bluuebunny/crossref_metadata_2025_split huggingface dataset:
Low‐temperature SEM for detection of fungicide activity
Authors: Guggenheim R. , Düggelin M. , Mathys D. , Grabski C. 
Published: August 2011

SUMMARYLow‐temperature scanning electron microscopy (LTSEM) combined with cryopreparation methods provided images of well‐preserved biological surfaces and structures on a routine basis. Fractures of wheat leaves revealed epidermal and parenchymatous cells and masses of fungal hyphae growing in intercellular spaces. Freeze‐fractured plant cells contained haustoria of the brown rust fungus Puccinia triticina. Extrahaustorial matrices were clearly distinguishable and at infection sites granular material was found. Activity of the triazole fungicide cyproconazole was mainly directed towards fungal hyphae and sporogenic tissue, resulting in a stronger branching and swelling of hyphal tips and collapse of fungal cells.Cryofixation methods combined with the use of a cryopreparation unit (Balzers SCU 020) were more reliable in interpreting the observed biological events through easier discrimination between evidence and artefacts. More advantages in the use of the cryopreparation unit are described in detail.
Contrast in high‐resolution scanning electron microscope images
Authors: Joy David C. 
Published: August 2011

SUMMARYCurrent scanning electron microscopes, equipped with field emission guns and high‐performance immersion lenses, can achieve spatial resolutions of the order of 1 nm in both secondary and backscattered imaging modes over a wide range of operating energies. The generation and interpretation of images with nanometre‐scale resolution relies on a detailed knowledge, and application, of electron‐solid interactions. This paper develops the practical steps required to produce a high‐resolution image, and discusses the principles which govern image interpretation. Attention is focused primarily on materials which are low in atomic number and density, such as biological tissue, but the results apply after appropriate scaling of the physical parameters to most other materials.
High‐resolution scanning electron microscopy of frozen‐hydrated and freeze‐substituted kidney tissue
Authors: Herter Peter , Tresp Gregor , Hentschel Hartmut , Zierold Karl , Walther Paul 
Published: August 2011

SUMMARYInner surfaces and fracture faces of rabbit kidney tissue were investigated with high‐resolution scanning electron microscopy using two different cryopreparation techniques: (i) for the observation of fracture faces, cryofixed tissue was fractured and coated in a cryopreparation chamber dedicated to SEM, vacuum transferred onto a cold stage and observed in the frozen‐hydrated state; (ii) for the observation of inner surfaces of the nephron, water was removed after freezing and fracturing by freeze substitution and critical‐point drying of the tissue.By both methods, macromolecular structures such as intramembranous particles on fracture faces and particles on inner surfaces were imaged. The latter method was used to investigate in more detail surface structures of cells in the cortical collecting duct. These studies revealed a heterogeneity of intercalated cells not described thus far.
Modelling and analysis of 3‐D arrangements of particles by point processes with examples of application to biological data obtained by confocal scanning light microscopy
Authors: König D. , Carvajal‐Gonzalez S. , Downs A. M. , Vassy J. , Rigaut P. 
Published: August 2011

SUMMARYWithin the concept of point processes, a review is presented of quantities which can be used in studies of three‐dimensional (3‐D) aggregates of particles. Suitable characteristics and estimators are given for both unmarked and marked point processes. To demonstrate the feasibility of such quantitative approaches, an application in histology, dealing with 3‐D arrangements of cell nuclei in rat liver, is described. Using a confocal scanning light microscope, 3‐D images are recorded and image analysis used to obtain the coordinates of the centroid, together with the volume and DNA content, of each cell nucleus. Examples of results are given, using both unmarked and marked point processes. In the latter case, cell type, nuclear volume and ploidy group are suitable marks.
Ultra‐rapid freezing by spraying/plunging: pre‐cooling in the cold gaseous layer
Authors: Chang Zhaohua , Baust John G. 
Published: August 2011

