bluuebunny/crossref_metadata_2025 huggingface dataset:
Microautoradiography of diffusible ions in plant tissues: problems and methods
Authors: Sanderson J.
Published: August 2011

A procedure for the preparation of autoradiography of diffusible ions in plant tissues is described. Special procedures are necessary to support the tissue, and minimize redistribution of ions during the various stages of the preparation.

The principal features of the procedure are: rapid freezing of tissues after harvest; tissue embedding in methyl cellulose; section cutting in the cryostat at −20°C and section mounting onto photographic emulsion at −20°C; photographic exposure at −25°C; and finally development and mounting so that the autoradiograph lies over the section.

By comparing the grain densities on autoradiographs of plant sections with autoradiographs prepared from standard sources, of known specific activity, it is possible to evaluate the results quantitatively.
High voltage electron microscopy of the central nervous system in Golgi preparations
Authors: Palay Sanford L. , Chan‐Palay Victoria
Published: August 2011

High voltage electron microscopy was applied to sections of cerebellar cortex prepared by the Golgi method and embedded in epoxy resin. The silver chromate deposit in the cells consists of a fine, fibrillar meshwork upon which dense globules of various sizes may be superimposed. At the light microscope level finer images are obtained from specimens in which the fibrillar meshwork is incrusted with the fewest globules or are without incrustation. The varicosities in nerve fibres display light circular patches which represent the sites of synaptic junctions. The velamentous processes of neuroglial cells are also well demonstrated in these preparations.
The geometry of dislocation loops generated in α‐iron by 1 MeV electron irradiation at 550°C
Authors: Little E. A. , Eyre B. L.
Published: August 2011

Rectilinear dislocation loops are observed to grow in thin foils of α‐iron during 1 MeV electron irradiation at 550°C in the beam of a high voltage electron microscope. A diffraction analysis indicates that the loops have sides parallel to <100> directions and are pure edge with a <100> Burgers vector. Only interstitial type loops have been identified. The geometry of the loops can be accounted for by an established model of loop nucleation.
Micromagnetic structures in single crystal specimens of intermediate thickness studied by Lorentz microscopy
Authors: Harrison C. G. , Leaver K. D.
Published: August 2011

Stripe domains in (001) nickel occur in a variety of structures due to the large number of easy directions present. We have given below a tentative analysis of two of these structures, not previously studied by Lorentz microscopy. In particular it has been shown that, as used hitherto, the technique has not been a reliable method of observing stripe domains since some structures may be invisible until the specimen is tilted.
The effect of room temperature pre‐strain on grain boundary cavitation in Nimonic 8oA
Authors: Dyson B. F. , Henn D. E.
Published: August 2011

Specimens of the alloy Nimonic 80A have been pre‐strained 11% in tension at room temperature and then heated to 750°C for various times either with or without the application of stress. Examination by means of a high voltage electron microscope revealed large numbers of spherical submicron diameter cavities. The cavities all lay on grain boundaries but were inhomogeneously distributed; a grain boundary usually contained either many cavities or none at all. It is concluded that the cavitation is a direct result of decohesion at the grain boundaries during the pre‐straining and not due to any creep process.
A high voltage electron microscope study of recrystallization in copper alloys
Authors: Ray R. K. , Hutchinson W. B. , Besag F. M. C. , Smallman R. E.
Published: August 2011

Recrystallization and recovery have been studied in the high voltage electron microscope (HVEM) in pure copper and two copper‐phosphorus alloys following heavy rolling deformation. Prior to the nucleation of recrystallized grains, extensive subgrain growth occurs by sub‐boundary migration. In the alloys, the first recrystallized grains are observed to nucleate within deformed matrix regions of the same orientation. The orientations of these grains are in close agreement with the crystallographic texture of the metal at complete recrystallization. Growth selection of orientation does not appear to be important. In pure copper, cube‐oriented recrystallized grains form after those of some other orientations, but grow more rapidly and absorb these during growth.
In situ superplasticity experiments in the 1 million volt electron microscope
Authors: Naziri H. , Pearce R. , Brown M. Henderson , Hale K. F.
Published: August 2011

In situ tensile straining was carried out in the 1 million volt electron microscope in an attempt to clarify the mechanism(s) of superplasticity in the Zn/Al eutectoid alloy. The dynamic microstructural observations in the 1 MV microscope at 100°C at an m value of ∼0.3 were consistent with 100 kV observations of specimens thinned from the bulk and showed that under the experimental conditions used, the predominant mechanism is diffusion of the zinc‐rich (β) phase. It has been theoretically predicted by Lifshitz that diffusional creep and grain‐boundary sliding are inseparable. This is verified by these in situ observations.
In situ observations of the nitriding of tantalum
Authors: Swann P. R. , Thomas G. , Tighe N. J.
Published: August 2011

A gas reaction cell developed for the AEI EM7 has been used to observe directly the structural changes which occur when tantalum is heated in nitrogen. It is demonstrated that sharp images of the microstructure can be obtained from specimens at temperature while surrounded by a 5.5 mm thick layer of the reaction gas at a pressure of 35 torr. The usefulness of the in situ technique for studying gas/metal reactions is assessed.
Transmission electron microscopy at 2.5 MeV
Authors: Thomas Gareth , Lacaze J.‐C.
Published: August 2011

