysis enables both pre-existing and irradiation induced defects in wide band gap materials (i.e. semiconductors and insulators) to be monitored and characterized with high sensitivity and spatial resolution. Depth resolution is achieved by varying the electron beam energy.

Charge Contrast Imaging (CCI) in the environmental scanning electron microscope (ESEM) has rapidly evolved to an important imaging technique for a range of materials. The investigation of the applicability of CCI to the examination of semiconductor and thin film (MCT) samples and devices has been highly successful. Dopant concentration and depth can be differentiated, as has been noted for variable pressure SEM. Quantification of concentration remains difficult due to the complexity of variables controlling CCI but should be achievable in the near future. The extreme sensitivity to surface contaminants previously noted on polished diamond surfaces is also seen on silicon. Monte Carlo modelling of this contrast indicates, for the examined materials, an information depth of a few nanometres for CCI, whilst using 30kV accelerating voltage. The modelling shows that CCI is capable of routine detection of contaminant layers on silicon at the nanometre scale. These results also provide the first data confirming initial suggestions that CCI is derived from the near-surface regions of the sample.

CCI examination of mercury cadmium telluride (MCT) devices has also provided images with new detail. This detail is currently being interpreted. Initial examination suggests an influence of the substrate on film growth (figure 1). Unusual corner structures are visible around the mask region.

We report the effects of scan rate/electron dose on the secondary electron contrast in liquid systems, studied by Environmental Scanning Electron Microscopy (ESEM). Understanding the mechanisms for SE contrast is essential: previous observations of oil-water emulsions have revealed useful SE contrast between the dispersed and continuous phases, attributed to differences in the electronic structure of these molecular liquids. However, it has been found that large oil droplets for example, have a propensity to give SE contrast that can be inverted (relative to the continuous water phase) as a function of scan rate/electron dose, as shown in Figures 1 & 2. In the ‘grabbed’ image, Figure 1, the scan rate is a relatively rapid 2.1 frames/sec. and results in the expected image of oil droplets with lower SE signal intensity than the surrounding water phase. However, the intensity of the oil droplets changes dramatically when the scan rate is slowed, such that the droplets appear bright. Figure 2 shows the same region of sample as in Figure 1, but with image acquisition over 30 seconds. The same contrast inversion can occur at rapid scan rate if higher magnification is used. Effects such as this could result in ambiguous interpretation of ESEM images. However, useful information may be gathered as to the dielectric nature of the medium — a property related to molecular structure. We note that similar scan rate-dependent effects have been observed in ESEM studies of solid samples, although the mechanisms responsible have yet to be rigorously established.

We have measured the secondary electron yield curve for liquid water using an Environmental SEM. The secondary electron emission coefficient, measured as a function of incident electron energy, is important for interpreting contrast in hydrated biological and inorganic specimens. This information is even more critical for water than other materials, as it is a factor of prime importance in understanding radiation damage in biological tissues.[1]

These measurements were taken using a Philips XL-30 field emission ,ESEM, and repeated on an Electroscan E3 ESEM, equipped with a CeB6 filament. A specially designed Faraday cup was fashioned from brass and fitted with a removable graphite cup having an inset for a platinum aperture. This assembly was placed into an electrically floating Peltier cooling stage, and connected to a KE Instruments probe current meter.

Three-dimensional reconstructions are a useful visualisation aid for understanding the volumetric relationships between features in an object. Typically, three dimensional reconstructions have evolved with regards to biological applications, where the collection and manipulation of the data has matured to a point where volumetric rendering has become almost routine. Two common data collection procedures involve either virtual sectioning using CLSM, where layers within the object are imaged successively without mechanical sectioning, or alternatively the projection of the object is imaged, via TEM while being rotated through a range of angles. This paper describes a novel application of three dimensional imaging by collecting 2 dimensional serial section images using a recently described technique for imaging materials using a VPSEM, known as Charge Contrast Imaging.

Figure 1 shows a typical image of a polished grain of gibbsite. Previous work has correlated the appearance of the concentric rings to laboratory controlled precipitation events, thus showing the technique provides information on the precipitation history

Environmental scanning electron microscopy (ESEM) has matured into a mainstream technique in many areas of microscopy. Instrumentation has evolved and our understanding of some of the imaging mechanisms has progressed. However the majority of laboratories where ESEMs are located are based around the materials sciences. Despite the fact that ESEM is the only SEM instrument that permits liquid water to be present whilst imaging, the housing of such a microscope in biological EM units has been relatively rare. This authors laboratory is a multi-user EM unit based in a School of Biological Sciences. There exists the opportunity for basic biological scientists, clinical and pre-clinical medical and dental researchers to make use of such a resource. Indeed as the ESEM is housed alongside a conventional high vacuum instrument and a cryo high vacuum instrument there exists the ideal opportunity to carry out comparative studies.

