bluuebunny/crossref_metadata_2025_split huggingface dataset:
Digital filters to restore information from fast scanning tunnelling microscopy images
Authors: Aguilar M. , Anguiano E. , Diaspro A. , Pancorbo M. 
Published: August 2011

SUMMARYA transfer function—similar to that used in optical cases to correct blurring effects due to the circular aperture of the system—is presented here to restore scanning tunnelling microscopy (STM) images. Due to the conical geometry of the tip‐sample system, we have established an analogy between the process of image formation in STM and in optical systems. The transfer function utilized, similar to that calculated by Stokseth, allows us to differentiate between the blurring effects introduced along the x and y axes. These effects are different due, mainly, to the different velocities achieved along the x and y directions. Furthermore we have measured the β parameter that characterizes the classical 1/fβ noise present in STM data, demonstrating its independence from experimental conditions. A Wiener filter is utilized to restore the images using the previous assumptions given for the transfer function and noise effects.
The quantification of fluorescent emission from biological samples using analysis of polarization
Authors: Entwistle A. , Noble M. 
Published: August 2011

SUMMARYThe quantification of fluorescent emission from biological specimens can only be carried out in cellular regions where the relationship between fluorophore concentration and fluorescent emission is linear. Using a confocal scanning laser microscope, we show that quantification of fluorescent emission from biological samples labelled with fluorescein and fluorescein analogues mounted in a viscous medium can be readily achieved. Where the distribution of fluorophore is highly localized, for example in cells labelled for immunofluorescence analysis, we demonstrate that analysis of fluorescence depolarization can identify regions in which fluorophore concentration exceeds the range in which the relationship to fluorescent emission is linear. We also demonstrate that, under the conditions examined, depth‐dependent effects, fading and quenching are either small enough to be ignored or can be corrected for mathematically when quantifying fluorescent emission.
Effect of high‐pressure freezing on plant microfilament bundles
Authors: Ding Biao , Turgeon Robert , Parthasarathy M. V. 
Published: August 2011

SUMMARYCell structures, particularly cytoskeletal elements, of tobacco (Nicotiana tabacum L. Var. Maryland Mammoth) leaf tissues were studied after high‐pressure freezing and freeze substitution. In general, the freezing quality was superb and all organelles including microtubules, single microfilaments and microtubule‐associated microfilaments were well preserved. However, when compared with those frozen with a propane jet freezer, the microfilament bundles in high‐pressure frozen cells often had a frayed or loosened appearance. We suggest that such an appearance is an artefact, presumably caused by the high pressure developed prior to freezing.
The new stereological tools in metallography: estimation of pore size and number in aluminium
Authors: Karlsson Lars M. , Cruz‐Orive Luis M. 
Published: August 2011

SUMMARYA number of simple, unbiased and efficient methods are now available in stereology for estimating the number and size of arbitrary particles or voids in a material, with the only assumption that particles must be identifiable on serial sections or confocal planes through the material. In recent years, these methods have been developed and applied mainly in a biomedical context: this paper reviews and illustrates them with the aid of a metallographic example, namely the pore population of a sand‐cast aluminium alloy. Our goal is to convey the fact that stereology is sampling in three dimensions, and therefore its principles remain valid and applicable in no matter what context. The disector, the selector, and an indirect method to estimate the distance between two parallel planes of polish are thereby illustrated. It is also shown how to split the error variance of the estimator of the pore volume fraction (‘porosity’) into the three components due to blocks, sections within blocks and systematic point counting on sections.
The efficient and unbiased estimation of nuclear size variability using the ‘selector’
Authors: McMillan Anna‐Marie , Sørensen Flemming Brandt 
Published: August 2011

SUMMARYThe selector was used to make an unbiased estimation of nuclear size variability in one benign naevocellular skin tumour and one cutaneous malignant melanoma. The results showed that the estimates obtained using the selector were comparable to those obtained using the more time consuming Cavalieri‐disector approach. Employing ‘optical sections’, the selector was found to be between five and ten times more efficient than the Cavalieri‐disector method when using physical sections.
Diamonds are a cryosectioner's best friend
Authors: Michel M. , Gnägi H. , Müller M. 
Published: August 2011