Measurements of penetration on silicon and austenitic stainless steel have been continued using the same criteria as previously reported (Thomas, 1968) up to 2.5 MeV using the 3 MeV Toulouse electron microscope. The results show that penetration increases to about 14 μm at 2.5 MeV for silicon although the curve starts to flatten out above 1.5 MeV, but no significant gain was found for stainless steel ≃2μm at 2.5 MeV). Primary knock‐on damage occurs readily in both materials. The critical voltages for 440 silicon and 422 tantalum were measured, and both found to be 1.4 MeV (± 10 kV).

Attempts to measure the variation of radiation damage with beam voltage in crystalline amino acids as indicated by the fading of diffraction patterns indicated that the damage rates apparently decrease as the voltage is raised to 2.5 MeV. However, it was not possible to determine this variation quantitatively because of difficulties in measuring the absolute values of the critical exposures.
A technique for the cytochemical demonstration in the electron microscope of glucose‐6‐phosphate dehydrogenase activity in erythrocytes of malaria‐infected animals
Authors: Theakston R. D. G. , Fletcher K. A.
Published: August 2011

A technique for the cytochemical demonstration in the electron microscope of glucose‐6‐phosphate dehydrogenase activity in malaria‐infected erythrocytes is described. Reaction product occurred as diffuse electron‐dense deposits of the formazan of MTT in the red cells. No activity was observed within the malaria parasites themselves, except in the contents of food vacuoles. The results of a series of control experiments are also described.
Methods for electron microscopy of the lamellated osmiophilic bodies of the lung
Authors: Schock C. , Pattle R. E. , Creasey J. M.
Published: August 2011

A study has been made of methods of fixing, staining, and embedding for electron microscopy the lamellated osmiophilic bodies (LOPBs) of the type II cells of the lung, which are probably connected with the lung surfactant. The preferred method for fixing and staining uses successively 2% glutaraldehyde, 1% osmium tetroxide, and a mixture of 10 vol M/40 lead nitrate with 10.5 vol M/60 potassium ferricyanide: all reagents are in cacodylate buffer. Unless the ferri‐cyanide is in excess, the sections show blotchy precipitation. The bodies are damaged by ethanol and 1:2 epoxypropane. Dioxan or (with a quick time schedule) acetone may be used as a dehydrant and thinner prior to Araldite embedding. Durcupan water‐soluble resin may also be used for dehydration and embedding; the preferred embedding mixture is: 5 ml resin A, 11.7 ml hardener B, 1.5 ml hardener C, and 0.4 ml plasticizer D, cured for 24–48 h at 60°C. Examination of LOPBs on a goniometer stage shows that they are composed of layers spaced at 4 nm, each lamella containing several such layers. Further evidence is adduced linking the LOPBs with the surfactant, and the application of the new methods to various problems involving the LOPBs is discussed.
Electron microscopy of cellulose in entire tissue
Authors: Cox Guy , Juniper Barrie
Published: August 2011

Microfibrillar structures can be seen in ultrathin sections of cell walls which have been post‐stained with uranyl acetate and lead citrate. This stain is removable by washing, suggesting that a physical mechanism is involved. It is suggested that these structures are the cellulose microfibrils of the wall; they appear to be 3.5 nm in diameter and not fasciated into larger units. Freeze‐etching of untreated tissue supports this conclusion. These two techniques seem to have many advantages over methods previously used for the study of microfibril arrangement and structure.
Permanent preparations of stained whole‐body sections using ‘Trycolac‘
Authors: Farebrother Derek A. , Woods Nicholas C.
Published: August 2011

Permanent preparations of whole‐body sections cut by Ullberg's technique can be prepared using a ‘spray‐on’ mountant (‘Trycolac’), which does away with the necessity for glass slides and coverslips. Routine staining methods can be used, providing the alcohol content of the stain is low. Glycerol is used for dehydration and clearing and a covering of ‘Trycolac’ is sprayed on.
Organic stains for electron microscopy
Authors: Stewart Murray
Published: August 2011

Evidence is presented to suggest that staining biological material prior to examination by electron microscopy with poly‐iodinated aromatic molecules may give contrast enhancement comparable to that obtained with uranyl acetate. This technique could enable the production of highly specific electron‐opaque stains analogous to the organic dyes employed in light microscopy.
Mica disc supports for particulate specimens for scanning electron microscopy
Authors: Desjardins Paul R. , Barkley Michael B.
Published: August 2011

Readily prepared mica discs have been found useful as supports for particulate specimens for the scanning electron microscope. The principal advantage is that they can be observed in the light microscope prior to further preparation for the scanning electron microscope.
Direct examination of chip formation during metal machining by scanning electron microscopy
Authors: Ramalingam S. , Bell A. C. , Black J. T.
Published: August 2011

New insights into the mechanics and morphology of chip formation in metal cutting have been obtained by direct observation of an orthogonal cutting process in progress in a scanning electron microscope. A specimen stage specifically designed for this purpose is described. This stage permits dynamic viewing of all accessible areas of interest at magnifications up to ×5000 during cutting. Permanent records are obtained by videotaping each experiment. A brief summary of results is given, and extensions of the technique of dynamic scanner observation are suggested.
Identification of elastic tissue in human skin viewed in the scanning electron microscope
Authors: Hunter John A. A. , Finlay Bryan
Published: August 2011

Elastic tissue has been identified in the scanning electron microscope by two independent methods. In one case specifically stained fibres were examined by both light and scanning electron microscopy and in the other chemically purified elastic fibres from ligamentum nuchae were studied.

