bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparative account of the isolated langerhans islets of the rat, pig and human pancreas: light- and Electron Microscopic studies
Authors: Syed Ali S , Syed Ali M.M. , Hering B.J. , Bretzel R.G , Federlin K.F.
Published: June 2020

The process of transplantation is accompanied by many complicated factors, each of which may be the cause of unsuccessful or failure in organ transplantation. Recently, we described the ultrastructural features of isolated is lets from the bovine pancreas. The present study deals with the comparative aspects of the isolated is lets from rat, pig and human pancreata. Rat pancreas was collected in the laboratory; pigmaterial was obtained from the local abattoir, and human pancreata were taken from biopsy material during surgical operation. The collected material was processed according to a recently described method in combination with the digestion-filtration technique, then further prepared for light- and electron microscopical investigations by fixation in 2% glutaraldehyde in 0.135 M phosphate buffer, 7.2 pH. After osmification (1% OsO4 in phosphate buffer) they were washed and dehydrated in acetone and embedded in Durcupan AMC. Semithin sections (0.5μm to 1.0μm) were used for light microscopical studies and ultrathin sections for electron microscopical observations.
The study of fine structure of wood of dove tree (davidia involucrata baill.) by SEM
Authors: Fangim Dai , Saoxin Zong , Dalin Dai , Jie Pan , Changhen Liang
Published: June 2020

The dove tree is a beautiful, ornamental tree which is endemic to western China mainly, both in Sichuan and Guizhou Provunces. The fine structure of its wood has not been studied under SEM, even under LM. Since this tree is a rare species and is suffering from the danger of extinguishment on the earth, which has been protected by RAW, we trace it into consideration and try to provide some information for further exploiting and utilizing it. The method used in this study and the results obtained are stated as follows.

The wood sample was furnished by Provincial Institute of Forestry Science, Sichuan, China. It was collected from Mongjing, Guizhou. The sample is cut with a single edge knife into a small cubic block which is mounted on metal stubs using double-sided tape, coated with a mixture of gold-palladium in a JFC-1100 sputter, and observed at 25 W, H-600 I analysis electron microscope with a H-6010A attachment.
Qualitative study on “hydropic degeneration” of The hepatocytes in viral hepatitis
Authors: Meizhao Le , Ming Ye , Xiaoming Song , Jiazhang Xu
Published: June 2020

“Hydropic degeneration” of the hepatocytes are often found in biopsy of the liver of some kinds of viral hepatitis. Light microscopic observation, compareted with the normal hepatocytes, they are enlarged, sometimes to a marked degree when the term “balloning” degeneration is used. Their cytoplasm rarefied, and show some clearness in the peripheral cytoplasm, so, it causes a hydropic appearance, the cytoplasm around the nuclei is granulated. Up to the present, many studies belive that main ultrastructural chenges of hydropic degeneration of the hepatocytes are results of the RER cristae dilatation with degranulation and disappearance of glycogen granules.

The specimens of this study are fixed with the mixed fluid of the osmium acidpotassium of ferricyanide, Epon-812 embed. We have observed 21 cases of biopsy specimens with chronic severe hepatitis and severe chronic active hepatitis, and found that the clear fields in the cytoplasm actually are a accumulating place of massive glycogen. The granules around the nuclei are converging mitochondria, endoplasm reticulum and other organelles.
Immunoelectron microscopic localization of plateins (alveolar plate proteins) during the morphogenesis of new cell cortex in the ciliate Euplotes
Authors: Kloetzel John A.
Published: June 2020

The ciliated protozoan Euplotes possesses a complex membrane-cytoskeletal surface layer, or pellicle, that appears to be a primary determinant of cell form. Flattened membranous sacs, termed alveoli, are arranged in a confluent monolayer just beneath the plasma membrane. Each alveolus contains a discrete polygonal alveolar plate (AP); it is likely that the tightly packed mosaic sheet of APs (together with subalveolar microtubules) provides the rigidity of the cell cortex and dictates the characteristic contours of the cell. This pellicular structure must be duplicated daily during vegetative growth of the cell, following a stereotypical program of cortical morphogenesis the stages of which have been well described at the light microscopic level. At the ultrastructural level, models of new pellicle formation have been presented; both suggest that new alveoli form beneath the expanding plasma membrane by the fusion of numerous specialized membranous vesicles, 0.1-0.4 (μm in diameter, termed “alveolar precursor vesicles” (APVs). It is proposed that the plates form within the growing alveoli by the condensation or polymerization of the electron-dense core material seen within the APVs.
Unusually large magnetite (Fe3O4) crystals inside a magnetotactic bacterium
Authors: Farina Marcos , Lins de Barros Henrique G.P. , Esquivel Darci M.S.
Published: June 2020

Magnetite crystals inside magnetotactic microorganisms have been shown to cover a wide range of dimensions and shapes. However in almost all cases reported until now, crystals width and length remain under the magnetite monodomain dimensions when using Butler and Banerjee theoretical curves. We report here one type of magnetotactic bacterium which produces very large magnetite crystals as compared with those described in the literature. In a previous communication we had notified this fact, but without detail.

