bluuebunny/crossref_metadata_2025_split huggingface dataset:
Resin development for electron microscopy and an analysis of embedding at low temperature*
Authors: Carlemalm E. , Garavito R. M. , Villiger W. 
Published: August 2011

SUMMARYWe have tested a wide range of acrylate and methacrylate resin formulations and have concluded that embedding resins can be developed which are usable within a broad range of environmental conditions. To demonstrate this versatility, we have designed two highly‐cross‐linked resins, one polar and the other nonpolar, that are usable to temperatures from 243 to 223 K. Both of these resins formulations, which are now commercially available, show that systematic experiments can be easily done to study the effects of environmental parameters, such as water content, temperature, or resin polarity, on biological material during embedding. Using these resins and aldehyde‐fixed protein crystals, it can be shown that low temperature minimizes the loss of molecular structure to an extent that is not often obtainable with conventional methods of dehydration and embedding. Embedded crystals of aspartate aminotransferase still retained molecular order to 0·6 nm. Embedded crystals of catalase show X‐ray diffraction maxima out to 0·8 nm. When sectioned, catalase revealed stain‐limited electron and optical diffraction patterns to 2·5 nm. Nevertheless, our work clearly demonstrates that low temperature embedding procedures are superior and that versatile, general purpose resins can be designed to take advantage of this fact.
Estimating the effect of etching agents on plastic sections
Authors: Horobin Richard W. , Proctor John 
Published: August 2011

SUMMARYThe effects on epoxy embedding media of certain etching procedures have been studied in two ways. Firstly the resin present in thick sections, before and after etching, was demonstrated by staining the plastic with Sudan Black B. Secondly the change in weight of plastic blocks after their exposure to etching procedures was measured. Methods involving halogens in acetone, alkoxides in ethanol or in methanol‐benzene, and performic acid resulted in gross removal of plastic. Additionally, in keeping with the widespread belief that etching may increase permeability of plastic sections to stains, most of the etch methods studied, including organic solvents alone, did remove some plastic from the blocks.
Investigation of fading and recovery of fluorescence intensity at 73·5 K
Authors: Tiffe H.‐W. , Hundeshagen H. 
Published: August 2011

SUMMARYFading and recovery of Af‐feulgen stained sperm heads are investigated at 73·5 K. The fast fluorescence signals are measured and stored by two coupled transient recorders. 100% recovery was reached after a dark time of 3 s. This shows that the photodecomposition is mainly caused by change of the probability density of energy level and not by destruction of the chromophore groups. The recovery effect allows measurement of the fluorescence intensity of the same sample more than 50 times. Therefore the unaffected spectrum can be measured directly, provided that between the short‐term measurements at constant wavelength an appropriate dark phase has been put into operation.
Low‐temperature X‐ray microanalysis of the differentiating vascular tissue in root tips of Lemna minor L
Authors: Echlin Patrick , Lai C. E. , Hayes T. L. 
Published: August 2011

SUMMARYThe fracture faces of bulk‐frozen tissue offer a number of advantages for the analysis of diffusible elements. They are easy to prepare, remain uncontaminated, and, unlike most frozen‐hydrated sections, can be shown to exist in a fully hydrated state throughout examination and analysis. Root tips of Lemna minor briefly treated with a polymeric cryoprotectant are quench frozen in melting nitrogen. Fractures are prepared using the AMRAY Biochamber, lightly etched if necessary to reveal surface detail and carbon coated while maintaining the specimen at 110 K. The frozen‐hydrated fracture faces are analysed at 110 K using the P/B ratio method which is less sensitive to changes in surface geometry and variations in beam current. The method has been used to investigate the distribution of seven elements (Na+, Mg++, P, S, Cl−, K+ and Ca++) in the developing vascular tissue of the root tip. The microprobe can measure relative elemental ratios at the cellular level and the results from this present study reveal important variations in different parts of the root. The younger, more actively dividing cells, appear to have a slightly higher concentration of diffusible ions in comparison to the somewhat older tissues which have begun to differentiate into what are presumed to be functional vascular elements.
On the size of small protein particles determined by electron microscopy of unidirectionally shadowed freeze‐etched preparations
Authors: Schmidt D. G. , Buchheim W. 
Published: August 2011