Elastic fibres above 2 μm diameter were found to have a central amorphous core and an irregularly corrugated and undulating outer surface. This model is in good agreement with that proposed on the basis of previous transmission electron microscopic studies.
Estimation of number and diameter of isodiametric spherical particles in microtome sections
Authors: Davies A. G.
Published: August 2011

In microtome sections small invisible and unrecognizable fragments cause errors in estimation of particle number and diameter. These errors can be reduced by rejecting fragments whose apparent diameter is larger than the true radius of the particles. Formulae for the calculation of diameter and number are given and the size of the errors is examined.
The principles and practice of the weak‐beam method of electron microscopy
Authors: Cockayne D. J. H.
Published: August 2011

The weak‐beam method of electron microscopy enables the high resolution capabilities of the electron microscope to be used in the study of lattice defects. In this paper, the principles of the method are outlined and the experimental procedures for obtaining weak‐beam images are discussed. The experimental diffraction conditions necessary for obtaining quantitative results and the limitations of the method are summarized, and applications of the method to various problems in defect studies are reviewed.
Investigation of dislocation geometries in the diamond cubic structure
Authors: Ray I. L. F. , Cockayne D. J. H.
Published: August 2011

The weak‐beam technique of electron microscopy (Cockayne, Ray & Whelan, 1969) has been applied to the examination of dislocations in germanium. These are shown to be dissociated into partial dislocations with a separation in the edge orientation of 5.5 ± 1·0 nm. A value for the stacking‐fault energy of γ = 60 ± 8 mJ m−2 (erg cm−2) is deduced from the measured dissociation width as a function of orientation, using anisotropic elasticity theory.
Transmission electron microscope observations of extended and unextended dislocation nodes in Si and Ge/Si layers using the weak‐beam technique
Authors: Cullis A. G.
Published: August 2011

The transmission electron microscope (TEM) weak‐beam method was employed in a study of dislocation nodes both in deformed single crystal Si and at the Ge/(lll)Si interface. All intrinsic nodes were observed to be extended, and measurements of their geometry in Si were used to determine the intrinsic stacking‐fault energy. However, only approximately half of the extrinsic nodes were extended and this was interpreted in terms of an energy barrier to node extension.
Four‐fold dissociations of super‐lattice dislocations
Authors: Crawford R. C. , Ray I. L. F. , Cockayne D. J. H.
Published: August 2011

The weak‐beam technique of electron microscopy (Cockayne, Ray & Whelan, 1969) has been used to image the four components of super‐lattice dislocations in a range of iron‐aluminium alloys, between 26 and 30 at % aluminium, with D03‐type long‐range order. Values of the first and second nearest neighbour interaction energies have been measured from the observed separations of the components.
Computer simulation of weak‐beam images of extended dislocations in copper
Authors: Perrin R. C. , Savino E. J.
Published: August 2011

Weak‐beam electron microscope images of a dissociated edge dislocation in copper have been calculated using core configurations computed both from an atomic model and elasticity. These images show two distinct intensity peaks the separation of which is considerably greater in those derived from the atomic model. A comparison of these images with recently reported experimental observations suggests that the intrinsic stacking fault energy in copper is at least 70 mJ m−2 (erg cm−2).
Mechanisms of contrast and image formation of biological specimens in the transmission electron microscope
Authors: Burge R. E.
Published: August 2011

A systematic account is given, in relation to amorphous and paracrystalline (especially biological) material, of electron scattering and image formation for the range of electron energy from 50 to 100 keV. Contrast mechanisms in both elastic and inelastic scattering are discussed and image formation is considered under both scattering contrast and phase contrast conditions.
Phase contrast electron microscopy of biological materials
Authors: Unwin P. N. T.
Published: August 2011

The properties of two electron microscope phase contrast imaging methods are compared. The first is the conventional bright‐field method in which dark phase contrast is created by defocusing; the second is a phase plate method in which bright phase contrast is created by means of a suitably shaped electric field.

