Comparative characterization of chemical vapor deposition diamond films by scanning cathodoluminescence microscopy

Thin diamond films grown by chemical vapor deposition (CVD) process on Si substrates under similar deposition conditions in the microwave-excited (MW) and direct current (DC) plasma discharges were taken for comparative examination. Raman spectra, photoluminescence (PL) spectroscopy, and color cathodoluminescence scanning electron microscopy (CCL-SEM) have been used for characterization of the structure and composition features of poly-crystalline diamond films. No essential difference in Raman spectra for the CVD diamond films was detected. A significant difference was revealed in the PL spectra and in CCL-SEM images.     

Topographic and material contrast in low-voltage scanning electron microscopy

Two scintillation backscattered electron (BSE) detectors with a high voltage applied to scintillators were built and tested in a field emission scanning electron microscope (SEM) at low primary beam energies. One detector collects BSE emitted at low take-off angles, the second at high takeoff angles. The low take-off detector gives good topographic tilt contrast, stronger than in the case of the secondary electron (SE) detection and less sensitive to the presence of contamination layers on the surface. The high take-off detector is less sensitive to the topography and can be used for detection of material contrast, but the contrast becomes equivocal at the beam energy of 1 keV or lower.     

Backscattered electron imaging of the undersurface of resin-embedded cells by field-emission scanning electron microscopy

In this study backscattered electron (BSE) imaging was used to display cellular structures stained with heavy metals within an unstained resin by atomic number contrast in successively deeper layers. Balb/c 3T3 fibroblasts were cultured on either 13-mm discs of plastic Thermanox, commercially pure titanium or steel. The cells were fixed, stained and embedded in resin and the disc removed. The resin block containing the cells was sputter coated and examined in a field-emission scanning electron microscope. The technique allowed for the direct visualization of the cell undersurface and immediately overlying areas of cytoplasm through the surrounding embedding resin, with good resolution and contrast to a significant depth of about 2 μm, without the requirement for cutting sections. The fixation protocol was optimized in order to increase heavy metal staining for maximal backscattered electron production. The operation of the microscope was optimized to maximize the number of backscattered electrons produced and to minimize the spot size. BSE images were collected over a wide range of accelerating voltages (keV), from low values to high values to give ‘sections' of information from increasing depths within the sample. At 3–4 keV only structures a very short distance into the material were observed, essentially the areas of cell attachment to the removed substrate. At higher accelerating voltages information on cell morphology, including in particular stress fibres and cell nuclei, where heavy metals were intensely bound became more evident. The technique allowed stepwise ‘sectional’ information to be acquired. The technique should be useful for studies on cell morphology, cycle and adhesion with greater resolution than can be obtained with any light-microscope-based system.     

Visualizing soft tissue in the mammalian cochlea with coherent hard X-rays

This paper concerns an important aspect of current developments in medical and biological imaging: the possibility for imaging soft tissue at relatively high resolution in the micrometer range or better, without tedious and/or entirely destructive sample preparation. Structures with low absorption contrast have been visualized using in-line phase contrast imaging. The experiments have been performed at the Advanced Photon Source, a third generation source of synchrotron radiation. The source provides highly coherent X-ray radiation with high photon flux (>10(14) photons/s) at high photon energies (5-70 keV). Thick gerbil cochlear slices have been imaged and were compared with those obtained by light microscopy. Furthermore, intact gerbil cochleae have been imaged to identify the soft tissue structures involved in the hearing process. The present experimental approach was essential for visualizing the inner ear structures involved in the hearing process in an intact cochlea.     

反射电子抑制技术和新型电子枪(BS-60050EBS)

在使用电子束真空镀膜时所产生的反射电子有可能射入基板,造成MgF2等不稳定化合物薄膜吸收的增加,另外在使用萤石(CaF2)基板时则会造成基板变色等问题。BS-60050EBS电子枪优化了反射电子发生部分的磁场分布、能够有效地抑制反射电子射入基板、提高镀膜产品的质量。     

The effect of subsurface topography on secondary electron images from chromate pretreated aluminium surfaces

In secondary and scanning transmission electron microscopes, secondary electron images of surface films can be dominated by an image derived from electrons back-scattered from the interface between the film and the substrate. The extent of the domination has been established by studying the variation in image obtained using primary beams of different energy and by platinum coating to enhance surface secondary electron emission. Studies of thicker films also established that chemical or structural difference within a film also lead to imaging effects. In general, 5 keV electrons are the most effective in producing subsurface and structural or chemical imaging effects.     

Errors in quantitative backscattered electron analysis of bone standardized by energy-dispersive x-ray spectrometry

Backscattered electron (BSE) imaging has proven to be a useful method for analyzing the mineral distribution in microscopic regions of bone. However, an accepted method of standardization has not been developed, limiting the utility of BSE imaging for truly quantitative analysis. Previous work has suggested that BSE images can be standardized by energy-dispersive x-ray spectrometry (EDX). Unfortunately, EDX-standardized BSE images tend to underestimate the mineral content of bone when compared with traditional ash measurements. The goal of this study is to investigate the nature of the deficit between EDX-standardized BSE images and ash measurements. A series of analytical standards, ashed bone specimens, and unembedded bone specimens were investigated to determine the source of the deficit previously reported. The primary source of error was found to be inaccurate ZAF corrections to account for the organic phase of the bone matrix. Conductive coatings, methyl-methacrylate embedding media, and minor elemental constituents in bone mineral introduced negligible errors. It is suggested that the errors would remain constant and an empirical correction could be used to account for the deficit. However, extensive preliminary testing of the analysis equipment is essential.     