Samples were collected from brakish water from ITAIPU lagoon near Rio de Janeiro (24° Southern hemisphere). In the same sample it was possible to find different magnetotactic microorganisms. After standard isolation treatments we obtained different groups of crystals identifyed as magnetite (Fe3O4) by electron diffraction.
An Electron Microscopic evaluation of the virus removability of regenerated cellulose hollow fibers (BMM)
Authors: Yamaguchi Kazuhito , Hamamoto Yoshiaki , Yamamoto Naoki , Sekiguchi Sadayoshi , Yamashiki Takashi
Published: June 2020

We examined the virus removability of a newly developed filter membrane, regenerated cellulose hollow fibers (BMM) manufactured by means of a microphase separation method. The BMM is proposed for clinical use of ridding infectious viruses from plasmas which are infused into patients, ie. hemophilia, for a treatment. A BMM hollow fiber has approximately 300um inner-diameter and 38um wall-thickness. The removability of BMMU80 (mean pore size 73nm) and BMMU40 (45nm) were examined in this study. For filtration, single modules consisting of a fiber (filtrating area 0.6cm2) and filter units consisting of 500 fibers (300cm2) were prepared. In order to evaluate the removability, colloidal gold solutions of 68, 36 and 22nm and human immunodeficiency virus (HIV), hepatitis B virus (HBV) and Japanses infectious encephalitis virus (JEV) were filtered at the pressure of 200mmHg.
Immunoelectron microscopy of the intracellular compartments of prespore cells of dictyostelium discoideum
Authors: Erdos Gregory W. , O'Brien Bonnie J. , West Christopher M.
Published: June 2020

In the amoeboid cellular slime molds two cell types are formed during differentiation, prestalk cells and prespore cells which ultimately become the stalk and spore cells of the mature sorocarp. The prespore cells can be identified ultrastructurally by the prespore vesicles (PSV), which have a dense layer lining the inside of the compartment (Fig. 1). The PSV's are known to contain several glycoproteins and a galactose polysaccharide that are released to the cell surface to become major components of the spore coat. Further investigations in this laboratory, by three dimensional reconstruction from serial thin sections and by confocal fluorescence microscopy have shown that the PSV's are not a collection of separate vesicles, but rather a tubular reticulum asymmetrically disposed in the cell. Similar geometry has been shown for some lysosomal systems. A recent report has shown that the lysosomal enzyme, α-mannosidase, can be localized to the PSV. These pieces of evidence suggest a possible lysosomal function or lysosomal origin for the prespore secretory system. In an effort to better characterize the PSV compartment we have chosen two monoclonal antibodies, 5G7 which recognizes a common lysosomal epitope and 81.2 which recognizes a carbohydrate epitope on the plasma membrane and on compartments of phagocytotic and autophagic origin. We further chose to characterize the distribution of acidic compartments using the DAMP method of Anderson, which would give us an indication of functional lysosomal activity.
Morphological changes of Helicobacter pylori caused in vitro by cytoprotective anti-ulcer drug, Sofalcone
Authors: Kondo Isamu , Yamaguchi Masashl
Published: June 2020

Since Helicobacter pylori has recently been regarded as a possible causative agent of a chronic gastritis and a peptic ulcer disease, the susceptibilities of this organism to anti-ulcer agents or antibiotics have attracted much interest of not only clinical docters but also medicobiologists.

We have reported that Sofalcone, a cytoprotective anti-ulcer drug, has anti-bacterial and bactericidal activity as well as TDS (tripotassium dicitrats bithmuthate) and Clarithromycin (a derivative of erythromycin), but the other anti-ulcer drugs Including anti-acid or H2 receptor antagonists such as cimetidine could not show such bacterial activity as those shown by the sofalcone (SFC), TDB, and clarithromycin (CLM). It was also found that therewere distinct differences between the morphological damage of H. pylori caused by SFC and those caused by TDB and CLM.
An ultrastructural study of ascospore delimitation and maturation in Ascodesmis nigricana using freeze substitution fixation
Authors: Mints C. W. , Richardson E. A. , Kimbrough J.
Published: June 2020

Fungi belonging to the class Ascomycetes are characterized by the production of sexually derived spores termed ascospores inside a microscopic, sac-like structure termed an ascus. In the Euascomycetidae, typically uninucleate spore initials are delimited within the ascus as the result of the invagination of membranes that arise from the so-called "ascus vesicle", a discontinuous cylinder of two closely spaced unit membranes that develops around the extreme periphery of the ascus. To date, there are conflicting data regarding the origin of the ascus vesicle. We therefore decided to address the problem using freeze substitution fixation. The advantages of this procedure over conventional chemical fixation protocols for preservation of ultrastructural details in fungi have been well-documented. In this study small pieces of dialysis membrane bearing developing ascocarps ofAscodesmis nigricanswere plunged into liquid propane and processed for TEM according to the procedures of Hoch and Mints et al.
Structures of cotton motes and mote fibers
Authors: Goynes W. R. , Ingber B. F. , Thibodeaux D. P.
Published: June 2020

A mote is a seedlike structure that is derived from an ovule, but that does not develop into a mature seed. In cotton, motes vary in size from tiny structures having no fibers, to those that appear to be full-sized seed with fibers approximately the length of fully-developed fibers. The size of the mote depends on the stage at which its development was arrested. In the cotton industry these undeveloped seed, as well as the quality of fibers produced by them, are of commercial importance. Results presented here are from an exploratory study to show differences between structures of normal seed and fibers, and motes and mote fibers.

