A simple microtome capable of cutting sections of plastic-embedded material down to 1 μm in thickness

Details of the design of an easily constructed microtome are presented. The instrument uses glass knives to cut sections down to about 1 μm in thickness, and is suitable for the examination of plastic-embedded material.     

A simple,practical ‘swiss roll’ method of preparing tissues for paraffin or methacrylate embedding

A simple method of preparing ‘swiss rolls' from strips of tissue up to 500 mm in length is described. This procedure has been applied to a variety of tissues and is suitable for the preparation of both paraffin- and methacrylate-embedded specimens.     

A new,less toxic polymerization system for the embedding of soft tissues in glycol methacrylate and subsequent preparing of serial sections

The paper describes a new polymerization system for embedding soft tissues in glycol methacrylate (GMA). The polymerization of GMA is initiated by means of a barbituric acid derivative in combination with chloride ions and dibenzoyl peroxide. The catalyst system contains no aromatic amines which constitutes a toxicological advantage over the commonly employed system of peroxide/aromatic amine. Clear blocks are obtained from which 1–2 μm sections are easy to cut. In combination with an appropriate softener, polyethylene glycol 400 , serial sectioning may be practised.     

A simple method to obtain en face sections of flat tissue culture cells

A method is described for embedment of multiple confluent sheets of flat tissue culture cells that permits sectioning for thin or semithin sections in precise planes. The technique is especially useful for obtaining en face sections.     

A simple device to aid tissue processing of large numbers of blocks for light or electron microscopy

A simple device is described that aids the processing of large numbers of tissue blocks for light or electron microscopy. The device has a number of compartments into which different tissue blocks can be placed. Each compartment has a fine mesh floor, allowing the free exchange of the reagents used in tissue processing, and a gentle flow of the reagents is promoted by a magnetic stirring bar, or a rotary mixer. All the materials used are resistant to solvents such as propylene oxide. The device is labor-saving in that many blocks can be processed as easily as a single block. The device may also be of value in quantitative microscopy, since it ensures that all tissue blocks examined experience an identical processing history.     

The α‐β circular scanning with large range and low noise

A circular‐route scanning method called α‐β circular scanning is proposed and realized using sinusoidal signals with a constant phase difference of π/2. Experiments show that the circular scanning range of α‐β circular scanning is 57% greater than the rectangular scanning range of raster scanning within an effective optical field of view. Moreover, the scanning speed is improved by 7.8% over raster scanning because the whole sine signal is utilized in α‐β circular scanning whereas the flyback area of the saw‐tooth signal needs to be discarded in raster scanning. The maximum scanning acceleration decreases by a factor of 44, drastically decreasing the high noise, which should considerably elongate the lifetime of the galvanometers while inhibiting internal vibration. The proposed α‐β circular scanning technique could be used in scanning imaging, optical tweezers and laser‐beam fabrication.     

Micro-structuring of glass with features less than 100 μm by electrochemical discharge machining

Micro-electrochemical discharge machining (ECDM) was studied in order to improve the machining of 3D micro-structures of glass. To minimize structures and obtain good surface microstructures, the effects of the electrolyte, the pulse on/off-time ratio, the voltage, the feedrate, the rotational speed, and the electrolyte concentration in the drilling and milling processes were studied.In ECDM, voltage is applied to generate a gas film and sparks on a tool electrode; however, high voltage produces poor machining resolution. To obtain a stable gas film over the whole surface of the tool at a low voltage, a new mechanical contact detector, based on a loadcell, was used; the immersion depth of the tool electrode in the electrolyte was reduced as much as possible. In this study, various micro-structures less than 100 μm in size, such as Ø 60 μm micro-holes, a 10 μm-thin wall, and a 3D micro-structure were fabricated to demonstrate the potential for micro-machining of glass by ECDM.     

Application of a new detector for cathodoluminescence measurements in the wavelength range to 1.8 μm

The energetic gap structure of semiconductors or insulators can strongly be influenced by the local appearance of inhomogenities, impurities, dopants or vacancies. A high spatial resolution cathodoluminescence (CL) measuring technique makes it possible to investigate this gap structure via spectral analysis of the emitted CL. This can lead to a detailed knowledge of the local defect distribution. The wavelength range which could be detected by CL measurements has, up to the present, been limited to values less than 1 μm, because no detectors were available for higher wavelengths. By use of a new germanium detector, the measuring range could be extended to 1.8 μm. This makes it possible to analyse the CL properties, both of materials with small gap energies and of deep impurities. The detector properties which are important for CL measurements are presented. The efficiency of the detector is demonstrated by CL investigations of barium titanate ceramics and silicon. The results are discussed and compared to results obtained using conventional detectors.     