SUMMARYHigh‐pressure frozen Golden Delicious apple leaves were cryosectioned at low temperature with diamond knives. Good cryosections were obtained by optimizing the cutting parameters, i.e. sectioning temperature, mechanical stability of the sample, and sectioning velocity. Cutting artefacts were minimized by reducing the electrostatic interactions between the knife surface and the cryosection. This was accomplished by sectioning the sample in the presence of an ionization electrode. The ionization device, with a primary voltage of 7–8 kV, produces positively and negatively charged nitrogen ions which neutralize the surface charges of the knife and the section. This minimizes the friction on the knife surface and results in ultrathin sections without crevasses or knife marks. Compression of the sections could be minimized, but not avoided, by reducing the knife angle to 30°. Improved contrast of the frozen‐hydrated sections was obtained with the Zeiss EM 902 energy‐filter microscope operated in the zero‐loss mode.
Imaging and measurement of cytosolic free calcium in plant and fungal cells
Authors: Read N. D. , Allan W. T. G. , Knight H. , Knight M. R. , Malhó R. , Russell A. , Shacklock P. S. , Trewavas A. J. 
Published: August 2011

SUMMARYCalcium plays a central role as a second messenger in plant and fungal cells and as such is involved in controlling numerous biological processes. Direct demonstration of signal‐response coupling via Ca2+requires the measurement and localization of changes in cytosolic free Ca2+, [Ca2+]i, during these processes in living cells. In recent years this has become possible with the introduction of a range of fluorescent dyes (e.g. Indo‐1 Fura‐2 and Fluo‐3) which have a high affinity and selectivity for free Ca2+. When used with recently developed microscope technologies (e.g. fluorescence ratio imaging or confocal scanning laser microscopy), subcellular localization and precise quantification of [Ca2+]idynamics in single cells can be achieved. This review describes the principles of [Ca2+]iimaging and measurement using fluorescent dyes, the equipment required to do it, the problems with botanical material and how they are being overcome, future developments for this approach in plant cell biology, and an entirely different strategy for the imaging and measurement of [Ca2+]iinvolving genetic transformation with the aequorin gene.
Confocal laser microscopy and three‐dimensional reconstruction of nucleus‐associated microtubules in the division plane of vacuolated plant cells
Authors: Lloyd Clive W. , Venverloo Catharina J. , Goodbody Kim C. , Shaw Peter J. 
Published: August 2011

SUMMARYThe way in which transvacuolar strands radiating from the cell nucleus reorganize to form the phragmosome, within which division occurs, has been thoroughly studied in epidermal explants of Nautilocalyx lynchii. In recent years it has been established that the movement of the nucleus into the centre of large vacuolated cells such as these, in preparation for division, involves actin filaments. In the present study, the appearance and gradual reorganization of nucleus‐associated microtubules (NAMTs) over the premitotic period is described. Epidermal explants fluorescently labelled with anti‐tubulin were optically sectioned by confocal scanning laser microscopy, the sections reconstructed by an image processing computer and projected as rotating stereo pairs. This revealed that the NAMTs are a major component of the phragmosome, and that they change from a radiating to a planar distribution concomitantly with the ‘bunching’ of cortical MTs to form the pre‐prophase band. The continuity of the two sets of MTs indicates that the band contains newly polymerized microtubules. Other recent studies on the division of vacuolated cells are reviewed and factors affecting the alignment of the division plane are discussed.
Complexity in the spatial localization and length distribution of plant cell‐wall matrix polysaccharides
Authors: McCann Maureen C. , Wells Brian , Roberts Keith 
Published: August 2011

SUMMARYDistinct polysaccharide fractions can be obtained from onion cell walls by sequential extraction methods. Pectins and xyloglucan molecules from these fractions have been imaged by electron microscopy and their lengths measured. Unexpectedly, instead of a simple distribution of lengths, both classes of matrix polysaccharide showed clear evidence of length periodicity, suggesting block polymer construction using subunits about 30 nm long, each containing a minimum of about sixty backbone sugar residues. These results are discussed in the light of our observation that the average hemicellulose cross‐link between cellulose microfibrils in the wall is also about 30 nm long, and that pectin forms a co‐extensive network with the cellulose/xyloglucan network. Using monoclonal antibodies to pectin we have further shown that even within a thin primary cell wall, containing only a few lamellae, distinct zones of pectin distribution are detectable, thus reinforcing our notion of the importance of domains in wall architecture. The chemical heterogeneity detected amongst pectins indicates that in a primary cell wall each lamella of cellulose microfibrils may be in a unique matrix environment.
Self‐incompatibility: insights through microscopy
Authors: McFadden G. I. , Anderson M. A. , Bönig I. , Gray J. E. , Clarke A. E. 
Published: August 2011