Using some negatively stained biological specimens which have a well‐known repeating structure as the test object, it is shown that the phase plate method has some important advantages over the bright‐field method. Its contrast transfer characteristics are such that it can provide a more faithful representation of the high resolution detail in the object. Moreover, by producing bright, rather than the normal dark, phase contrast it is able to simultaneously enhance the detail in the specimen and weaken the detail in the stain; this latter property enables the method to display information about the specimen that it would not be possible to detect with the bright‐field method.
Aspects of dark‐field electron microscopy
Authors: Perrier Frantz
Published: August 2011

This paper endeavours to give all the necessary practical information to operate with the contrast‐stop method as far as fairly thick objects (from 10 to 100 nm) are concerned; including working diagrams, practical operation out of the stops and also to define the range of application of this method. It is shown that false detail is not to be feared in these rather thick specimens, which is not at variance with the results obtained by Hanszen (1967, 1969) and Thon (1966) as far as very thin specimens are concerned.
Recent developments in analytical electron microscopy
Authors: Chandler John A.
Published: August 2011

Recent years have seen the way in which the analytical electron microscope has been applied to problems involving thin specimens in metallurgy, mineralogy and many branches of biology. The limits of sensitivity have been explored and its potential usefulness in these fields investigated.

Problems concerning the interaction of electrons with the specimen are discussed in relation to the correct choice of operating conditions and specimen preparation.

In biological work, frozen sections provide new information about subcellular elemental localization of mobile electrolytes, while analysis of tissue prepared by conventional means is used to detect physiological levels of some naturally occurring elements.

Examination of mineral dispersions provides analysis of particles just 10 nm thick, visible only in transmission electron microscopy, and further work with thin metal foils confirms the value of high resolution transmission imaging as a complementary facility to micro‐analysis.

Work has been done to investigate the possibilities of improving sensitivities both by changing operational parameters and instrumental design, and the value of quantitation in thin specimen analysis explored.
Optimization of stereoscopic and high angle tilting procedures for biological thin sections
Authors: Willis Robin A.
Published: August 2011

The use of stereoscopic and high angle tilting facilities in electron microscopes to optimize information retrieval from thin sections of biological material is reviewed. Some of the technical problems involved are analysed in detail and problems of interpretation of stereo‐pairs and the necessity of ‘learning to see depth’ in such unfamiliar visual situations is emphasized. A table suggesting methods of optimizing various parameters is presented. An indication of some of the unanticipated results which have arisen from the work and of the consequent reservations about currently accepted routine evaluations of biological sections, especially as regards their thickness, are considered. The validity of serial section reconstruction at the 10–50 nm level needs careful consideration.
High voltage microscopy of biological specimens: Some practical considerations
Authors: Beeston B. E. P.
Published: August 2011

Techniques are now available for the preparation of specimens giving excellent image contrast at high voltages, either by block‐staining, staining an already embedded block or by post‐staining. Further techniques are being developed for selectively staining certain features of the specimen thereby reducing the stain content of the specimen and permitting the examination of very thick specimens at adequate resolution.

Specimens having inadequate contrast, particularly preparations of macro‐molecules, may be imaged in one of the various dark‐field modes, giving a considerable enhancement in image contrast on the viewing screen. Further enhancement of the recorded image contrast can be achieved by exposing the photographic plate to a much higher density than is normal.

The complexity of image detail which occurs when the many features of a thick specimen are contracted into a two‐dimensional micrograph may be elucidated by stereo‐imaging techniques. Care must be taken to optimize the stereo‐tilt angles, the orientation of the tilt axis and the register of the two micrographs in the stereo‐pair, for the maximum depth in the specimen to be observed.
Quantitative photogrammetric analysis and qualitative stereoscopic analysis of SEM images
Authors: Boyde A.
Published: August 2011

This paper is concerned with methods for obtaining three‐dimensional (3D) information from scanning electron microscope (SEM) images. It may come as a surprise that this topic merits separate discussion, because, it is generally accepted that every SEM image of a rough surface using the secondary electron emission mode gives apparent 3D information. This is true to some extent, since SEM images are comparable with photographs of rough surfaces illuminated with light coming from one direction, but our main concern here will be with 3D visualization from stereoscopic pairs of images. However, there are other ways in which 3D information may be obtained, even in the SEM, and these are considered first. The remainder of the paper is largely concerned with the use of stereoscopic parallaxes, whether or not they are visualized as such. Commonly used methods for viewing stereo‐pairs are described, followed by simple methods for measuring stereoscopic parallaxes. Formulae for reducing linear measurements in the two photographs to real height differences are given.
Field‐ion microscopy of interphase interfaces
Authors: Hildon A. , Howell P. R. , Youle A. , Taunt R. J. , Page T. F. , Ralph B.
Published: August 2011

This paper demonstrates the use of the analytical techniques, developed in the companion paper, to determine interfacial parameters from a variety of multiphase structures. It is shown that accurate orientation relationships and the structure of unknown phases can be derived from this approach. In some cases assumptions have to be made concerning some of the interfacial parameters, but these assumptions can always be checked from the self‐consistency of the full analysis.
Scanning electron microscopy of transfer cells: A new method for preparation of plant tissues
Authors: Chambers T. C. , Hamilton Catherine D.
Published: August 2011

The xylem transfer cells of the wheat node have been examined in the scanning electron microscope following a preparative procedure of partial embedding in epoxy resin sufficient to prevent delicate structures from collapse during polymerization and scanning electron microscopy examination.
A method of microscopic evaluation of effects of demineralizers on complex carbohydrates of rat tissues
Authors: Weatherford Thomas W. , Mann Wallace V.
Published: August 2011

Mandibles, salivary glands, trachea, costal cartilages and kidneys of young adult, male, sibling rats were fixed in chilled, modified Lavdowsky's solution. The mandibles and blocks from the other tissues were then exposed to one of four demineralizers: Win 3000 and 20% formic acid, Greep's solution, 5% nitric acid and 10% EDTA. For comparison with demineralized tissues, a replicate set of tissues, other than mandibles, from each animal was processed without exposure to demineralizers.