SUMMARYThe influence exerted by the thickness of the deposited metal layer and the ionic strength of the solution on the apparent size of particles of bovine serum albumin in unidirectionally shadowed freeze‐etch preparations of spray‐frozen specimens was investigated. It appeared that the size increase due to shadowing is nearly twice the thickness of the deposited metal layer. Apparent particle size was shown to increase linearly with the inverse square root of the ionic strength of the solution. At ionic strength 0·001 the particles appeared about 30% larger than at infinite ionic strength.
On‐line digital computer techniques in electron microscopy: general introduction
Authors: Smith K. C. A. 
Published: August 2011

SUMMARYDigital computers may be interfaced to electron microscopes in a variety of ways to provide on‐line facilities for the acquisition, storage, processing and display of data. The data may be in the form of either images or spectra, the former necessitating a large amount of storage or memory space within the computer system. This has led to the development of special‐purpose digital memory systems, known as framestores, capable of holding one or more complete TV size images. Framestores can be used for both image acquisition and image display at normal TV frame rates. On‐line computer systems with, and without, framestores are being applied increasingly in scanning and conventional electron microscopy. The introduction of new computer technology, coupled with a reduction in costs, appears likely to accelerate this trend.
Multiple signals in STEM
Authors: Burge R. E. , Browne M. T. , Charalambous P. , Clark A. , Wu J. K. 
Published: August 2011

SUMMARYA brief description is given of the eight image signals that can be simultaneously recorded in digital form using an appropriate data recording system and high resolution STEM; this involves a set of detectors covering bright and dark field. Details are provided of the collection and manipulation of image data for STEM, including on‐line estimation of off‐focus distance, on‐line assessment of astigmatism, on‐line signal to noise ratio in any or all detectors; examples of all three procedures are given. The use of the data recording system is discussed together with some aspects of image display, including the use of colour. A method is introduced of providing, using the appropriate hardware, a prescribed colour map (image intensity converted to colour) using replaceable PROMS.Two off‐line procedures are considered. One is the Karhunen‐Loeve transform and its application to bringing together the information contained within the multiple signals, and the other is the discrete cosine transform by which the storage problem for digital image storage may be reduced, according to the image, by between 15 and 30 times.
An integrated approach to scanning microscope data acquisition
Authors: Jones A. V. , Unitt B. M. 
Published: August 2011

SUMMARYThis paper describes an integrated data acquisition system built for a cryogenic STEM which is currently under construction in the authors' laboratory. Advantage was taken of modern digital and microprocessor techniques to construct a versatile and expandable system in the minimum time. The system is able to operate in conjunction with an external image processing computer or as an independent system with built‐in image processing facilities.
Topographical analysis in the SEM using an automatic focusing technique
Authors: Holburn D. M. , Smith K. C. A. 
Published: August 2011

SUMMARYA novel technique for automated topographical analysis in the SEM has been investigated. It utilizes a 16‐bit minicomputer arranged to act as an automatic focusing unit. The computer is coupled to the objective lens of the microscope, by means of a digital to analogue converter, and may regulate the excitation of the lens under program control. Further digital‐to‐analogue converters allow the computer to act as a programmable scan generator by applying ramp waveforms to the scan amplifiers, permitting the beam to be swept over a small sub‐region of the field of interest. The video signal is sampled and applied to an analogue‐to‐digital converter; the resultant binary numbers are stored in computer memory as an array of values representing relative image intensities within a subregion. A differencing algorithm applied to the collected data allows the level of objective lens excitation to be found at which the sharpness of the image is optimized, and the excitation may be related to the working distance for that subregion through a previous calibration experiment. The sensitivity of the method for detecting small height changes is theoretically of the order of 1 μm.
A review of on‐line processing of chemical analysis data using a conventional CTEM/STEM
Authors: Nicholls A. W. , Jones I. P. 
Published: August 2011