SUMMARYThe contributions of microscopy to our understanding of gametophytic self‐incompatibility in the ornamental tobacco Nicotiana alata are reviewed. Self‐incompatibility is a genetically based system which prevents self‐fertilization and thereby promotes outbreeding in plants. Pollen bearing an S‐allele identical to either of the S‐alleles in the female sporophytic tissues is rejected. Several S‐allele‐specific style glycoproteins have been isolated from Solanaceous species. All have ribonuclease activity and are termed S‐RNases. The cDNA clones encoding N. alata S‐RNases have been used in in situ hybridization experiments which show that the genes are expressed in the transmitting tract of the style and in the epidermis of the placenta. Immunocyto‐chemistry using an antibody to S2‐RNase demonstrates extracellular accumulation of the enzyme in the same tissues. These tissues are the pathway through which pollen tubes must grow to reach the ovules. The microscopy data are congruent with a model in which self‐pollenation is susceptible to an allele‐specific cytotoxin produced by the sporophyte.
Ultrastructural localization of titanium in chemically modified chrysotile asbestos
Authors: Berghmans Patrick , Jacob Wim , Adams Freddy 
Published: August 2011

SUMMARYThe ultrastructural localization of Ti in chemically modified chrysotile fibres has been studied. TiCl3‐treated fibres were analysed by electron energy‐loss spectroscopy and electron spectroscopic imaging (ESI) along directions parallel and perpendicular to the fibre axis. ESI of ultrathin sections of conventionally embedded chrysotile fibres was made possible with the use of a cryo‐stage. The ESI images observed in fibre cross‐sections revealed that the Ti is located not only at the surface of the fibres but also inside the fibril tubes. This combination of microanalysis and imaging in the analytical electron microscope is the most suitable technique for relating the microchemistry and microstructure on a submicrometre scale.
The application of electron spectroscopic imaging for quantification of the area fractions of calcium‐containing precipitates in nervous tissue
Authors: Körtje Karl‐Heinz , Körtje Doris 
Published: August 2011

SUMMARYEnergy‐filtering transmission electron microscopy has been applied to the quantification of area fractions of calcium‐containing cytochemical reaction products in central nervous tissue and the retina of fish. The method of electron spectroscopic imaging using electrons with an energy loss of 250 eV produces images with a very high, structure‐sensitive contrast. This is a suitable imaging condition for the reliable detection of reaction products and structural details in unstained ultrathin sections. The images were recorded with a sensitive TV camera and evaluated with the integrated digital image‐analysis system of the Zeiss CEM 902 energy‐filtering electron microscope. An empirical procedure was developed which objectively detects reaction products and calculates characteristic values, taking into account different staining intensities. This new and sensitive method enabled an assessment to be made of the influence of temperature and light adaptation on cytochemically detectable calcium in nervous tissue of fish. Higher amounts of calcium‐containing reaction product were detected in synaptic clefts of the optic tectum in warm‐adapted fish than in cold‐adapted fish. In synaptic vesicles of photoreceptor cells in the fish retina, higher amounts of reaction product were found in dark‐adapted fish than in light‐adapted fish.
A variable cell culture chamber for ‘open’ and ‘closed’ cultivation, perfusion and high microscopic resolution of living cells
Authors: Pentz S. , Hörler H. 
Published: August 2011

SUMMARYA multipurpose chamber is described for growing and testing cultured cells. The chamber can be converted from a perfusion chamber to an ‘Open’ or ‘closed’ culture system. The chamber provides optimum conditions for microscopy using all common objectives and condensers for different microscopic methods.
Fixation for biological ultrastructure. I. A viscometric analysis of the interaction between glutaraldehyde and bovine serum albumin
Authors: Luft John H. 
Published: August 2011