Paraffin sections of all tissues were stained with Alcian blue at pH 2–6 and at pH 0.5, with PAS and with Mowry's revised colloidal iron. The effect of the demineralizers on the staining intensities, tinctorial qualities, and cellular details were studied by pairwise comparison of each slide of a common tissue‐stain group with every other slide of the group. A comparison microscope was used and a numerically expressed preference ranking was obtained for each slide.

Analysis of variance tests and Duncan's new multiple range tests were applied to determine the preferred demineralizers. Scoring results of the mandibles indicated that Win 3000 and 20% formic acid was either the preferred demineralizer or in each case not significantly different from the preferred one. Reliability of the findings was reduced because of inconsistencies among soft tissues not exposed to any demineralizer.
The application of optical microscopy and electron‐probe microanalysis to a mineral processing topic—the high‐temperature reduction of ilmenite
Authors: Jones D. G. , Stephens J. F.
Published: August 2011

A combination of optical microscopy and electron‐probe microanalysis (EPMA) has been used to study the phases which occur during reduction of the mineral ilmenite to metallic iron and titanium oxides. Quantitative EPMA of natural and chemically‐processed minerals frequently suffers from variable specimen conductivity caused by the porosity and varying texture of the mineral grains. It has been found, however, that the element concentration ratios are largely independent of porosity and texture and can be used to identify the phases with precision.

The microprobe has revealed some hitherto unsuspected effects arising from small concentrations of magnesium and manganese dissolved in the ilmenite lattice. The morphology and composition of the reduced product are in fact controlled by the way in which these impurity solutes segregate within the mineral grains during reduction.
Wetting agents for biological electron microscopy
Authors: Gregory D. W. , Pirie B. J. S.
Published: August 2011

When preparing specimens for shadowing it is frequently necessary to add a wetting agent to aqueous suspensions to enable them to spread on hydrophobic support films. However high‐resolution platinum/carbon shadowing is capable of visualizing wetting agent molecules, so the effect of added wetting agent on the final appearance of the specimen must be considered. To facilitate interpretation of the specimen fine detail the background should be smooth and flat. This is achieved by using the concentration of wetting agent which produces a close‐packed monolayer of wetting agent molecules after drying. Bacitracin is a better wetting agent for use when shadowing than the commonly‐used bovine serum albumin (BSA). Because its molecules are smaller than those of BSA, the size of the background granularity is smaller, which facilitates interpretation of the specimen, and the layer of wetting agent molecules is thinner, so small specimen details are less likely to be buried and rendered invisible to the shadowing process. These points are described in detail and illustrated with the help of models where necessary.
Profiles of jet polished electron microscope specimens
Authors: Ali H. O. , Adams R. , Wann P. R. S.
Published: August 2011

The cross‐sectional profiles of jet polished electron microscope disc specimens have been measured as a function of (1) jet diameter, (2) electrolyte velocity, (3) jet to specimen distance, (4) lip thickness of clamping disc and (5) material used for the clamping disc. It is found that the optimum values of these parameters varies according to the material/electrolyte combination. However, a fault‐finding table has been devised which enables the optimum conditions to be found rapidly for any material.
Stereological correction procedures for estimating true volume proportions from biased samples
Authors: Mayhew T. M. , Cruz L.‐M.
Published: August 2011

Experimental evaluation of component volume proportions may require the introduction of a component‐biased sampling approach in order to facilitate the unequivocal identification of cell type. Such is certainly the case for macrophages, neurons and cells of the lymphocyte‐plasma cell family. With these systems, mathematical corrections are necessary so that reliable morphometric information can be computed from the biased data. The type of correction procedure adopted will depend, to some extent, on the spatial relationships between the component of interest (nucleus, nucleolus) and its containing volume (cell, nucleus). In this communication, correction formulae for the special cases of spatial concentricity and eccentricity are derived. Their usefulness is illustrated by application to a number of well‐known model systems, and a reference table for obtaining the appropriate true volume proportion from observed (biased) data is provided. The reliability of the component‐biased morphometric approach is thereby increased.
Metal oxide systems studied by analytical electron microscopy
Authors: Fursey Anita , Kent B. , Saunders S. R. J.
Published: August 2011