SUMMARYA review of the on‐line processing of X‐rays and the energetically complementary electron intensities to yield chemical compositions is followed by a discussion of X‐ray mapping, energy filtered images and some extended uses of conventionally supplied equipment associated with these analytic techniques.
Glycol methacrylate embedding of bone and cartilage for light microscopic staining
Authors: Cole Madison B. 
Published: August 2011

SUMMARYA method is described for embedding bone and cartilage in glycol methacrylate (GMA) for light microscopy. Dehydration‐infiltration of the hard tissue is with aqueous GMA solutions minimizing solvent and dehydration artefact, and polymerization is by UV light in the cold to minimize thermal damage. Over fifty stains, enzyme localizations and related histochemical methods for 0·5–3·0 μm thick sections of GMA embedded tissue are listed. The increased resolution plus the localization of cellular and extracellular chemical moieties is now easier and more accurate providing an improved method for the study of the musculo‐skeletal system by light microscopic histochemistry.
An improved double‐tilt stage for the AEI EM7 high‐voltage electron microscope
Authors: Turner J. N. , Ratkowski A. J. 
Published: August 2011

SUMMARYVibration transmitted by the specimen rod of a side‐entry stage frequently decreases image resolution, and the length of the rod in the high‐voltage electron microscope can make the problem severe. A detachable tip clamped to the translation stage minimizes the effect, but eliminates the rod as a means of tilting. Furthermore, the second tilt mechanism is usually built into the rod. Thus the sample is coupled not clamped to the rod, increasing the effect. The image resolution attainable with our double‐tilt stage was 1·5 nm along the rod's axis and 5·0 nm perpendicular to it before modification.An isotropic resolution of 0·5 nm was achieved by attaching the specimen tip to the rod with a flexible coupling and clamping both ends of the tip to the translation ring. The couplings used transmit the torque to rotate the specimen holder but dampen vibration. Ion‐pumping the vacuum system with all other pumps off also improved specimen stability.
Improved handling of embedding plastics for electron microscopy
Authors: Shannon W. A. 
Published: August 2011

SUMMARYAn improved, safer, rapid method for preparing embedding plastics for electron microscopy is described. The method consists of contained storage and dispensing of individual plastic components on an automatic tare balance. The proportions are based on weight measurements and may be calculated from volume or proportion recipes. The usual problems in and resulting from embedding plastic handling have been eliminated.
Automated cell counting in tissue sections: a new approach by ‘multiple grey‐level analysis‘
Authors: Wynford‐Thomas D. , Garrahan N. , Jasani B. , Williams E. D. 
Published: August 2011

SUMMARYA system is described for the automated quantitation in tissue sections of the follicular and stromal cell populations of the thyroid gland, using a computer‐linked TV image analyser. The paper illustrates how the problems of variable staining intensity and clustering of images (nuclei) may be overcome by a novel approach, in which multiple analyses of each field are made at increasing grey‐level thresholds, and then ‘synthesized’ by computer to give a composite image. Discrimination between cell (nuclear) types was by a single minimum width criterion.The results were highly reproducible and correlated well with counts obtained by a comparable manual method.
Freeze‐substitution
Authors: Harvey Diana M. R. 
Published: August 2011

SUMMARYFreeze‐substitution is a technique suitable for the preparation of unicellular and multicellular plant and animal specimens for conventional light microscopy, TEM and SEM. It is also widely used as a means of preparing animal and plant tissues for the localization of water soluble substances by analytical electron microscopy, autoradiography or visual detection of precipitates.The technical requirements of preparation, together with an evaluation of the procedures, are presented for various applications. Careful selection and evaluation of freezing technique, substitution solvent and regime are required for meaningful results.
Applications of secondary ion mass spectrometry (SIMS) in biological research: a review
Authors: Burns Margaret S. 
Published: August 2011