SUMMARYThe increase in viscosity resulting from mixing concentrated solutions of albumin with dilute glutaraldehyde has been investigated. The viscosity changes are slow at first, very sensitive to small changes in glutaraldehyde concentration, and non‐linear with time. The cross‐linking of the albumin by glutaraldehyde seems to be retarded by the weak mechanical shear forces produced by the viscometer. The significance of these findings to the events in tissue fixation is discussed.
Improved preservation of bacterial exopolymers for scanning electron microscopy
Authors: Vandevivere P. , Baveye P. 
Published: August 2011

SUMMARYTwo methods of sample preparation, originally developed for transmission electron microscopy and better at preserving bacterial exopolymers than the traditional fixation with glutaraldehyde, were adapted for scanning electron microscopy. The first involves a glutaraldehyde‐lysine mixture as a fixative. The second, based on the use of polycationic ferritin, was modified to include a post‐fixation step with either glutaraldehyde or a glutaraldehyde‐lysine mixture. These techniques were found to improve considerably the preservation of exopolymers secreted by three bacterial strains.
Texture analysis of ceramic films by image processing
Authors: Prod‘Homme Murielle , Chermant Liliane , Coster Michel 
Published: August 2011

SUMMARYThis paper presents an application of automatic image analysis to the morphometry of ceramic sheets. The sheets can be characterized by granulometrie and stereological texture parameters of the granular and the porous phases. Grey level granulometry, automatic thresholding and filtering have thus been used. The results show that the morphology of green sheets is strongly dependent on the ball‐milling conditions.
Estimation error of Leydig cell numbers in atrophied rat testes due to the assumption of spherical nuclei
Authors: Mendis‐Handagama S. M. L. C. 
Published: August 2011

SUMMARYIn this study, Leydig cell numbers in control and atrophied testes (induced via subcutaneous implants of testosterone plus 17β estradiol for 16 weeks; TE‐implanted) of rats, estimated via the fractionator method (independent of any assumptions) were compared to those estimated via the disector (unbiased, but dependent on shrinkage) and Floderus (assumes spherical particles, dependent on shrinkage) methods. Estimates of Leydig cell numbers in control rats produced by all three stereological methods were similar. In rats with atrophied testes, both the fractionator and the disector methods produced significantly lower (P < 0.01; 47% and 41 % with fractionator and disector, respectively) Leydig cell number estimates per testis than in the controls. By contrast, the estimates of Leydig cell number in atrophied testes derived via the Floderus equation were not significantly different from those of controls, but larger than those obtained via the fractionator and the disector methods. These results suggested that the assumptions of the Floderus method were violated in the atrophied rat testes.Why was the Floderus method of estimating Leydig cell number applicable to control rats but not to the TE‐implanted rats? In an attempt to answer this question the diameter measurement together with its correction factor used in the Floderus equation (i.e. D + t − 2H) was also derived from the data collected for the disector method. The values for D + t − 2H used in the Floderus method and also calculated via the disector method were found to be identical in controls, but for the TE‐implanted rats a 32% lower value was obtained with the Floderus equation when compared to the disector. These findings suggested that this estimation error caused an overestimation of Leydig cell numbers in the TE‐implanted rat testes.
Pattern reconstruction in ultrastructural morphology
Authors: Bellon Pier Luigi , Lanzavecchia Salvatore 
Published: August 2011

SUMMARYA method of image reconstruction which can be used in ultrastructural morphology when a number of comparable structures are available is described. The method is suitable for quasi‐regular arrays of objects, and an extension of it allows the average of equivalent objects which could be orientated at random to be obtained. When dealing with crystalline arrays it is first necessary to obtain a leakage‐free pattern in which the coarse distortions are suppressed; this pattern is then filtered to obtain a reference. When dealing with several individual objects which must be brought to the same orientation and enlargement, one of the objects is fixed as a reference. In both cases the distortions relative to the reference are eliminated by an iterative process, making a final average possible. In crystalline arrays this method enhances the bandwidth of the pattern in a way that can be estimated quantitatively by figures of merit. Two examples of reconstructions are reported and discussed: a ctenophore macrocilium, regarded as a crystalline structure, and a sectioned flagellum of an insect spermatid.
Combined cathodoluminescence and X‐ray analysis of paint pigments
Authors: Koschek Georg 
Published: August 2011