A fast experimental beam hardening correction method for accurate bone mineral measurements in 3D μCT imaging system

Bone mineral density plays an important role in the determination of bone strength and fracture risks. Consequently, it is very important to obtain accurate bone mineral density measurements. The microcomputerized tomography system provides 3D information about the architectural properties of bone. Quantitative analysis accuracy is decreased by the presence of artefacts in the reconstructed images, mainly due to beam hardening artefacts (such as cupping artefacts). In this paper, we introduced a new beam hardening correction method based on a postreconstruction technique performed with the use of off‐line water and bone linearization curves experimentally calculated aiming to take into account the nonhomogeneity in the scanned animal. In order to evaluate the mass correction rate, calibration line has been carried out to convert the reconstructed linear attenuation coefficient into bone masses. The presented correction method was then applied on a multimaterial cylindrical phantom and on mouse skeleton images. Mass correction rate up to 18% between uncorrected and corrected images were obtained as well as a remarkable improvement of a calculated mouse femur mass has been noticed. Results were also compared to those obtained when using the simple water linearization technique which does not take into account the nonhomogeneity in the object.     

3D reconstruction of prior β grains in friction stir–processed Ti–6Al–4V

The prior β grain structure and orientations in the central stir zone of friction stir–processed Ti–6Al–4V were reconstructed from measured α phase orientations obtained by three‐dimensional serial sectioning in a dual‐beam focused ion beam scanning electron microscope. The data were processed to obtain the α colony and β grain size distributions in the volume. Several β grains were individually analysed to determine the total number of unique α variants and the respective volume fractions of each. The analysis revealed that some grains experienced overwhelming variant selection (i.e. one variant dominated) whereas other β grains contained a more evenly distributed mixture of all 12 variants.     

Microstructure grooves with a width of less than 50 μm cut with ground hard metal micro end mills

Mechanical microengineering is an easy and cheap way to fabricate microstructures; for example, molds for injection molding or hot embossing. Restrictions remain in the selection of materials and in the minimum structure size. Especially for cutting microstructures in steel, these limitations included the lack of available small tools and the poor surface quality. We discovered that microstructures can be cut in both brass and stainless steel workpieces by using ground hard metal micro end mills. The minimum groove width achieved is less than 50 μm for cutting brass and about 100 μm for cutting stainless steel. Burrs are removed by a subsequent diamond-cutting step or electrochemical polishing, respectively. Our results represent the first step toward a microstructured, resistant, and cheap mold made of steel, which is then used for mass production of plastic microstructures.     

3-D vibration analysis of skew thick plates using Chebyshev–Ritz method

The free vibration characteristics of skew thick plates with arbitrary boundary conditions have been studied based on the three-dimensional, linear and small strain elasticity theory. The actual skew plate domain is mapped onto a basic cubic domain and the eigenvalue equation is then derived from the energy functional of the plate by using the Ritz method. A set of triplicate Chebyshev polynomial series multiplied by a boundary function chosen to satisfy the essential geometric boundary conditions of the plate is developed as the trial functions of the displacement components. The vibration modes are divided into antisymmetric and symmetric ones in the thickness direction and can be studied individually. The convergence and comparison studies show that rather accurate results can be obtained by using this approach. Parametric investigations on rhombic plates with fully clamped edges and completely free edges are performed in detail, with respect to the thickness-span ratio and skew angle. Some results known for the first time are reported, which may serve as the benchmark values for future numerical technique research.     

A flexure-based tool holder for sub-μm positioning of a single point cutting tool on a four-axis lathe

A tool holder was designed to facilitate the machining of precision meso-scale components with complex three-dimensional shapes with sub-μm accuracy on a four-axis lathe. A four-axis lathe incorporates a rotary table that allows the cutting tool to swivel with respect to the workpiece to enable the machining of complex workpiece forms, and accurately machining complex meso-scale parts often requires that the cutting tool be aligned precisely along the axis of rotation of the rotary table. The tool holder designed in this study has greatly simplified the process of setting the tool in the correct location with sub-μm precision. The tool holder adjusts the tool position using flexures that were designed using finite element analyses. Two flexures adjust the lateral position of the tool to align the center of the nose of the tool with the axis of rotation of the B-axis, and another flexure adjusts the height of the tool. The flexures are driven by manual micrometer adjusters, each of which provides a minimum increment of motion of 20 nm. This tool holder has simplified the process of setting a tool with sub-μm accuracy, and it has significantly reduced the time required to set a tool.     

A study of the effects of non-isothermal source conditions on the transfer of radiance temperatures in the temperature-wavelength domain from 180 to 30 000 μm K

The calibration of infrared pyrometers against a commercially available cylindroconical black-body system, operating in the range from 900 to 2900 K, forms the basis for the study presented in this paper.

The main features of the black-body system, in use at VSL, are briefly described. In practice, uniformity in temperature is difficult to achieve, especially at very high temperatures. For this reason, the effect of a non-uniform temperature distribution on the calibration accuracy was studied by model calculations, varying the relative temperature difference ΔT/T_b along the cylindrical wall of the cavity with respect to its bottom temperature T_b up to a maximum of 5%. Spectral emissivities = (λ, T_b) are calculated for a cylindro-conical geometry that approximates the real system. For some specific cases the effects for actually measured temperature distributions are also evaluated.

It is demonstrated that when transferring radiance temperatures from a lower wavelength (λ_s 0.6 μm), characterising the reference pyrometer, to the higher effective wavelength of the (spectral) infrared pyrometer to be calibrated (λ_x 6 μm), compensation of errors leads in general to a considerable reduction of the overall error, arising when neglecting the effect of departures from temperature uniformity, the reduction being more pronounced the higher the temperature.     