Cotton seed develop within a closed boll that is separated into compartments or locules. Embryo development is normally completed within the seed in approximately 35 days. Fiber growth initiates on the seedcoat surface as cells elongate to become thin-walled tubes until approximately 18-20 days postanthesis when the main body of the fiber, the secondary wall, begins to be laid down.
Freeze-substituted fungal cells of Nectria haematococca: A comparison of the macroconidial walls of the adhesion-competent wild type with an adhesion-reduced mutant
Authors: Caesar-Ton That Thecan , Epstein Lynn
Published: June 2020

Nectria haematococca mating population I (anamorph, Fusarium solani) macroconidia attach to its host (squash) and non-host surfaces prior to germ tube emergence. The macroconidia become adhesive after a brief period of protein synthesis. Recently, Hickman et al. (1989) isolated N. haematococca adhesion-reduced mutants. Using freeze substitution, we compared the development of the macroconidial wall in the wild type in comparison to one of the mutants, LEI.

Macroconidia were harvested at 1C, washed by centrifugation, resuspended in a dilute zucchini fruit extract and incubated from 0 - 5 h. During the incubation period, wild type macroconidia attached to uncoated dialysis tubing. Mutant macroconidia did not attach and were collected on poly-L-lysine coated dialysis tubing just prior to freezing. Conidia on the tubing were frozen in liquid propane at 191 - 193C, substituted in acetone with 2% OsO4 and 0.05% uranyl acetate, washed with acetone, and flat-embedded in Epon-Araldite. Using phase contrast microscopy at 1000X, cells without freeze damage were selected, remounted, sectioned and post-stained sequentially with 1% Ba(MnO4)2 2% uranyl acetate and Reynold’s lead citrate. At least 30 cells/treatment were examined.
SEM and fluorescence imaging of callose deposition in root hair tips and suspension cultures of Streptanthus tortuosus
Authors: Walters K.S. , Sjolund R.D. , Moore K.C.
Published: June 2020

Callose, B-1,3-glucan, a component of cell walls, is associated with phloem sieve plates, plasmodesmata, and other cell wall structures that are formed in response to wounding or infection. Callose reacts with aniline blue to form a fluorescent complex that can be recognized in the light microscope with ultraviolet illumination. We have identified callose in cell wall protuberances that are formed spontaneously in suspension-cultured cells of S. tortuosus and in the tips of root hairs formed in sterile callus cultures of S. tortuosus. Callose deposits in root hairs are restricted to root hair tips which appear to be damaged or deformed, while normal root hair tips lack callose deposits. The callose deposits found in suspension culture cells are restricted to regions where unusual outgrowths or protuberances are formed on the cell surfaces, specifically regions that are the sites of new cell wall formation.

Callose formation has been shown to be regulated by intracellular calcium levels.
Electron Energy Loss Spectroscopy as an alternative method to follow up the route of activators and drugs in cells and tissues
Authors: Wolf B. , Uhl B. , Schwinde A. , Hauschildt S. , Bessler W.G.
Published: June 2020

Investigations on uptake and distribution of small molecules and drugs in cells and cells from solide tissues are very important for the evaluation of their physiological behaviour and/or therapeutical potency. In electron microscopy localizations of molecules are usually performed by immunocytochemical techniques using antibodies coupled to electron dense markers (e.g. ferritin, gold). Problems concerning labelling precision and the production of highly specific antibodies to small molecules resulted in the evaluation of a more sensitive and more physical technique.

We applied analytical electron energy loss spectroscopy (EELS) on ultrathin cryosections of different cells incubated with various substances (lipopeptide, 5’fluoro-uracil, cis-platinum, etc.) to follow up the route of these molecules from the plasma membrane to ultrastructural target regions. Using cryosections, investigations on almost native material, without the use of disintegrative methods, are possible. The main advantage of this procedure is the exclusion of metabolic interferences after the incubation period.

Here, we demonstrate analytical energy loss spectroscopy as a possibility to analyze the distribution of small molecules within cells directly without the use of labelled antibodies.
Study on erythrocytes by SEM photogrammetry (SEMP) technique
Authors: Wensheng Tong , Lianhuang Lu , Zhijun Zhang
Published: June 2020

This is a combined study of two diffirent branches, photogrammetry and morphology of blood cells. The three dimensional quantitative analysis of erythrocytes using SEMP technique, electron computation technique and photogrammetry theory has made it possible to push the study of mophology of blood cells from LM, TEM, SEM to a higher stage, that of SEM P. A new path has been broken for deeply study of morphology of blood cells.

In medical view, the abnormality of the quality and quantity of erythrocytes is one of the important changes of blood disease. It shows the abnormal blood—making function of the human body. Therefore, the study of the change of shape on erythrocytes is the indispensable and important basis of reference in the clinical diagnosis and research of blood disease.

The erythrocytes of one normal person, three PNH Patients and one AA patient were used in this experiment. This research determines the following items: Height;Length of two axes (long and short), ratio; Crevice in depth and width of cell membrane; Circumference of erythrocytes; Isoline map of erythrocytes; Section map of erythrocytes.
Ultra grids for observation of same wide specimen at low and high magnification
Authors: Hirose Haruko , Shirota Kohei , Makita Takashi
Published: June 2020

Inconvenience in observation of wide view of biological sections was to prepare two sections from same sample; one thicker and heavily stained for contrasity at lower magnification and another ultrathin and lightly stained for higher magnifications. High contrasity and less deformative characteristics of currently developed electron microscopes,EM-002A(120kv) and EM-002B(200kv) from ISI.ABT,allow us to examine same section at both low and high magnifications.Besides to superwide(Fig.1-a) and wide (Fig.1-b) grid of 7mmØ those which have been utilized for super wide field electron microscopy using a specially constructed conventional electron microscope(LEM-2000, ISI,ABT)(1), a new type of ultra grid was designed in this laboratory(Fig. 1-c) and it became commercially available.