This study will examine a range of biological samples using ESEM, cryo SEM and dry high vacuum SEM.

Contrast in secondary electron images of p-n junctions has been observed using a FESEM operating at 1kV. This contrast has been attributed to energy band-bending effects and dopant level variations. In this paper, we report that contrast from the depletion layer of a p-n junction can be obtained in an ESEM using a gaseous secondary electron detector (GSED). The contrast is caused by a signal induced in the GSED by charging and discharging of the device.

A 1N4002 p+pn silicon power diode was cross-sectioned along the lead axis to expose the Ag ohmic contacts and the semiconductor device. The section was mechanically polished to an optical finish using diamond abrasives and mounted in a holder which allowed electrical connection to both the p and n sides of the diode. Imaging experiments were performed in a Philips Electroscan XL30 ESEM and a JEOL 6300F below-lens FESEM. Current images were obtained with a GW Electronics 103 specimen current amplifier.

The environmental scanning electron microscope (ESEM) is not typically used for quantitative analysis by energy dispersive x-ray spectrometry (EDS) because the electron beam is scattered by the chamber gas forming a broad tail or skirt rather than a focused spot. Scattered electrons can contribute x-rays from areas not directly under the beam, which compromises the spectrum for quantitative analysis. For specimens compatible with high vacuum, quantitative EDS analysis in a conventional electron microscope is preferred. However, the ESEM has found its niche in providing a vehicle for investigating those specimens that for any of a variety of reasons are not suitable for high vacuum electron microscopy and microanalysis. For these specimens, it is important that we find the most accurate way to perform EDS analysis in the low vacuum environment.

Research has focused on reducing, accounting for, predicting, or measuring the electron scattering on a simplified system. Instrumental tricks to reduce the scattering of the primary electron beam have been reported in the past.

The purpose of this study is to evaluate the variable pressure correction technique (VPCT) as a solution to the problem of extraneous x-ray peaks due to electron beam broadening in the chamber gas of the ESEM. The basis of VPCT is the observation that target x-ray counts decrease with increasing chamber pressure; whereas, x-ray counts due to beam broadening increase. If data are collected at two or more chamber pressures, the number of x-ray counts for an element can be corrected to that expected at zero gas pressure (high vacuum). Tests of NIST SRM 482 have shown EDS x-ray analysis in the ESEM (within the chamber pressure range of 1 to 8 torr) to have comparable accuracy and precision values to those of EDS in the traditional SEM. The samples used in the these studies, however, were quite large (ca. 500 μm in diameter) and so extraneous EDS peaks, due to the electron beam broadening effect of the chamber gas, were minimized.

The 60% Au / 40% Cu wire of SRM 482 was pressed into a hole in the surface of an Al specimen stub so as to produce a flat surface with a sharp interface between the wire and the stub. Spectra were collected at 5 and 150 μm from the junction of the wire and the Al stub at chamber pressures of 2, 4, and 8 torr.

The environmental scanning electron microscope (ESEM) allows imaging and analysis in a gaseous environment at pressures up to 20 torr. Gas phase scattering of primary electrons leads to a loss of useable probe current and produces an electron skirt around the beam that can degrade the spatial resolution in microanalytical studies. Progress in ESEM microanalysis requires accurate modeling and measurement of the beam intensity profile. Previously, x-ray techniques and beam sensitive materials have been used to measure the extent of the skirt. These investigations have yielded inconsistent skirt distances possibly due to dose dependence of the measurement techniques. We present a new, high gain method to measure the useable probe current and skirt profile using the electron beam induced current (EBIC) from a P-N junction.

The interface of an N- and a P-type semiconductor contains an electrostatic donor-acceptor dipole (“depletion layer”) created by an electron-hole recombination energy minimization mechanism.

It is well known that the interaction of the electron beam with the gas in the VP-SEM or ESEM generate the so-called skirt as a result of the elastic collisions between the electrons and the molecules. Since the electrons in the skirt hit the specimen far away from the electron beam, they degrade the resolution of the analyses performed in the VPSEM or ESEM. However, the magnitude and the shape of the skirt are still a matter of controversy despite the fundamental importance of knowing these two factors. In this context, a Monte Carlo program has been developed to simulate the interaction of the electron beam with a gas as a function of the gas composition, gas pressure, electron beam energy and working distance (in reality, we should talk of the total distance traveled by the electron beam in the gas). This Monte Carlo program used a single scattering approach considering elastic collisions only since energy loss is negligible owing to the low density of the gas. 10 millions electron trajectories have been simulated for each conditions.