Detailed analysis of thin oxide films is providing a better understanding of the mechanisms of oxidation and corrosion. The analytical electron microscope used to study thin layers on iron‐chromium alloys has shown that even after oxidation times as short as 10 min at 600°C the ratio of iron to chromium in the surface oxide will be different from that in the underlying alloy and discrete nodules may well have a higher iron content than the protective film in which they are growing. Progressive changes in different morphological features of the oxide films grown at varying times and temperatures can be followed by micro‐probe analysis in EMMA 4. Results obtained from the oxidation in air of iron with 10% and iron with 20% added chromium are presented.
The use of the analytical electron microscope EMMA‐4 to study the solute distribution in thin foils: some applications to metals and minerals
Authors: Lorimer G. W. , Razik N. A. , Cliff G.
Published: August 2011

Results are reported concerning the application of the analytical electron microscope EMMA‐4 to the quantitative analysis of thin foils. A simple procedure by which the observed characteristic X‐ray intensities from a thin foil sample can be converted into chemical weight fractions is described. This technique has been applied to study the partitioning of manganese between the carbide and ferrite phases during the isothermal decomposition of a eutectoid steel. The results show that it is possible to analyse the manganese content of the ferrite and carbide phases independently and the partitioning of manganese to the carbide phase has been followed as a function of time at the isothermal transformation temperature.

The analysis of a calcium‐rich pyroxene which contains calcium‐poor precipitate lamellae is described. The results are consistent with the precipitation of a pigeonite, poor in calcium and rich in iron, from an augite host.

The third example is the ‘in situ’ analysis of nitride precipitates in a stainless steel. The precipitates are known to contain either titanium or chromium, but it is impossible from morphological or diffraction evidence to differentiate between the two types of nitride. Combined electron microscopy and microprobe analysis is shown to be a rapid and positive means of identifying the precipitates.
Localization of acid and alkaline phosphatase
Authors: Bauer Heinz , Sigarlakie Elsje
Published: August 2011

Ultrathin frozen sections of glutaraldehyde fixed yeast cells have been successfully used for the demonstration of acid and alkaline phosphatase.

Acid phosphatase was localized over the cell wall of both the mother cell and the bud as well as over the newly forming cross wall (septum). Cytoplasmic vesicles (vacuoles, lysosomes?) located close to the cell wall showed a positive reaction for acid phosphatase as well. After 3 h glutaraldehyde fixation an activation of the nuclear acid phosphatase was observed. Lead precipitates were predominantly found over the nucleolar material of ‘resting’ and budding cells.

Alkaline phosphatase could be demonstrated in the ‘yeast‐mesosome’ and within the plasmalemma invaginations. After separation of the bud, small vesicles, probably derived from the endoplasmic reticulum showed a strong positive reaction for alkaline phosphatase.

In frozen sections incubated for alkaline phosphatase, non‐specific lead precipitates were found in the nucleus and along the plasmalemma invaginations.
Growth of ice crystals in frozen specimens
Authors: Nei Tokio
Published: August 2011

The formation of ice crystals, which might be the possible artefact in cryo‐techniques for electron microscopy, was examined during the rewarming process of rapidly frozen erythrocytes.

Intracellular ice formation, which is usually found in cells suspended in saline by rapid freezing, was inhibited by the addition of 30% glycerol. When such glycerinated cells, having no ice crystals at liquid nitrogen temperature, were rewarmed to higher temperatures above − 80° C, recrystallization of ice occurred. Ice particles became visible within the cells even at − 80°C and grew larger with a temperature rise.

From the results obtained in the morphological and physiological investigations, it also became evident that the recrystallization of ice appeared prior to the increase in haemolysis during the rewarming process.
Performance characteristics of a microreflectometer for measuring autoradiographic grain density
Authors: Entingh Dan
Published: August 2011

An incident light microphotometer for measuring reflectance has been constructed from Wild M20 microscope components and Photovolt 520‐M photometer. The instrument measures autoradiographic silver grain densities in either small regions (5–30 μm diameter) or large regions (25–150 μm diameter) with an accuracy of greater than 90% compared to visual grain counts. Grain density measurements can be made in the presence of histological stains. The device speeds the collection of autoradiographic data by a factor of 10 or more, and can be used for 6 or 7 h per day without producing significant strain on the operator.
Preparation of wood specimens for the scanning electron microscope
Authors: Exley R. R. , Butterfield B. G. , Meylan B. A.
Published: August 2011

Techniques used by the authors for preparing wood samples for the scanning electron microscope are detailed. Topics covered include: procedures for cutting, cleaning, dehydrating, mounting and storage of the specimens.
Some observations on knife properties and sectioning mechanics during ultramicrotomy of plastic embedding media
Authors: Helander Herbert F.
Published: August 2011

The edge and the trough surface of ultramicrotome glass knives were studied by interference light microscopy and electron microscopy. Particular attention was paid to the region which is commonly used for microtomy; here the bevel angle was calculated to 50–55°. The edge was serrated, even in the region used for sectioning, where the indentations measured up to 20 nm in depth. The imprints of edge serrations left on the sectioned material (Vestopal) were studied by electron microscopy. Different embedding materials displayed different degrees of compression when sectioned in the ultramicrotome: with Epon the compression was less than 10%, whereas methacrylate was compressed by more than 30%. Different trough fluids were tested: aqueous solutions of ethanol, acetone and dioxane gave less compression than pure water. However, the compression could also be reduced by exposing compressed Vestopal sections to these solutions. The direction and magnitude of the cutting force was measured with transducers during the sectioning of Vestopal blocks. Thus ∼0·3 μm wide sections cut at a feeding rate of 80 nm and a speed of 2 mm/sec required a cutting force of about 8 mN. The direction of the forces was 8–15° from the plane of cutting.
Histoquantitation using computer data cards
Authors: Meinhard Elizabeth
Published: August 2011

This paper describes a new approach to histoquantitation which makes fuller use of the visual expertise of the microscopist, provides a permanent visual two‐dimensional coded record, and allows mathematical exploitation of the collected data.