SUMMARYSecondary ion mass spectrometry (SIMS) is a potentially valuable but not fully exploited technique for problems in biological research. It is valuable because of: (1) detection of all elements and isotopes from mass 1, hydrogen; (2) high sensitivity for physiologically important elements, Na, K, Mg, Ca; (3) ion imaging of elements in areas as large as 250 μm in diameter with a lateral resolution of 0·5 μm; (4) promising efforts at quantitation at levels as low as 0·1 mmol/kg; (5) ability to analyse sequential layers to form a three‐dimensional analysis. The problems which complicate its use are primarily variations in ion emission, presence of polyatomic interferences and tissue preparation. Examples are cited of SIMS analysis of biomineralization, botany, toxicology and physiology, mostly by ion imaging techniques. SIMS has not yet been fully exploited for any single biological problem. In particular, studies using isotope discrimination, quantitative analysis and depth profiling will enhance the usefulness of SIMS as a technique for biological research.
A simple device for recording serial optical sections
Authors: Haarlem R. Van , Lagerweij C. , Horn L. C. J. E. M. Ten 
Published: August 2011

SUMMARYA stepper motor was attached by way of a speed reducer to the coarse focusing spindle of a light microscope. After exposure of a film frame the stepper motor displaces the microscope stage according to the programmed instructions laid down in the electronic control unit. In this manner the focal plane is changed and serial optical sections of the developing fish blastoderm were recorded on 16 mm film. This design facilitates the analysis of cell behaviour during early development of annual fishes, of either large numbers of cells simultaneously or of individual cells in detail.
Coalignment of the muscle cell and nucleus, cell geometry and Vv in the tunica media of monkey cerebral arteries, by electron microscopy
Authors: Canham Peter B. , Henderson Ruth M. , Peters Mark W. 
Published: August 2011

SUMMARYWe undertook to ascertain how well aligned is the rod‐shaped nucleus within the spindle‐shaped cell of vascular muscle in order that we might use the darkly staining nucleus in histological sections to indicate precisely the directional alignment of the cell. We fixed cerebral arteries from five monkeys under physiological pressure and embedded portions of the tissue so that mid‐plane longitudinal sections of the arteries were obtained; the circumferentially arranged muscle cells were cut in cross‐section. From the electron micrographs we obtained the cross‐sectional profile of the cell and its nucleus, determining that the centre of the nucleus was on average 9·5 ± 5·8% (SD) away from the centre of the cell (expressed as a ratio of the cellular diameter). We calculated the alignment between the cell and nucleus to be from 0 to 3°, and obtained a volume fraction of 59% for muscle tissue in the tunica media of these arteries.
Water in malignant tissue, measured by cell refractometry and nuclear magnetic resonance
Authors: Ross K. F. A. , Gordon R. E. 
Published: August 2011

SUMMARYIt has long been known that tumour‐bearing tissues often have a significantly higher water content than the normal tissues from which they have been derived. Most of the evidence suggesting this in recent years has been obtained from methods employing nuclear magnetic resonance (NMR), which, although undoubtedly indicative of hydration, cannot at present be precisely quantified. Furthermore it has not been possible by these means to determine whether this overall increase in the water content of the tissue is principally an increase in the extracellular fluid or whether the water content of the tumour cells and the cells immediately adjacent to the tumours increases also. In this investigation, which is the first of its kind, a combination of NMR and immersion refractometry techniques have been used to examine the water content of normal and tumour bearing tissues. NMR measurements were made on pieces of normal and tumour bearing tissue from rat livers: intact living cells were also isolated from these pieces and their refractive indices measured by immersion refractometry from which the water content of their cytoplasm was calculated. It was found that all the cells so measured obtained from hepatomas had more water in their cytoplasm (usually over 5% more water) than any of the cells from normal livers; and that normal‐looking cells taken from the vicinity of hepatomas also all had more water in them than those of normal liver cells, although the differences in this case were less. These results were closely parallel to those obtained by NMR measurements. It is therefore concluded that an appreciable proportion of the increase in the water content of the tissue as a whole that occurs during carcinogenesis, must, in this tissue, be an increase in intracellular water.
Autoradiography: its methodology at the present time
Authors: Williams M. A. 
Published: August 2011