SUMMARYA thermophysical model is established to analyse the influence of pre‐cooling of a biological specimen in the cold gas layer associated with spray‐freezing techniques. The basic principles governing the process of pre‐cooling are provided. It is concluded that pre‐cooling is one of the major limiting steps in attaining an overall ultra‐rapid cooling rate. Pre‐cooling has a substantial influence on the nature of the final frozen specimens. In order completely to avoid crystallization before entry into the liquid cryogen and maximize the overall cooling rate of the specimen, precautions should be taken to control the height of the gaseous layer and the size of the specimen. The probability of the specimen being frozen in the cold gaseous layer is reduced by increasing the entry speed. The effectiveness, however, becomes less marked at speeds greater than 10 m/s. In order to minimize the risk of misinterpreting the measured cooling rate, it is necessary to specify the pre‐cooling conditions. The pre‐cooling effect is much more evident in liquid helium than in cryogens such as propane, ethane, Freon 12, 13 and 22.
Surfaces of cryosections: is cryosectioning ‘cutting’ or ‘fracturing’?
Authors: Kirk R. Gary , Knoff Laura , Lee Ping 
Published: August 2011

SUMMARYIn order to determine if cryosectioning involves ‘fracturing’ or ‘cutting’ we examined the surfaces obtained in cryosectioning by a metal‐replicating procedure commonly used in freeze‐fracture microscopy. Platinum‐carbon replicas were made of the surfaces of both the sections and the complementary surfaces of the sample stubs from which the sections were cut. When samples of frozen red cells were sectioned at −120°C with large knife advancements (1 μm), the chips produced did not resemble sections. Membrane fracture faces, produced by splitting of the lipid bilayer, were found in electron micrographs of replicas of the sample stubs. This demonstrates that a cryomicrotome can be used to produce large intact replicas. When dull knives were used with small knife advancements, both smooth and fractured regions were found. The sections produced with dull knives had a snowflake appearance in the light microscope. When sharp knives were used with small advancements (0·1 μm), replicas of the surfaces were free of fracture faces and the sections had a cellophane‐like appearance in the light microscope. Therefore, in cryosectioning a different process other than ‘fracturing’ is responsible. This ‘cutting’ process may be micromelting of a superficial layer by the mechanism of melting‐point depression from the pressure exerted by the sharp edge of the knife.
Radiation damage in soft X‐ray microscopy of live mammalian cells
Authors: Shinohara Kunio , Ito Atsushi 
Published: August 2011

SUMMARYWhen specimens are observed by soft X‐ray microscopy, they always absorb many photons, causing radiation damage at the imaged site. The problems of radiation damage were studied in view of the principle of image formation; absorption contrast, scattering (holography), or phase contrast. In all cases, photons with a wavelength of 1–10 nm interact with the specimen mainly through the photoelectric effect followed by the transfer of energy to the imaging site either directly (absorption imaging) or indirectly (holography or phase contrast). This absorbed energy will cause structural changes to the imaging site. From a review of the literature the absorbed dose is estimated to be as high as 107 Gy when the expected resolution of the specimen (1–10 thick) is 10 nm. This dose is far in excess of the amount required for cells to be able to survive when live mammalian cells are exposed. The levels of radiation effects were extrapolated to the estimated absorbed dose from the reported values for cell survival, chromosome aberrations, and DNA strand breaks with respect to observations on mammalian chromosomes. The extrapolated results show that some damage will occur in every 10 times 10‐nm (expected resolution) size unit. Although these studies focused only on the effects on mammalian chromosomes, the present results are more or less common phenomena in the observation of biological specimens. Hence, the results suggest that dynamic observations will be difficult. On the other hand, a time‐scale study of the effects of radiation on structural integrity suggests that single‐shot imaging with short‐pulsed (probably shorter than a few milliseconds) X‐rays may be appropriate for the observation of intact live biological specimens in the hydrated condition, before they have deteriorated.
The use of adhesive‐coated grids for the X‐ray microanalysis of dry‐cut sections in the TEM
Authors: Fritz Eberhard 
Published: August 2011

SUMMARYSections cut dry for the X‐ray microanalysis of diffusible elements were fixed to adhesive‐coated single fine‐bar grids. The drawbacks of folding grids normally used for this purpose can thus largely be avoided.
Quantitative analysis of electron energy‐loss spectra from ultrathin‐sectioned biological material
Authors: Sorber C. W. J. , Ketelaars G. A. M. , Gelsema E. S. , Jongkind J. F. , De Bruijn W. C. 
Published: August 2011