Though wide- and superwide- grids have wider squares than those in ordinary 3mmØ grids, we have to cut off some bars with a razor blade to obtain further wider vision in some cases.(2-6). Ultra grid(Fig.1-c,Fig.2) eliminated that inconvenience since it has several types of large compartments.The largest square is 1.5×2.Omm and equivalent to 9 squares in super wide grid.
En bloc staining available for stereoscopic observation of epoxy resin Quetol 651-embedded thick sections under a 300 kV TEM
Authors: Kushida Tsuyuka , Iijima Haruyuki , Kushida Hiroshi , Tsuruta Chusei
Published: June 2020

A staining method has been devised for easy en bloc staining for stereoscopic observation of epoxy resin Quetol 651-embedded thick sections under a 300 kV transmission microscope (TEM). In order to enhance staining properties in thick section, osmium tetroxide-fixed tissue blocks are stained only en bloc, since the images of both sides in thick section give high contrast and the image of an intermediate layer shows low contrast by double staining.

This method uses carbohydrazide (Polysciences, Inc., U.S.A.) as osmium bridging agent, and both osmium tetroxide and uranyl acetate as electron staining agents.

The following procedure is suitable for en bloc staining.

1.

Fix small tissue blocks in 2% cacodylate-buffered osmium tetroxide (pH 7.4) for 3 hours at 4°C.

2.

Wash well in buffer for 1 hour.

3.

Transfer in 1% aqueous carbohydrazide for 2 hours at room temperature.

4.

Wash well in distilled water for 1 hour.

5.

Stain in 1% aqueous osmium tetroxide for 2 hours at room temperature.

6.

Wash well in distilled water for 1 hour.

7.

Dehydrate in 50% alcohol for 1 hour.

8.

Stain in a 2.5% solution of uranyl acetate in 50% alcohol for 3 hours at room temperature.

9.

Wash in 50% alcohol for 1 hour.

10.

Dehydrate with 60%, 70%, 80%, 90% and 100% (2 changes) alcohols for 30 minutes each.

11.

Embed in a mixture of Quetol 651 (Nissin EM Co., Ltd., Japan), nonenyl succinic anhydride, methyl nadic anhydride and DMP-30 according to the method of Kushida et al.
Localization of α1-proteinase inhibitor binding to extracellular matrix by tandem scanning reflected light and transmission electron microscopy
Authors: Roemer E.J. , Rinehart A.R. , Simon S.R.
Published: June 2020

α1-Proteinase Inibitor (α1-PI), a major inhibitor of human leukocyte elastase (HLE), has been shown to bind saturably to extracellular matrix (ECM), composed of 51% glycoproteins and proteoglycans, 37% types I and III collagen and 12% elastin, produced by R22 rat aortic smooth muscle cells. To determine the components of the ECM to which α1-PI binds we have employed both tandem scanning reflected light (TSM) and transmission electron (TEM) microscopy on whole mount and sectioned material immunolabeled with antibodies to α1-PI and to various ECM components.

R22 cells were grown on glass coverslips for TSM, on coverslips with formvar and carbon coated gold grids for whole mount TEM, and in 60 mm dishes for embedment. At confluence ascorbic acid was added for 6-18 days. The cells were lysed with 5 mM NH4OH at 25°C and the remaining ECM was rinsed with PBS and dH2O. ECM was blocked with 0.1% BSA in PBS for 30 min at 25°C and then incubated with 1.3 mg α1-PI/ml in 0.1% BSA/PBS for 1.5-2 h at 37°C.
Effects of ultrasound treatment of the arteria thoracica (A. mammaria) interna (ATI) during preparation for coronary artery bypass surgery (CABG): A correlative light microscopic, scanning and Transmission electron microscopy study
Authors: Muss Wolfgang , Lametschwandtner A. , Schistek R. , Ghosh P.K. , Thurner J. , Unger F.
Published: June 2020

Ultrasonic Aspirators (USA) or Ultrasonic Dissectors, respectively, are used in a variety of surgical disciplines . The functional principle is to fragment and aspi rate tissue components using a piezoelectric PZT (lead, zirconium, tantalum) ceramic electrostrictive transducer tip, which "vibrates" in Tts long axTs at about 23-24 kHz producing longitudinal waves.

This study deals with the preparation of the ATI for CABG and compares the effects of conventional preparation of ATI by forceps, scalpel and cautery vs. USA-Application (SONOCORR; SOERING Ltd, Quickborn/FRG), whereby two different energy levels (5%, 25% of the maximal available energy (100W) ) were tested. The aim of the study was to check the effects of the USA on endothelial morphology. Two experiments were done: first, the distal ends of ATIs from four patients immediately prior to CABG were prepared either (two of them) conventionally (cf. above) or by use of USA (one at 5%, one at 25% of max. available energy).
A quantitative study of silver enhancement of differently sized gold labeling particles for postembedding indirect immunogold EM
Authors: D'Andrea Michael , Forte Michele , Gu Jiang
Published: June 2020

The post-embedding indirect immunogold staining method has been widely used in electron microscopy. In this technique, colloidal gold labeling particles are round, electron dense and uniform in size with diameters from 5nm to 40nm. At optimal conditions, the smaller the labeling gold particles, the higher the maximum detecting sensitivity (MDS) for the immunostaining. Janssen Pharmaceutical (Piscataway, NJ), developed 1 nm gold, perhaps the most sensitive gold probe to date, in 1989. Even at the higher electron microscopic magnifications, x100,000 - 200,000, the resolution of the lnm gold particles is not clear enough for direct viewing. To overcome this limitation, a silver treatment procedure is used to enlarge the gold particle size. Several studies demonstrated the applications of gold-silver staining but not quantitatively. We tried to correlate the enlargement of gold-silver particle sizes to the silver staining duration using as a model atrial natriuretic peptide (ANP), a newly discovered cardiac hormone located in specific electron dense granules in the cytoplasm of atrial myocardiocytes.
New support film by the plasma polymerization method
Authors: Yamaguchi Masashi , Tanaka Akira , Suzuki Teruo
Published: June 2020