One of the unique abilities of the environmental scanning electron microscope is that it permits realtime dynamic observations of materials under controlled temperature/atmosphere conditions [1]. The need for accurate and precise measurements of dimensional changes at elevated temperatures arise in many material areas including determining the longitudinal and transverse coefficient of thermal expansion of fibers, sintering studies, hydration and dehydration of absorbent polymers, and strain measurement of materials under an applied load. The purpose of this research is to evaluate the capabilities of an ElectroScan 2020 environmental scanning electron microscope (ESEM™) in performing precise in situ spatial measurements at elevated temperatures. Identification and proposed solution and/or compensation of imaging difficulties such as specimen drift, sample stability, and thermal strain of a reference grid are addressed.

In general, the accuracy of spatial measurements in a scanning electron microscope (SEM) depends on several factors including calibration of the scanned area, the effects of lens hysteresis and reproducibility in the working distance and column alignment.

A variety of porous materials are used in a number of tissue-engineering applications such as cell transplantation and drug delivery. They provide a temporary scaffolding for a large cell mass, however they must have a fully interconnected pore structure. One such material is sodium alginate, a naturally derived polysaccharide which is isolated from seaweed. Sodium alginate can de dissolved in water to form a highly viscous solution that can be cross-linked in the presence of divalent cations (e.g., Ca2+) to form what is known a as hydrogels. We have developed a method for the fabrication of porous beads of sodium alginate and have attempted to measure the pore size, density and distribution in the beads. Characterization of the pore size and distribution is necessary to determine if the material is appropriate to promote cellular in-growth. The characterization procedures have been limited to conventional scanning electron microscopy (SEM) and mercury intrusion porosimetry. There has been some concern as to the potential introduction of artifacts (spatial distortion) during dehydration and sample preparation during these characterizations.

Biological scanning electron microscopy is increasingly performed at low beam energies in order to improve image contrast, reduce charging artifacts, and minimize beam induced damage to the sample. It is natural then to wish to also use these same low accelerating voltage for X-ray microanalysis using an energy dispersive spectrometer (EDS) but a variety of fundamental physical effects affect the performance that can be achieved. An X-ray photon can only be emitted when the incident beam Eo energy exceeds the critical excitation energy Ecrit for that line. As the beam energy is reduced the number of elements that can be excited falls and it is becomes necessary to use L- and M-lines rather than the K-lines accessible at higher energies. At 5keV, the upper limit of ‘low voltage’ microscopy, K-lines can be excited from elements up to calcium, L-lines can be detected up to cesium, and the rest of the periodic table is available using M-lines. The entire periodic table is therefore, in principle, available at an overvoltage U>2, where U = E0/Ecrit But at lower energies the number of accessible excitations falls and some elements cannot, with current technology, then be analyzed and for general purposes an incident energy lower than about 3keV is probably too limiting to be useful.

Microanalysis performed at low electron beam energies (≤ 5 keV) is limited by the physics of x-ray generation and the performance of existing semiconductor energy dispersive spectrometry (EDS) and wavelength dispersive spectrometry (WDS). Low beam energy restricts the atomic shells that can be excited for elements of intermediate and high atomic number, forcing the analyst to consider using unconventional M- and N-shells for elements such as Sn and Au. Unfortunately, these shells have very low fluorescent yield, which results in inherently low spectral peak-to-background ratios. The modest energy resolution of semiconductor EDS leads to poor limits of detection for these weakly emitted photons. The situation is further complicated by the inevitable interferences with the much more strongly excited K-shell x-rays of the light elements, particularly carbon and oxygen. WDS has the spectral resolution to overcome the resolution limitations of semiconductor EDS. However, WDS has a low geometric efficiency, and because of its narrow instantaneous spectral transmission, spectral scanning is required to detect and analyze x-ray peaks. Moreover, the high resolution field-emission-gun scanning electron microscope (FEG-SEM) provides only a few nanoamperes of beam current.

The use of a high-brightness field-emission gun and a high-resolution lens system in modern SEM instruments allows us to perform high-resolution chemical analysis and atomic-number contrast imaging of bulk materials at low voltages. Low voltage microanalysis is especially useful for analyzing surface coatings or deposits, catalyst particles, small inclusions or precipitates, and biological samples. In this report, we present useful applications as well as limitations of low voltage microanalysis in studying surface-related problems.

In the study of film formation on metal coupons for inhibiting metal corrosion in drinking water supplies, highly porous films were found to form on fresh metal coupons after treatment with various corrosion inhibitors. Figure 1a shows a low voltage (1.5 kV) SE image of a copper test coupon that was exposed to flowing water for a period of two months. The SE image clearly shows the porosity of the surface film and the presence of different phases.