SUMMARYThe suitability of cathodoluminescence (CL) measurements for differentiating between different paint pigments is demonstrated with an example of a combined energy‐dispersive X‐ray (EDX) and CL analysis of different zinc oxide (ZnO) pigments in a paint fragment where the EDX spectra are virtually identical, but where the CL spectra show significant differences. Consequently, it is possible to distinguish different pigments on the basis of CL spectra and monochromatic CL micrographs.
An environmental sample chamber for X‐ray microscopy
Authors: Goncz K. K. , Batson P. , Ciarlo D. , Loo B. W. , Rothman S. S. 
Published: August 2011

SUMMARYThe manufacture, construction and performance of a special specimen holder for biological samples suspended in aqueous solution is described. The holder was designed for use in a scanning transmission X‐ray microscope. Using it, we have successfully obtained high‐resolution images of fresh cellular and subcellular specimens in suspension and have been able to vary the sample environment during viewing in the microscope.
A Monte Carlo code to simulate the effect of fast secondary electrons on κAB factors and spatial resolution in the TEM
Authors: Gauvin Raynald , L'Espérance Gilles 
Published: August 2011

SUMMARYFast secondary electrons (FSE), which result from inelastic scattering of incident electrons, are known to generate a significant number of X‐rays for light elements, and also to degrade the spatial resolution of X‐ray microanalysis in thin foils. A Monte Carlo program simulating the generation and diffusion of FSE in binary systems has been developed to study the effect of composition on κAB factors and spatial resolution. The effect of accelerating voltage and thickness is also presented.
High‐resolution scanning electron microscopy of frozen‐hydrated cells
Authors: Walther Paul , Chen Ya , Pech Louis L. , Pawley James B. 
Published: August 2011

SUMMARYCryo‐fixed yeast Paramecia and sea urchin embryos were investigated with an in‐lens type field‐emission SEM using a cold stage. The goal was to further develop and investigate the processing of frozen samples for the low‐temperature scanning electron microscope (LTSEM).Uncoated frozen‐hydrated samples were imaged with the low‐voltage backscattered electron signal (BSE). Resolution and contrast were sufficient to visualize cross‐fractured membranes, nuclear pores and small vesicles in the cytoplasm. It is assumed that the resolution of this approach is limited by the extraction depth of the BSE which depends upon the accelerating voltage of the primary beam (V0). In this study, the lowest possible (V0) was 2.6 kV because below this value the sensitivity of the BSE detector is insufficient. It is concluded that the resolution of the uncoated specimen could be improved if equipment were available for high‐resolution BSE imaging at 0.5–2kV.Higher resolution was obtained with platinum cryo‐coated samples, on which intramembranous particles were easily imaged. These images even show the ring‐like appearance of the hexagonally arranged intramembranous particles known from high‐resolution replica studies. On fully hydrated samples at high magnification, the observation time for a particular area is limited by mass loss caused by electron irradiation. Other potential sources of artefacts are the deposition of water vapour contamination and shrinkage caused by the sublimation of ice.Imaging of partially dehydrated (partially freeze‐dried) samples, e.g. high‐pressure frozen Paramecium and sea urchin embryos, will probably become the main application in cell biology. In spite of possible shrinkage problems, this approach has a number of advantages compared with any other electron microscopy preparation method: no chemical fixation is necessary, eliminating this source of artefacts; due to partial removal of the water additional structures in the cytoplasm can be investigated; and finally, the mass loss due to electron beam irradiation is greatly reduced compared to fully frozen‐hydrated specimens.
A simple way to encapsulate small samples for processing for TEM
Authors: Mariné M. C. Hernández 
Published: August 2011

SUMMARYA variation of the technique of encapsulating samples in agar cubes is presented. The samples are enclosed by bubbles in the agar. The hollow space is occupied by the specimen and a drop of melted agar is used to fill the remainder of the cavity and to cover it. The solidified agar is cut into a small cube containing the bubble, and is processed for TEM in the usual way.