The plasma polymerization replica method and the apparatus required were deviced by Tanaka. In this technique, ionized gas molecules from a glow discharge are instantly polymerized on the surface of a specimen. The polymerized molecules make a uniform, ultrathin film which precisely copies the three-dimensional ultrastracture of the specimen's surface. In the course of our study, we soon realized that the polymerized film would serve as a good support film. We report here a method of preparation of the new support film by plasma polymerization and its application.

Method of preparation: Newly cleaved NaCl or KC1 crystals of 1-2 mm thickness, newly cleaved mica, or coverglass were placed on the cathod plate in a bell jar of the Plasma nano-replica apparatus(PNR-110, Ushio Inc., Tokyo). After evacuation of air, the bell jar was filled with hydrocarbon monomer gas(naphthalene etc.) to about 10 Pa. A high voltage (2-2.5 kV D.C.) was then applied across the electrodes to generate a glow discharge, and a very thin, threedimensionally polymerized film was formed on the surface of substrates.
The luminescent probes europium and terbium-DTPA localize in cells and organs and serve as analogues for gadolinium-DTPA
Authors: Allen Nina Stromgren , Jackels Susan C. , Elster Allen D. , Marrache Ron , Sizemore Julia S.
Published: June 2020

The Europium (III) and Terbium (III) complexes of diethylene-triaminepenta-acetate (DTPA) were prepared to be luminescent, histochemical analogues for the recently developed specific paramagnetic resonance contrast agents such as Gadolinium (III)DTPA. Electron probe microanalysis (energy dispersive X-ray spectroscopy) demonstrates that the Eu and Gd-DTPA colocalize in various tissues. Eu, Tb and Gd are closely related as seen in the Periodic Table (Fig. 1). The structures of Eu and Gd-DTPA (Fig. 2) as well as Tb-DTPA are very similar. Fig 3 shows the synthetic path of one of these compounds, Na2Eu (DTPA)H2O. Fig. 4 depicts a Europium(III) complex fluorescence-excitation scheme, where UV (360 - 440 nm) excites electron energy to the 5D7 state, followed by a rapid (less than 5 μec) non-radioactive relaxation to the luminescent D0 state. There is a slow return to the ground 7F manifold with emission from 575-790 nm. Crystals of the Europium complex exhibit intense, non-fading fluorescence (Fig. 5) in a narrow red band when observed with a UV excitation filter (360-440 nm), a midrange chromatic beam splitter (460 nm), and a long pass barrier filter (590 nm). The same crystal is seen in Differential Interference Contrast (DIC) light microscopy (LM) (Fig. 6).
Electron probe microanalysis of calcium and potassium in cryo-preserved cerebral cortex in situ: A comparison of wet and dry sectioning
Authors: Lightfoot F.G. , Rosenthal R.E. , Fiskum G.
Published: June 2020

The sub-cellular concentration of calcium and potassium are of particular interest, and have been elusive to investigators not utilizing cryo-ultramicrotomy and cold stage microscopy. By using improved methods in cryo-fixation and tissue drying, one now has an alternate approach to determining ion concentration within the cell. Under certain experimental regimens, metal-mirror fixation is not plausible, and modifications must be adapted.

Adult male Beagles, with ischemic brain damage, were used in this study. After the appropriate experimental design of each dog, following anesthesia, a left parietal craniotomy was done and samples of cerebral cortex were removed and processed for biochemical analysis. For analytical electron microscopy (AEM) the dura was removed and the endomarginal gyrus of the parietal lobe was cryofixed using cryo-pliers specifically designed for this procedure. Once the gyrus had been rapidly frozen it was transferred to a cold block for freeze drying under high vacuum (10-5Torr) for 48 hours. After drying, the tissue was fixed for 30 minutes in osmium vapors and infiltrated with Araldite resin and polymerized.
Radial mass analysis of unstained STEM images of molluscan hemocyanins
Authors: Hamilton M.G. , Herskovits T.T. , Wall J.S.
Published: June 2020

The hemocyanins of molluscs are aggregates of a cylindrical decameric subparticle that assembles into di-, tri-, tetra-, penta-, and larger multi-decameric particles with masses that are multiples of the 4.4 Md decamer. Electron micrographs of these hemocyanins typically show the particles with two profiles: circular representing the cylinder viewed from the end and rectangular representing the side-view of the hollow cylinder.

The model proposed by Mellema and Klug from image analysis of a didecameric hemocyanin with the two decamers facing one another with collar (closed) ends outward fits the appearance of side-views of the negatively-stained cylinders. These authors also suggested that there might be caps at the ends. In one of a series of transmission electron microscopic studies of molluscan hemocyanins, Siezen and Van Bruggen supported the Mellema-Klug model, but stated that they had never observed a cap component. With STEM we have tested the end cap hypothesis by direct mass measurements across the end-views of unstained particles.
An integrated screening method for evaluating the pulmonary toxicity of inhaled particulates: Role of microscopic techniques in assessing pulmonary macrophage clearance functions
Authors: Warheit David B. , Achinko Lena , Hartsky Mark A.
Published: June 2020

There is a great need for the development of a rapid and reliable bioassay to evaluate the pulmonary toxicity of inhaled particles. A number of methods have been proposed, including lung clearance studies, bronchoalveolar lavage analysis, and in vitro cytotoxicity tests. These methods are often limited in scope inasmuch as they measure only one dimension of the pulmonary response to inhaled, instilled or incubated dusts. Accordingly, a comprehensive approach to lung toxicity studies has been developed.