SUMMARYElectron energy‐loss spectroscopy (EELS) has been used to determine elemental concentrations in biological specimens, consisting of ultrathin‐sectioned cells and tissues. Chelex100‐based Ca‐ and Fe Bio‐standards are used for elemental quantification to establish iron and calcium concentrations. These Bio‐standards, as well as the biological materials, are treated in a standard EM procedure such that ‘known’ and ‘unknown’ sites are located in one cross‐section.Uncertainties and variabilities present in the equations for calculating the concentration in the ‘unknown’ site (determined by comparing simplex‐fitted EEL spectra from Bio‐standards with those from tissue) are outlined in two examples. Using an H+ Bio‐standard, the matrix composition of such biological cell material is analysed, leading to values which approach each other closely. Quantitative EELS, using Chelex100‐based Bio‐standards, is advocated.
A simplified preparation procedure of plant material for elemental analysis by ESI and EELS techniques
Authors: Lehmann H. , Kunz U. , Jacob A. 
Published: August 2011

SUMMARYLiving plant cells were embedded in the water‐soluble resin Nanoplast FB 101 and cut into ultrathin sections. Non‐vacuolated cells of algae and fungi showed a satisfactory preservation of fine structure. First results of elemental mapping by the ESI technique indicate that phosphorus and sulphur are neither extracted nor dislocated within the cells.The authors are members of the ‘Study Group for Electron Microscopy’ at the Hannover School of Veterinary Medicine.
The glycosomes of trypanosomes: number and distribution as revealed by electron spectroscopic imaging and 3‐D reconstruction
Authors: Tetley L. , Vickerman K. 
Published: August 2011

SUMMARYComputer‐aided 3‐D reconstruction of typanosomes from 0·35‐μm‐thick sections imaged on the Zeiss 902 electron microscope are being used to study the dynamics of cell organization. Segregation of glycolytic enzymes into glycosomes raises questions concerning the distribution and biogenesis of these organelles. Direct counts of glycosomes from Trypanosoma evansi indicate 30–40 per cell and for the closely related T. brucei, 65 per cell. These figures contrast with the estimates of others who have used model‐based morphometric methods to obtain a value of 230 per cell.
Ultrastructural aluminium detection in amphibian tissues by electron spectroscopic imaging and electron energy‐loss spectroscopy
Authors: Böhmer J. , Rahmann H. 
Published: August 2011

SUMMARYAluminium causes a variety of toxic effects in living organisms but very little is known about its uptake, pathways and locations of deposition. We have applied electron spectroscopic imaging (ESI) and electron energy‐loss spectroscopy (EELS) to locate aluminium at the ultrastructural level in amphibian larvae from acidic ponds. It is found diffusely bound or precipitated in cell organelles. The spatial resolution of aluminium detection is high. The elemental composition of small areas can be demonstrated by EELS. Three different fixation procedures give similar results. The two‐ and three‐window methods at the K‐ and L‐edges are compared.
Demonstration of calcium in dermal melanocytes of Xenopus laevis and Poecilia reticulata with electron energy‐loss spectroscopy and electron spectroscopic imaging
Authors: Müller Thomas , Bereiter‐Hahn Jürgen 
Published: August 2011

SUMMARYThe subcellular distribution of calcium in dermal melanocytes of Xenopus laevis and Poecilia reticulata has been analysed. Using two cytochemical methods, phosphate precipitation and a combined oxalate‐pyroantimonate technique, electron energy‐loss spectroscopy and electron spectroscopic imaging have been applied for elemental analysis. Both precipitation techniques revealed a high calcium content in the melanosomes of both species. Calcium was also located in the vicinity of collagen fibrils and in the plasma membrane.
Electron spectroscopic diffraction and imaging of the early and mature stages of calcium phosphate formation in the epiphyseal growth plate
Authors: Barckhaus R. H. , Höhling H. J. , Fromm I. , Hirsch P. , Reimer L. 
Published: August 2011

SUMMARYA review of different models of biomineralization in collagen‐rich hard tissues shows that further investigations of crystal formation are necessary. The electron spectroscopic diffraction (ESD) mode of operation of an energy‐filtering electron microscope offers the possibility of being able to avoid the background from inelastic scattering in selected‐area electron diffraction patterns. First experiments on the different stages of mineralization in the epiphyseal growth plate have only indicated the presence of apatite. The ESD mode can be complemented by the electron spectroscopic imaging mode and by elemental mapping of calcium.
A quantitative computer simulation of microscopic imaging
Authors: Goldstein D. J. 
Published: August 2011