The microscopic image of the specimen is traced directly onto computer data cards. A simple sketch records accurate quantitative data for areas and boundaries of as many components of the image as required, in a matrix of 840 points. Spatial relationships are maintained. The matrix is transferred by an Optical Mark Data Card Reader into a computer for extraction of a maximum of information.
A method for the fabrication of thin‐foil apertures for electron microscopy
Authors: Schabtach Eric
Published: August 2011

A method is described for making thin foil apertures by piercing a plastic film with a heated microneedle and subsequently metallizing the plastic film by vacuum evaporation. Apertures can be made with diameters from 5 μm to 100 μm. They are round and very sharp‐edged and can be made in various patterns and combinations of sizes.
An anti‐roll plate for flattening of ‘dry’ ultrathin frozen sections
Authors: Hellström Sten , Sjöström Michael
Published: August 2011

A simple anti‐roll plate has been designed for use in preparing ‘dry’ ultrathin frozen sections. The plate, which is made of a strip of a glass cover slip and attached close to the edge of a glass knife, prevents curling of the sections.
The etching of thick Araldite‐embedded sections for scanning electron microscopy
Authors: Thurley Kenneth W. , Mouel Walter C.
Published: August 2011

A rapid method of correlation of information from the images of the scanning electron microscope, the transmission electron microscope and the light microscope, from biological tissues embedded in epoxy‐resin. It allows a perception in depth at the SEM level of areas already defined by the other two modes. One micrometre sections of Araldite‐embedded tissues are cut, dried onto glass cover slips, and the embedding medium etched away, with a saturated solution of NaOH in absolute ethanol. After washing in absolute ethanol and distilled water the resultant osmicated ‘skeleton’ is coated and viewed in the SEM.
Routine collection of flat large‐area sections for electron microscopy as applied to a detailed study of axon dimensions
Authors: Arbuthnott Elinor R.
Published: August 2011

A convenient method is described for the routine collection of flat large‐area sections for electron microscopy. The technique, which is of general application, has been applied to a survey of the dimensions and location of selected axons within a whole nerve.
A method for accurately determining lattice parameters using electron diffraction in a commercial electron microscope
Authors: Lodder J. C. , Berg K. G. v.d.
Published: August 2011

In this paper an electron diffraction method is discussed by which the lattice constants of polycrystalline thin films can be accurately determined (0.1%).

The method involves the sequential examination of a standard material and the unknown material mounted on separate grids. The error which can arise through the possible difference in height of the two grids can be corrected by means of height and tilting adjustments. The analytical approach of the dependence of the camera constant on the ring diameter as given in literature appears to be insufficient. An experimental correction factor for this dependence is introduced. The accuracy of this method, and the influence of relevant sources of error, are discussed and explained in terms of some experimental results.
Complementary freeze‐etch replicas by adapting the Denton apposed specimen tooling to the Balzers unit
Authors: Demsey Anthony , Frank Hermann , Schwarz Heinz , Henne Erhard
Published: August 2011

The complementary replica technique for freeze‐etching was developed to help answer certain questions concerning fracture planes and contamination artefacts. In order to obtain this capability in our freeze‐etch laboratory we have chosen, for scientific as well as financial reasons, to adapt the Denton apposed specimen tooling to our already‐existing Balzers facilities. The method employed for this adaption and results obtained with it are reported herein.
Beam‐induced loss of organic mass under electron‐microprobe conditions
Authors: Hall T. A. , Gupta B. L.
Published: August 2011

The loss of mass from biological tissue sections during electron microprobe irradiation was observed by means of a silicon semiconductor detector set to record continuum X radiation. The rate of loss depended on several factors, conspicuously on specimen temperature. In accord with earlier observations with non‐focused beams, most of the ‘immediate’ loss occurred within an electron dose of the order of 10−10 C/μm2.
Importance of controls for the demonstration of carbohydrates in electron microscopy with the silver methenamine or the thiocarbohydrazide‐silver proteinate methods
Authors: Courtoy R. , Simar L. J.
Published: August 2011