SUMMARYThe autoradiographic methods used in biological and medical research are reviewed from an historical viewpoint. It is pointed out that early methodological research was directed at the selection of suitable detector substances for radiation (nuclear emulsions) and at the preparation and application of suitable layers of these detectors to biological samples. On the other hand, later research was more concentrated on aspects of the preparation of the biological components of the specimens. In particular, the retention of biochemicals in the tissue sample was studied in detail. Research in the quantification of the emulsion response to α and β radiation paved the way for certain quantitative applications of autoradiography. This has been followed by the derivation of models for the measurement of image spread and the evolution of image analysis methods which allow refined determinations of the specific radioactivity of fine structures taking into account losses and gains of silver grains by image spread.The sum of all these advances is a methodology capable of localizing and quantifying radioactivity and hence marking chemical processes from an organism and organ level down almost to the present limits of anatomical delineation.
The distribution of sodium and potassium in toad (Bufo bufo) oocytes estimated by a stereological method
Authors: Ibrahim K. B. , Dick D. A. T. 
Published: August 2011

SUMMARYAs the toad oocyte grows, the intracellular Na concentration rises and the K concentration falls. By stereological analysis using a point counting method in a projection apparatus, it is shown that as the oocyte grows the volume density of the nucleus, endoplasmic reticulum and mitochondria falls, while that of the yolk platelets and cytoplasmic vesicles rises. A multiple regression analysis applied to the sodium and potassium concentrations and to the volume densities of the subcellular components indicates that sodium is most concentrated in cytoplasmic vesicles while potassium is most concentrated in a component composed of the cell cytoplasmic ground substance, the mitochondria and the cell nucleus.
Frequencies of n‐polygons in planar sections of polyhedrons
Authors: Voss Klaus 
Published: August 2011

SUMMARYIn geology and metallography thin or polished sections of crystalline aggregates are a main subject of microscopic investigations. It is important to know what kinds of polygons are generated by planar sections through polyhedrons. In recent years computer simulation was the common method to determine the frequencies of n‐polygons. In this paper an integral geometric approach to derive exact formulas for these frequencies is described. For some families of simple polyhedrons results are given in analytical form and as numerical values.
Labelled‐replica techniques: post‐shadow labelling of intramembrane particles in freeze‐fracture replicas
Authors: Rash John E. , Johnson Timothy J. A. , Hudson C. Sue , Giddings F. Dennis , Graham William F. , Eldefrawi Mohyee E. 
Published: August 2011

SUMMARYThree methods are described for direct post‐fracture, post‐shadow labelling of individual classes of intramembrane particles (IMPs) in freeze‐fracture replicas of biological membranes. The P‐face IMPs corresponding to the acetylcholine receptor complexes (AChRs) of vertebrate neuroeffector junctions are identified by post‐replication labelling with ferritin‐antibody complexes and with neurotoxin‐biotin‐avidin‐colloidal gold affinity ligands. (The freeze‐etch nomenclature of Branton et al., 1975, is used in this report.) These post‐shadow labelling techniques resemble conventional en bloc labelling techniques except that the labelling reagents must penetrate a thin but discontinuous layer of platinum superimposed on the molecules of interest. In the ‘sectioned labelled‐replica technique’, the replicated and labelled tissues are stained, embedded in plastic and sectioned parallel to the replica‐tissue interfaces. In the direct ‘labelled‐replica techniques’, the replicated and labelled samples are freeze‐dried or critical point dried, the labelled surfaces are stabilized by carbon coating, and the underlying tissues are dissolved, allowing the labelled‐replicas to be examined as conventional freeze‐fracture replicas. The unshadowed side of each AChR IMP is shown to retain sufficient biochemical information to permit both immunospecific and neurotoxin specific labelling despite formaldehyde fixation, freezing, fracturing, platinum shadowing, and thawing in aqueous media. A new mixed ferricyanide‐osmium staining method reveals electron opaque structures spanning the membrane bilayer in the same size, number and distribution as the labelled IMPs. These experiments demonstrate the feasibility of identifying individual IMPs in freeze‐fracture replicas and may allow the identification of specific membrane lesions in human disease.