To validate the method, rats were exposed for 6 hours or 3 days to various concentrations of either aerosolized alpha quartz silica (Si) or carbonyl iron (CI) particles. Cells and fluids from groups of sham and dust-exposed animals were recovered by bronchoalveolar lavage (BAL). Alkaline phosphatase, LDH and protein values were measured in BAL fluids at several time points postexposure. Cells were counted and evaluated for viability, as well as differential and cytochemical analysis. In addition, pulmonary macrophages (PM) were cultured and studied for morphology, chemotaxis, and phagocytosis by scanning electron microscopy.
Intraneuronal accumulations of lysosomes in the cerebellum of rats and dogs after dosing with MDL-832
Authors: Barnard S.D. , Warner S.D.
Published: June 2020

1, 2, 9, 10-tetramethoxyaporphine phosphate (MDL-832) was once considered a potential human antitussive. MDL-832 was administered orally in the diets of Sprague-Dawley rats at dose levels of 0, 5, 10, 20, 40, 80 and 160 mg/kg/day for 3 and 6 months and in gelatin capsules to Beagle dogs at 0, 5, 10, 15, 30 and 60 mg/kg/day for 3, 6 and 12 months. Histopathologic examinations of hematoxylin and eosin-stained cerebellar sections revealed intracytoplasmic brown pigment accumulations in large fusiform neurons (presumably the motor type) of the pons. The pigment granules were found to be PAS-positive, non-acid fast, iron-free, Sudan B-positive and fuchsinophilic. Intraneuronal pigment accumulations were seen in rats after 3 months of treatment at 80 mg but not at 40 mg and after 6 months at 20 mg but not at 10 mg. For dogs the effect was observed after 3 months at 60 mg but not at 30 mg and after 12 months at 10 mg but not at 5 mg.
SEM of a self-expanding vascular stent in a canine animal model
Authors: Laliberté Verdon S. , Gould K. E. , Termin P. L. , Connolly R. J. , Belkin M. , Breckwoldt W. , Allen M. J. , Hayes L. L. , Callow A. D.
Published: June 2020

Since the first reported use of a coilspring endovascular prosthesis to improve the outcome of recanalization and ballon angioplasty in peripheral atherosclerotic vessels, a nonthrombogenic, autologous lining has been considered critical for longterm patency. We have investigated the morphology of canine illiac arteries by scanning electron microscopy (SEM) at 3, 7, 21 and 28 days after placement of self-expanding stainless steel stents (Wallstent®).

Purpose-bred mongrel dogs, purchased from USDA licensed vendors were housed in the AAALAC approved animal facility at New England Medical Center. Stents (n=18) were placed by percutaneous cannulation into the illiac arteries under fluoroscopic control. The area of each artery containing the stent was excised and perfusion fixed with 3% 0.1M cacodylate buffered glutaraldehyde. Samples were rinsed in 0.1M cacodylate buffer and post-fixed for 60 min. in 0.1M cacodylate buffered 1% Osmium tetroxide. Samples were then rinsed in double-distilled water and fixed for 60 min. in 1% aqueous tannic acid. Samples were dehydrated through graded alcohols, placed in hexamethyldisilazane (HMDS) for 10 minutes and subsequently air dried overnight. Dried specimens were mounted on aluminum stubs with silver paste and examined at 10-20 kv on an Amray 1000B scanning electron microscope.
Visualization of capsular polysaccharide of coagulase negative staphylococci (CNS) associated with vascular prosthetic graft material
Authors: Goheen Michael P. , Edmiston Charles E. , Khan Saeed A. , Maguire Nancy K. , Schmitt David D.
Published: June 2020

The coagulase negative staphylococci (CNS) are a ubiquitous group of microorganisms which comprise a major component of the microbial flora of the skin and adjacent glands. Historically, the CNS were viewed as insignificant contaminants with little clinical importance. However, over the last decade these organisms have emerged as important etiologic agents of biomaterial associated infections. Furthermore, many of the Staphylococcus epidermidis strains recovered from catheter or prosthetic infections exhibit the capacity to produced an extracellular capsular polysaccharide (slime) which is viewed as an important factor in biomaterial adherence and persistence. Bacterial capsular material is composed of hydrated polysaccharides and has been visualized by transmission electron microscopy when lysine is added to the fixation protocol. This study reports on the use of a lysine-ruthenium redglutaraldehyde fixation technique for visualizing the polysaccharide capsule on two coagulase negative staphylococcal reference strains, S. epidermidis RP62A (ATCC 35984) and S. hominis SP2 (ATCC 35982).
Preservation and microscopy of bacterial biofilms causing corrosion on steel surfaces
Authors: Aldrich H.C. , McDowell L. , Choate A. , Bono P.
Published: June 2020

Steel methane transmission lines are sometimes attacked by anaerobic bacteria which cause deep corrosion pits and represent a serious economic problem. We have developed a model laboratory system to follow the process of bacterial attachment and colonization of test specimens of pipeline steel. Test coupons of X and B grades of steel approximately 5mm x 10mm x 20mm were exposed to combinations of the bacteria Enterobacter aerogenes, Desulfovibrio desulfuricans, and Clostridium acetobutylicum in an aerobic culture for periods ranging from 2 hr. to 30 days.