The periodic acid‐silver methenamine method and the periodic acid‐thio‐carbohydrazide‐silver proteinate method have been used for the demonstration of carbohydrates in various plant and animal tissues. The comparison of the results obtained with both methods indicates that the periodic acid‐thiocarbohydrazide‐silver proteinate method is more specific and gives better quality pictures than the PA‐silver methenamine method. It is pointed out that adequate controls are necessary for a valid cytochemical interpretation of the results. With the PA‐thiocarbohydrazide‐silver proteinate method, no positively stained structure can be assumed to be of carbohydrate nature unless it is found unstained in non‐oxidized thiocarbohydrazide‐silver proteinate treated sections. Both methods have their limitations and do not demonstrate the total carbohydrate amount in tissue sections.
Detection of haemoglobin in red cells of Xenopus laevis by immunofluorescent double labelling
Authors: Jurd Richard D. , Maclean Norman
Published: August 2011

Red cells from metamorphosing Xenopus laevis were treated with a fluorescein‐conjugated guinea‐pig antibody to Xenopus tadpole haemoglobin, a rabbit antibody to Xenopus adult haemoglobin, and a rhodamine‐conjugated goat anti‐rabbit globulin antibody. The presence of both red and green fluorescence in some cells indicated that at metamorphosis a proportion of the red cells contained both adult and tadpole haemoglobins.
Lung fixation by perfusion—a simple method to control the pressure in the perfusion circuit
Authors: Assimacopoulos A. , Kapanci Y.
Published: August 2011

A simple method which maintains the hydraulic pressure at a constant level during fixation of organs by perfusion is described. This is obtained by means of a glass tube interposed between a peristaltic pump and the organ to be perfused, the specially designed glass tube acting as a safety valve. The efficiency of the system is tested by electromagnetic pressure recording.
Selection of the best method in stereology
Authors: Weibel Ewald R.
Published: August 2011

A brief historical survey of the origin of stereological methods is followed by a synoptic characterization of their nature. They fall into two main classes depending on the structural models at their origin. Those methods which aim at a size analysis of the particle population provide more information, but are more subject to limitations, than those which estimate bulk parameters of the structure as a whole. The choice between the different methods must be guided by a clear definition of the most informative parameters and of the geometric properties of the structure.
Improved cathodoluminescence microscopy
Authors: Bond E. F. , Beresford Diana , Haggis G. H.
Published: August 2011

Methods are described for the use of a sensitive photomultiplier, improved photomultiplier voltage supply circuitry, quartz lens and a half‐parabolic mirror to increase the amount of light collected from a luminescing specimen examined in the scanning electron microscope. The combined effect of these improvements increases cathodoluminescence detection twenty times. Sources of background light are investigated and recommendations are made for decreasing this noise component, which seriously limits sensitivity of cathodoluminescence signal detection.

An appendix describes the natural luminescence (autoluminescence) of plant tissues under the impact of the electron beam. The biological applications of this mode of scanning microscopy have been little explored, but autocathodoluminescence is found to be of wide occurrence in plant tissues.
The control of the configuration of nucleic acid molecules deposited for electron microscopy, by ionic bombardment of carbon films
Authors: Highton Peter J. , Whitfield Morag
Published: August 2011

DNA molecules bind to carbon films placed on the surface of aqueous solutions and are extended and oriented by the flow forces on removal from the surface. The number of molecules bound increases with ionic strength suggesting that the bonding is hydrophobic.

If carbon films are subjected to ionic bombardment in a flow discharge, they can be modified so that molecules bind strongly, and are deposited in a tangled, random‐coil configuration, uninfluenced by flow forces. The strength of binding is decreased by reducing the discharge time and current, and by addition of salt (NaCl), suggesting that the bonding is ionic. When weakly bound, the molecules are condensed by ethanol. The strength of binding can be adjusted so that the molecules are largely untangled by flow, and in a form suitable for the mapping of bound enzymes, or other identifiable markers.

Aggregation and extension of molecules, that occur on unmodified films, and the binding of cytochrome c, that occurs in methods based on that of Kleinschmidt & Zahn (1959) are avoided. Both double and single‐stranded molecules can be deposited and seen after rotary shadowing with Pt. The discharge can be performed in a vacuum produced by a standard oil‐filled rotary pump.
Problems in the use of selective optical spatial filtering to obtain enhanced information from electron micrographs
Authors: Taylor C. A. , Ranniko J. K.
Published: August 2011

Possible hazards in the use of optical spatial filtering applied to the analysis of electron micrographs are illustrated. Two experiments show how information is either lost or spuriously introduced into reconstructed images of micrographs when (a) only one part of a two‐part micrograph image is reconstructed and (b) when noise is eliminated from the reconstruction.
Staining of biological material for the scanning electron microscope
Authors: Carr K. E. , McGadey J.
Published: August 2011

A histochemical technique has been used in scanning electron microscopy to localize specific areas in biological material. The specific contrast introduced by the staining procedure depends on the interaction of complex variables such as the coating material, the accelerating voltage and the use of a primary or secondary electron signal.
Adaptation of a rotary microtome for sectioning with glass knives
Authors: Fineran B. A. , Johnson I. A.
Published: August 2011

A glass knife holder for sectioning plastic embedded material, using a Jung microtome, is described. The knife holder and the device to clamp the LKB specimen chuck on the microtome are designed to fit the instrument without its modification.