SUMMARYA versatile program is described for a BBC‐ or IBM‐compatible microcomputer, which uses a fast Fourier transform in calculating diffraction‐limited microscopic images. The defined ‘object’ can be a diffraction grating or a discrete specimen seen in cross‐section, of arbitrary interval (or width), transmittance and retardation. Many microscopical techniques can be simulated including bright‐field, phase contrast, central and peripheral dark‐field, schlieren, apodization, fluorescence, differential interference contrast and confocal scanning. Instrumental settings which can be varied include the aperture, focus and primary spherical aberration of the objective, the coherence and obliquity of the illumination, and in phase contrast the width, transmittance and retardation of the phase plate. The Fourier transform and various other algorithms used in the program are explained.
Study of potassium deficiency in cardiac muscle: quantitative X‐ray microanalysis and cryotechniques
Authors: Pogorelov A. , Allachverdov B. , Burovina I. , Mazay G. , Pogorelova V. 
Published: August 2011

SUMMARYAn imbalance of potassium in cardiac muscle causes an alteration of heart function. The distribution and concentration of potassium in rat papillary heart muscle was studied using cryofixation and X‐ray microanalysis. Freeze‐dried cryosections and sections of freeze‐dried, embedded tissue were analysed. Bulk frozen specimens were freeze‐dried either in a vacuum or by a new technique using liquid propane as a cryodehydration medium. These two methods of freeze‐drying were tested for elemental retention in other specimens, with comparable results. A potassium concentration of 120 mmol/l was measured in normal myocytes of cardiac papillary muscle compared to 80 mmol/l in myocytes of animals stressed by a temperature of 45°C for 1 h. The presumed physiological significance of the findings is discussed.
An improved analysis of experimental data from 125 I hot‐line autoradiographs: allowing for the effects of background grains
Authors: Aykroyd R. G. , Anderson C. W. , Junger E. , Williams M. A. 
Published: August 2011

SUMMARYIn this paper we suggest a new statistical method of correcting the results of hot‐line experiments for the effects of background sources and we use the new method to reassess the adequacy of three probability distributions proposed in the literature for image spread from line sources. The data are from sources labelled with 125I in semi‐thin resin sections 0·4‐0·8 μm in thickness. The new method reveals that two of the models describe the empirical distributions more closely than earlier analysis had suggested, and it confirms an increasing relationship between half distance of image spread and the thickness of the source. However, it also confirms that considerable ‘inter hot‐line’ experimental variation remains, even after background correction. This suggests that multiple experiments are needed to produce reliable estimates of half distance.
Structural characterization of air‐oxidized chromium particles by extended energy‐loss fine‐structure spectroscopy
Authors: Diociaiuti Marco , Paoletti Luigi 
Published: August 2011

SUMMARYExtended energy‐loss fine‐structure spectroscopy (EXELFS) and electron energy‐loss spectroscopy (EELS) microanalysis have been used to study the structural and chemical modifications caused by interaction with the atmosphere of small, very reactive chromium (Cr) particles whose diameters range from 150 to 1300 Å.Analysis of the data allows us to deduce that the particles consist of a Cr core with a body‐centred cubic structure and with a lattice parameter reduced by about 4% with respect to the bulk value by surface stresses (for the smallest diameter), surrounded by a Cr2O3 shell of about 15‐Å width.
Comparison between the conventional method and the simple desiccation method in porcine sperm processing for scanning electron microscopy
Authors: Bonet S. , Briz M. 
Published: August 2011

SUMMARYSperm quality has been studied using different microscopic methods. Light microscopy gives similar percentages of sperm typologies (mature, immature and aberrant) as scanning electron microscope (SEM) techniques using both normal (critical‐point drying) and simple desiccation techniques. Artefactual forms, which are easily recognizable, are not taken into account. The artefactual forms observed by the simple desiccation SEM method were spermatozoa with tails folded in a simple loop, and spermatozoa with short tails coiled at their extreme. The simple desiccation SEM method is a useful and rapid technique for the testing of porcine sperm quality.
Alkaline hydrolysis/methylation‐acetylation: a new technique for ultrastructural DNA cytochemistry
Authors: Romero J. B. , Ferrer J. M. , Tato A. , Tandler C. J. , Llorente A. R. , Del Castillo P. , Stockert J. C. 
Published: August 2011