A method is described for preserving bacterial films on the surface of the coupons, so that both the bacteria and the corrosion may be studied with light microscopy and scanning and transmission electron microscopy (LM, SEM, TEM). Upon removal from the culture vessel, coupons are fixed in 2.5% glutaraldehyde, dehydrated in ethanols, and embedded in Spurr’s embedding resin, all within a glove bag purged with nitrogen to prevent oxidation of the steel. After polymerization, the plastic is separated by chilling to -80 C. and dropping into hot tap water. The plastic surface next to the metal carries the bacterial film intact, along with impressions of corrosion sites.
Cytochemical study in ischemia injury of renal proximal tubule epithelia cells and the preventive effect of inosine and Ligusticum wallichi
Authors: Zun Zheng , Jihong Sha
Published: June 2020

The cytochemical methods were used to evaluate the changes of intracellular calcium,plasma meabrane permeability and their relationship during ischemia and reflow in rat renal proximal tubule epithelia cells. In addition, the preventive effects of inosine and Ligusticum Wallichi(LW) on ischemia and reperfusion cell injury were studied and compared.

Renal ischemia was induced by 60 min. of renal pedicle clamping in male Spraque-Dawley rats (180-230g) and 2,4, and24 hr. reflow after 60 min. ischemia. The inosine and LW were administed 10 min. and 30 min. respectively before ischemia (Inosine 100mg/100g IV : LW 0.2ml/100g IM;LW is a traditional Chinese medicine, supplied by Chang Hai hospital, Shanghai, China). Calcium was selectively localized with oxalate-pyroantimonate precipitation procedures and analysed by EDX. Lanthanum (La) was employed as a tracer of plasma membrane permeability.

Result, Ultrastructural morphology, By the end of 60 min. ischemia, there were no significant differences of cell damage between drug treated groups and untreated groups (Fig.1) but the cell damage would be more and more serious following extension of reflow time.
Ultrastructural localization of immunoglobulin M in activated B-cells in patients with epidemic hemorrhagic fever using gold probe
Authors: Chen Dehui , Guo Ning , Zhang Heqiu , Wang Guohua , Sun Yingxun , Bai Yan
Published: June 2020

Our previous electron microscopic studies have shown for the first time that the atypical lymphoid cells in the peripheral blood of patients with epidemic hemorrhagic fever (EHF) during the acute febrile phase of the illness appeared to be activated B-cells and plasma cells of various developmental stages. But it is still not a tall clear whether those initially activated cells were actually B-cells. In this study, an indirect immuno gold staining method for the detection of B-cell surface antigen was used.

Peripheral blood mononuclear cells were isolated from anticoagulated venous blood of 3 EHF patients (2—7 days of illness) and control donors by centrifugation at 2,500 Xg for 15 min. The cells obtained from the buffy coat were washed in TBS and resuspended in TBS-5% BSA. Unfixed cells were in cubated insuspension in step I with monoclonal antibody (MS27) directed against human IgM (μ Chain) for 30 min at 37±C. After washing in TBS—BSA, the gold probe was applied to the cells in step II incubation with GAM-G15 reagent for 45 min at RT. The gold probe was prepared in our laboratory according to Slot and Geuze. After the third washing in TBS-BSA, the cell pellets were fixed, dehydrated and embedded in Epon 812.
Immunolocalization of Type VI collagen in calfskin: A correlative study
Authors: Kronick Paul L. , Maleeff Beverly E. , Dahms Matthew P.
Published: June 2020

Although fine details of the fundamental structural units of calfskin, the types I and III collagen fibrils, have been well elucidated, little is known about the location of “minority” collagens in the skin before and after processing to leather. Using microscopic methods, we investigated one such collagen, type VI, in calfskin to determine its presence and distribution.

Four different methods of identification of type VI collagen were employed: immunofluorescence and peroxidase-antiperoxidase (PAP) staining using light microscopy, and transmission (TEM) and scanning (SEM) immunoelectron microscopy. In all cases, the primary antibody used was a polyclonal anti-type VI collagen raised in rabbitss. For light microscopy, 12 μm frozen sections were prepared. Fluorescent labeling was carried out with anti-rabbit IgG conjugated to r-phycoerythrin as secondary antibody. PAP staining was performed with the modified techniques of Amenta et al. Tissue for SEM was prepared by a modified protocol of Keene et al. without prefixation: the secondary antibody used was goat anti-rabbit IgG conjugated to 30 nm colloidal gold; it was then fixed in glutaraldehyde-paraformaldehyde and OsO4, dehydrated, critical point dried and sputter coated with gold.
Immunocytochemical mapping of the biosynthetic pathways of two major proteins expressed in terminally differentiated epidermal keratinocytes
Authors: Bisher M.E. , Roop D.R. , Steinert P.M. , Steven A.C.
Published: June 2020

In the process of constant renewal of mammalian epidermis, cells divide and move upwards from the basal layer as they embark on a program of terminal differentiation. Cells located higher in the epidermis - the spinous, granular, and cornified layers - represent progressively later steps in this pathway. Commitment to terminal differentiation is accompanied by major changes in gene expression and morphology.Inter alia, large-scale synthesis of several proteins is initiated: these include filaggrin, a protein thought to be responsible for aggregating keratin filaments, and a recently discovered protein called loricrin, which has been implicated as a likely major component of the covalently cross-linked protein lining of the cornified cell envelope. We have used immunoelectron microscopy to investigate the biosynthetic pathways of filaggrin and loricrin.