Peculiarities of imaging one- and two-dimensional structures using an electron microscope in the mirror operation mode

Measurements performed in an electron microscope with the mirror operation mode are most sensitive to local electric fields and geometrical roughness of any kind of the object being studied. The object with a geometrical relief is equivalent to a smooth surface with an effective distribution of microfields. Electrons forming the image interact with the local microfields for an extended time: during approach to the object, deceleration and acceleration away from the object. As a result, the electron trajectories can be strongly distorted, and the contrast changes essentially, leading to image deformation of details of the object under investigation and to lowering of the resolution. These effects are theoretically described and are illustrated by experiments. An analysis of these effects enables the real size and the shape of the object involved to be reconstructed.     

Dual-channel heterodyne microscope: new functions

Quantitative assessment of critical dimensions of sub-wavelength structures, dark-field and bright-field modes of operation, recognition of nonpatterned anisotropic surfaces, and three-dimensional profiling of opaque and transparent samples are the new features of a super-resolution dual-channel heterodyne microscope that was introduced in a previous publication in this journal. The aforementioned modes of operation can be activated by simple rotation of a polarizer or insertion of a polarization-separating prism. Various experimental results are presented, demonstrating new performance capabilities of the heterodyne microscope.     

The method of increasing COMPO contrast by linearization of backscattering characteristic η =f(Z)

The backscattered electron signal (BSE) in the scanning electron microscope (SEM) has been used for investigation of a specimen surface composition (COMPO mode). Creation of a material composition map is difficult because the dependence of backscattering coefficient η on the atomic number Z for Z > 40 is nonlinear. The method of increase in SEM resolution for the BSE signal by use of digital image processing has been proposed. This method is called the linearization of the η =f(Z) characteristic. The function approximating the experimental η =f (Z) dependence was determined by numerical methods. After characteristics linearization, the digital image in COMPO mode allows to distinguish between two elements with high atomic numbers if their atomic numbers differ by ΔZ = 1.     

Influence of topography and specimen preparation on backscattered electron images of bone

Backscattered electron (BSE) images of bone exhibit graylevel contrast between adjacent lamellae. Mathematical models suggest that interlamellar contrast in BSE images is an artifact due to topographic irregularities. However, little experimental evidence has been published to support these models, and it is not clear whether submicron topographical features will alter BSE graylevels. The goal of this study was to determine the effects of topography on BSE image mean graylevels and graylevel histogram widths using conventional specimen preparation techniques. White-light interferometry and quantitative BSE imaging were used to investigate the relationship between the BSE signal and specimen roughness. Backscattered electron image graylevel histogram widths correlated highly with surface roughness in rough preparations of homogeneous materials. The relationship between BSE histogram width and surface roughness was specimen dependent. Specimen topography coincided with the lamellar patterns within the bone tissue. Diamond micromilling reduced average surface roughness when compared with manual polishing techniques but did not significantly affect BSE graylevel histogram width. The study suggests that topography is a confounding factor in quantitative BSE analysis of bone. However, there is little quantitative difference between low-to-moderate magnification BSE images of bone specimens prepared by conventional polishing or diamond micromilling.     

Mirror lenses in light microscopy—Theoretical considerations and practical implications

Various types of mirror lenses were developed some decades ago designed for several special tasks in microscopy. When compared with high‐end glass lenses, mirror lenses lead to an extraordinary image quality because of their supraapochromatic color fidelity and their high planarity; the working distances are significantly longer, and in most cases, the depth of field and resolution are higher than in concurrent glass lenses. In this respect, the microscopic advantages of mirror lenses seem to be comparable with the advantages of mirror telescopes in astronomy; for in observations of celestial bodies, mirror telescopes lead to better results than refractors based on glass lenses. When mirror lenses are available in light microscopy, not only common illumination modes can be carried out in transmitted or incident light, but also new and specific axial illumination techniques can be utilized, which cannot be achieved with normal glass lenses. This axial illumination, called “luminance contrast,” can be carried out in various modes, so that the resulting images can be compared with dark‐field, phase or interference‐contrast images. In all variants, especially, fine details within transparent specimens can be visible in maximized contrast and resolution, and blooming or haloing artifacts are significantly reduced or absent. These findings are based on theoretical consideration, and intensive practical tests carried out with several mirror lenses constructed in various optical designs. All in all, supramicroscopic image qualities could be expected if mirror lenses were produced based on the optimized manufacturing techniques available nowadays. Microsc. Res. Tech. 2010. © 2009 Wiley‐Liss, Inc.