SUMMARYA new technique for the visualization of DNA‐containing structures in electron microscopy is described. Samples of glutaraldehyde‐fixed bone marrow from rats were subjected to alkaline hydrolysis to remove RNA and the phosphate of phospho‐proteins, followed by a combined blockage of protein carboxyl and amino groups through methylation‐acetylation. After uranyl acetate staining of epoxy‐embedded ultrathin sections, chromatin from all cell types showed a highly selective and intense electron opacity. Staining methods for DNA were also positive in semithin sections. This simple procedure could be very useful in ultrastructural cytochemistry of DNA and chromatin.
Electron probe X‐ray microanalysis of intracellular element concentrations in cryosections in the presence of changes in cell volume
Authors: Bostrom T. E. , Field M. J. , Györy A. Z. , Dyne M. , Cockayne D. J. H. 
Published: August 2011

SUMMARYThe interpretation of element concentration data for X‐ray microanalyses of biological tissues, which are subjected to some experimental treatment, can be complicated by changes in cell volume and total cell dry matter induced by the treatment. We have examined the manner in which such changes would affect the values measured in frozen‐dried cryosections of soft tissues, and how they may be taken into account in the interpretation of the results. The element content (mass per unit dry weight) measured by the peak‐to‐continuum or Hall method is independent of changes in cell volume, but is sensitive to a change in the local dry mass. Conversely, intracellular concentrations in terms of mass per unit volume, as determined by the peripheral or internal standard technique, are dependent on volume changes but independent of dry mass. The estimated dry weight fraction is affected by changes in both volume and dry mass. The results obtained from both quantification methods can therefore provide information on the combination of changes in cellular element levels, volume and total dry mass that may occur following the experimental treatment.In a study of the late effect of the drug cisplatin on electrolyte concentrations in kidney proximal tubules, both quantification methods have been used to obtain wet weight and dry weight concentrations. By applying the above considerations, the analytical results have been interpreted as a combination of changes in element levels and a shrinkage of the tubule cells. Cell shrinkage was confirmed by morphometric analysis of tubular cross‐sections.
A device for freeze‐substituting a large number of samples under controlled conditions
Authors: Marshall A. T. , Kent M. 
Published: August 2011

SUMMARYAn inexpensive device for reproducible freeze‐substitution is described. The equipment consists of a reciprocating shaker, holding vials of substitution fluid, mounted in the gas phase of a large liquid nitrogen refrigerator. The shaker temperature is controlled by a programable microprocessor.
Advances in ALCHEMI analysis
Authors: Turner P. S. , White T. J. , O'Connor A. J. , Rossouw C. J. 
Published: August 2011

SUMMARYThe ALCHEMI method for locating the sites of foreign atoms within crystals is known to be sensitive to the delocalized emission of X‐rays. This can result in large errors in some cases through differences in delocalization for different excitations or by error amplification in the ratio method of analysis. An alternative approach to the analysis of ALCHEMI data, using multivariate statistical analysis, is extended to the case of multiple impurities. Initial results from zone‐axis channelling experiments for a Yb‐doped zirconolite (CaZrTi2O7) are shown to confirm the improved accuracy of this method, especially for axial orientations. Data were collected using a 400‐keV analytical electron microscope fitted with an intrinsic Ge X‐ray detector. The potential advantages for ALCHEMI analysis of Ge detectors are considered.
Electron microscopy in synthetic polymer membrane research
Authors: Kim K. J. , Dickson M. R. , Fane A. G. , Fell C. J. D. 
Published: August 2011

SUMMARYThe use of electron microscopy (EM) to study microporous membranes is described. Application of EM to polymer membranes can be classified into two major areas: elucidation of membrane formation mechanisms (the effect of preparative parameters on morphology of the final membrane) and physical characterization of membranes.Conventional transmission and scanning electron microscopes have been used successfully to clarify details of membrane organization (structure) and formation mechanisms as well as supplying some information on the pores. However, neither transmission electron microscopy nor scanning electron microscopy could resolve the fine surface morphology of ultrafiltration membranes due to instrumentation limitations. The recent development of field‐emission scanning electron microscopy has opened new opportunities in membrane research.