Reconstruction of 3D grain boundaries from rock thin sections,using an advanced polarised‐light microscopy method

Grain boundaries play a significant role in materials by initiating reactions and collecting impurities. Here we present FAGO (Fabric Analyser Grain boundary recOnstruction), a first step towards the automatic determination of three‐dimensional (3D) grain boundary geometry using polarised light. The trace of the grain boundaries from 2D rock thin sections are determined primarily from data acquired using an automatic fabric analyser microscope and the FAME software (fabric analyser‐based microstructure evaluation; Peternell and colleagues and Hammes and Peternell). Based on the Fabric Analyser G50's unique arrangement of nine differently oriented light sources the retardation can be determined independently for each light direction and at each pixel in the field of view. FAGO combines these retardation datasets for each individual pixel together with retardation profiles across grain boundaries, to determine the orientations of the boundaries. The grain boundary traces are then broken up into segments of equal orientation, using the profile‐obtained orientation data. Finally, a 3D grain boundary model is reconstructed. The data processing is almost fully automatic using the MATLAB® environment. Only minor manual inputs are required.     

A high-resolution electron microscopic study of defects in sodium β′″-alumina

A high-resolution electron-microscopic study of sodium β′″-alumina, a polytype of the more-widely-studied sodium β- and β″-alumina, has been undertaken using the 600 kV instrument at Cambridge University. Images revealed the loss of sodium-containing planes, which had caused crystals to collapse and shear into defect layers. A model for the structure of these defects is proposed, based on the use of computed images and by comparison with high-resolution images of silver β″-alumina.     

FE‐SEM observation of swelled seaweed using hydrophilic ionic liquid; 1‐butyl‐3‐methylimidazolium tetrafluoroborate

The method to observe the exact morphology of swelled seaweed as an example of biological material by field emission scanning electron microscopy (FE‐SEM) with the aid of hydrophilic ionic liquid (IL); 1‐butyl‐3‐methylimidazolium tetrafluoroborate is reported. Seaweed was first swelled in 3.5% NaCl solution and then treated with the IL and water mixture in 1:7 weight ratios and centrifuged to remove the excess IL solution. Thus treated seaweed maintained its morphology even at high magnification and did not show drying in the FE‐SEM chamber. This observation technique might be useful for various kinds of biological materials to be observed under FE‐SEM. Microsc. Res. Tech., 2013. © 2012 Wiley Periodicals, Inc.     

A tribological study of (2‐sulphurone‐benzothiazole)‐3‐methyl‐dibutyl borate

An organic compound containing S, N, B, and O was synthesised by reacting 2‐mercaptobenzothiazole and formalin in ethanol solution, the resulting product then being reacted with butanol and boric acid in toluene solution. The tribological performance of the novel compound when added to liquid paraffin was evaluated using a four‐ball tester and a ring‐on‐block machine. The relationship between performance and concentration was analysed, and the results show that the compound possesses good antiwear and load‐carrying abilities. The mechanism of action of the additive was investigated using X‐ray photo‐electron spectroscopy (XPS). Surface analysis indicated the formation of a protective film containing FeSO₄, an organo‐sulphur compound, FeS₂, borate, and an organonitrogen compound. This protective film formed during sliding processes contributes to the increase in wear resistance.     

Can scanning near‐field optical microscopy be compared with confocal laser scanning microscopy? A preliminary study on α‐sarcoglycan and β1D‐integrin in human skeletal muscle

The dystrophin–glycoprotein complex and the vinculin–talin–integrin system constitute, together a protein machinery, called costameres. The dystrophin–glycoprotein complex contains, among other proteins, also dystrophin and the sarcoglycans subcomplex, proteins playing a key role in the pathogenesis of many muscular dystrophies and linking the cytoplasmic myofibrillar contractile elements to the signal transducing molecules of the extracellular matrix, also providing structural support to the sarcolemma. The vinculin–talin–integrin system connects some components of the extracellular matrix with intermediate filaments of desmin, forming transverse bridges between Z and M lines. In our previous reports we always studied these systems by confocal laser scanning microscopy (CLSM). In this paper we report on the first applications of optical near‐field fluorescence microscopy to the spatial localization of α‐sarcoglycan and β1D‐integrin in human skeletal muscle fibres in order to better compare and test the images obtained with conventional CLSM and with scanning near‐field optical microscopy (SNOM). In addition, the analysis of the surface morphology, and the comparison with the fluorescence map is put forward and analyzed for the first time on human muscle fibres. In aperture‐SNOM the sample is excited through the nanometre‐scale aperture produced at the apex of an optical fibre after tapering and subsequent metal coating. The acquisition of the topography map, simultaneously to the optical signal, by SNOM, permits to exactly overlap the fluorescence images obtained from the two consecutive scans needed for the double localization. Besides, the differences between the topography and the optical spatial patterns permit to assess the absence of artefacts in the fluorescence maps. Although the SNOM represented a good method of analysis, this technique remains a complementary method to the CLSM and it can be accepted in order to confirm the hypothesis advanced by CLSM.