Immediately after sacrifice, skin samples were taken from newborn mice, and diced into blocks of 1-2 mm3. They were then either (i) fixed with 1% glutaraldehyde, 0.2% picric acid, in PBS and embedded in LR White, or (ii) fixed with 1% glutaraldehyde in PBS, and embedded in Lowicryl K4M. Essentially the same conclusions were reached from the experiments with both resins.
Al localization in plant roots using EDXA and EELS techniques
Authors: Marienfeld Stephan F.-W. , Stelzer Ralf , Lehmann Heiner
Published: June 2020

Aluminum, which is soluble at pH 5.0 is toxic to plants and frequently the most important yield limiting factor of crop production, on acid soils. Moreover, Al has been associated with human mental diseases (e.G. Alzheimer).

Phytotoxicity of Al primarily results in adecline of mitotic activity of the root meristem, an evident reduction of root growth and subsequently affects mineral nutrition and water supply. Several authors assume a shifting of Alin to the nucleus and a direct influence on DNA-synthesis to be the reason for this growth reduction. However, Al in nucleus has only been demonstrated after preparations allowing Al to shift during this preparation. Using SEM and microprobe-analysis, it was only possible to demonstrate Al in roots mainly to be absorbed (as Al-phosphate) in cell walls. A quantification in cytoplasma or even a localisation in distinct organelles (e.g. the nucleus) fails due to the low tracability and the in sufficient resolution.

Therefore, in addition to EDX-analysis we try to localize Al by using electron spectroscopic imaging (ESI) and electron energy loss spectroscopy (EELS) techniques, which are distinguished by an excellent spatial resolution and a high sensitivity. So element scan be analyzed and localized in small cell compartments. For those observations the plant material has to be fixed, embedded and cutin ultrathin sections. During chemical fixation procedure, however, the membranes lose their semipermeability and substances may be extracted or shifted inside the tissue or the cells. There fore elemental analysis must be done only under special conditions.
Application of video EM and image processing for evaluation of barrier tissues endocrine activity after environmental chemical exposure
Authors: Ptashekas J.
Published: June 2020

Biotesting procedure of new environmental chemicals is unavoidable within framework of environmental medicine and public health. Micrographs in transmission electron microscopy are indispensable for evaluation of fine details of ultrastructure of diffuse endocrine system cells located in barrier epithelial tissues of gastrointestinal tract. At the same time it is insufficient to obtain integrated image and tissue functional activity analysis. Videoprocessed electron microscopy provides for full section (s) visual information and is applicable in environmental medicine.

Wistar rats 250-280 gr b.w. had been once tube fed with 0.1 DL50 solution of piperasin derivative herbicide saprol (Celamerck, FRG) and investigated 15 min., 4 h and 24 h after treatment. Vertical samples of pyloric mucosa were prepared for transmission electron microscopy (TEM). Separate types of gastric endocrine cells had been verified ultrastructurally from micrographs according to lausanne (1981) Enteroendocrine Cells Classification. TEM image had been videoprojected to the monitor and computer processed to obtain distinct electron optical density, maturity level and number of secretory granules within 1 sq mkm of endocrine cell cytoplasma. Granules size (nm) frequency distribution histograms provided as well.
Radioactive and Non-Radioactive In Situ Hybridization Techniques
Authors: Durrant I.
Published: June 2020

In situ hybridization is a powerful technique that has found multiple applications in the biological sciences. The most widely used technique is in the analysis of mRNA species in cell populations. This is particularly useful for the analysis of relative amounts, site of transcription, timing and induction of transcription. Recent advances have lead the technique into the areas of relative quantification, dual signal detection and disease diagnosis. DNA targets can also be visualised by in situ hybridization both in cells and tissues and on isolated chromosomes and nuclei. There are a variety of systems that can be used to detect hybridization signals in situ. These choices are based in two areas, probe type and label type and this selection process holds true for both radioactive and non-radioactive systems.

Probe selection is important and plays a part in the overall success of the application (see Table
1). RNA probes and oligonucleotide probes are the most widely used in cells and tissues whereas
DNA probes are routinely used for chromosome and nuclei targets. RNA probes are used due to
the higher sensitivity obtained but oligonucleotide probes are gaining in popularity due to ease
of use. The lower sensitivity seen with these probes, due to lower labelling capacity, can be
overcome by use of probe cocktails.
Magnetic Resonance Analysis of Mouse Embryos with Altered Gene Expression
Authors: Smith B. R.
Published: June 2020

Techniques to assess and analyze phenotypes of experimentally altered animals have not kept pace with our ability to manipulate the genome. Assessment of the shape, position, and timing of the appearance of anatomical structures is indispensable to identifying and understanding the mechanisms of normal and abnormal morphogenesis. Tools are needed to effectively diagnose the condition of the hundreds of experimentally altered animals being produced.

Magnetic resonance (MR) microscopy is a non-destructive technique capable of generating three-dimensional images of developing structures from whole embryos. It facilitates the evaluation of complex anatomical structures by allowing them to be studied in any plane of sectioning for cut-away views and from any angle for surface views.

MR microscopy has been used to assess the following mutant mouse embryos and mouse embryos with altered gene expression: Splotch, Curly-tail, Loop-tail, TS 16, HOX 2.2 transgenic, MARKS knockout, LH-2 knockout, and Dlx-2 knockout. Imaging was performed on a modified 9.4 Tesla GE NMR Instruments Omega system with echo times of 6 ms and repetition times of 200 ms.