bluuebunny/crossref_metadata_2025 huggingface dataset:
Ultrastructural investigation of an adenoma of the pituitary post mortem
Authors: Siew Shirley , deMendonca W. C.
Published: June 2020

The deleterious effect of post mortem degeneration results in a progressive loss of ultrastructural detail. This had led to reluctance (if not refusal) to examine autopsy material by means of transmission electron microscopy. Nevertheless, Johannesen has drawn attention to the fact that a sufficient amount of significant features may be preserved in order to enable the establishment of a definitive diagnosis, even on “graveyard” tissue.

Routine histopathology of the autopsy organs of a woman of 78 showed the presence of a well circumscribed adenoma in the anterior lobe of the pituitary. The lesion came into close apposition to the pars intermedia. Its architecture was more compact and less vascular than that of the anterior lobe. However, there was some grouping of the cells in relation to blood vessels. The cells tended to be smaller, with a higher nucleocytoplasmic ratio. The cytoplasm showed a paucity of granules. In some of the cells, it was eosinophilic.
Visualization of the polymerization dynamics of individual 25nm-diameter microtubules in real time
Authors: Salmon E. D. , Walker R. A. , O'Brien E. T. , Pryer N. K. , Voter W. A. , Erickson H. P.
Published: June 2020

Microtubules (Mts) of the interphase cytoplasmic Mt complex (CMTC) and the mitotic spindle are dynamic polymers assembled from a cellular pool of tubulin by end-dependent association-dissociation reactions (1,2). Mts have an intrinsic structural polarity, which orients the direction of cell motility. Mts also play a major role in the active translocation of vesicles and chromosomes. Quantitative fluorescence microscopy studies using fluorescently labeled tubulin microinjected into living cells have shown that the great majority of Mts in the CMTC and the spindle exchange rapidly with tubulins in the cellular pool (2,3). Dynamic instability, as proposed by Mitchison and Kirschner, provides the best explanation for this rapid turnover of microtubules (1,2).

Three major temporal phases of Mt polymerization and three major abrupt transitions between these phases characterize the dynamic instability behavior of pure tubulin (Fig. 1).
3D biological imaging using TSRLM confocal microscopy
Authors: Loren Buhle E. , Himpens Bernard , Somlyo Avril V. , Shuman Henry , Somlyo Andrew P.
Published: June 2020

Tandem scanning reflected light (TSRLM) confocal microscopy permits the imaging of optical sections through translucent material in real time. We are currently using a TSRLM microscope to examine the morphological components of biological tissue. Optical sections of either stained or unstained biological specimens can be rapidly obtained by a through focus series of a potentially dynamic specimen. Each optical section shown here is an average of 256 512X512x8 bit frames collected from a silicon intensified target camera at video rates.

Unstained living frog muscle fibers were imaged with sufficient contrast to obtain 1μ optical sections traversing a total distance of approximately 80μ. Fig. 1 shows 4μ sections through a blood vessel. Frog red cells can be clearly seen in panels 1c-e. Frog muscle fibers were mounted and stretched to a defined length. The total fiber volume was easily imaged by progressive 1μ sections.
Removal of inelastically scattered electrons substantially increases phase contrast on frozen-hydrated molecules
Authors: Langmore John P. , Athey Brian D.
Published: June 2020

Although electron diffraction indicates better than 0.3nm preservation of biological structure in vitreous ice, the imaging of molecules in ice is limited by low contrast. Thus, low-dose images of frozen-hydrated molecules have significantly more noise than images of air-dried or negatively-stained molecules. We have addressed the question of the origins of this loss of contrast. One unavoidable effect is the reduction in scattering contrast between a molecule and the background. In effect, the difference in scattering power between a molecule and its background is 2-5 times less in a layer of ice than in vacuum or negative stain. A second, previously unrecognized, effect is the large, incoherent background of inelastic scattering from the ice. This background reduces both scattering and phase contrast by an additional factor of about 3, as shown in this paper. We have used energy filtration on the Zeiss EM902 in order to eliminate this second effect, and also increase scattering contrast in bright-field and dark-field.
Cryosectioning plant roots prepared by high-pressure freezing
Authors: Crang R.E. , Mueller M. , Zierold K.
Published: June 2020

Obtaining frozen-hydrated sections of plant tissues for electron microscopy and microanalysis has been considered difficult, if not impossible, due primarily to the considerable depth of effective freezing in the tissues which would be required. The greatest depth of vitreous freezing is generally considered to be only 15-20 μm in animal specimens. Plant cells are often much larger in diameter and, if several cells are required to be intact, ice crystal damage can be expected to be so severe as to prevent successful cryoultramicrotomy. The very nature of cell walls, intercellular air spaces, irregular topography, and large vacuoles often make it impractical to use immersion, metal-mirror, or jet freezing techniques for botanical material.

However, it has been proposed that high-pressure freezing (HPF) may offer an alternative to the more conventional freezing techniques, inasmuch as non-cryoprotected specimens may be frozen in a vitreous, or near-vitreous state, to a radial depth of at least 0.5 mm.
Microprobe analysis of inclusion bodies related to two benzofuran derivatives
Authors: Benning T. L. , Ingram P. , Shelburne J. D.
Published: June 2020

Two benzofuran derivatives, chlorpromazine and amiodarone, are known to produce inclusion bodies in human tissues. Prolonged high dose chlorpromazine therapy causes hyperpigmentation of the skin with electron-dense inclusion bodies present in dermal histiocytes and endothelial cells ultrastructurally. The nature of the deposits is not known although a drug-melanin complex has been hypothesized. Amiodarone may also cause cutaneous hyperpigmentation and lamellar lysosomal inclusion bodies have been demonstrated within the cells of multiple organ systems. These lamellar bodies are believed to be the product of an amiodarone-induced phospholipid storage disorder. We performed transmission electron microscopy (TEM) and energy dispersive x-ray microanalysis (EDXA) on tissue samples from patients treated with these drugs, attempting to detect the sulfur atom of chlorpromazine and the iodine atom of amiodarone within their respective inclusion bodies.

A skin biopsy from a patient with hyperpigmentation due to prolonged chlorpromazine therapy was fixed in 4% glutaraldehyde and processed without osmium tetroxide or en bloc uranyl acetate for Epon embedding.
Synaptic mechanisms in the light of structural insights: Quantal release in the central nervous system and its physical constraints
Authors: Korn H. , Triller A.
Published: June 2020

Morphological features of chemical synapses can help to understand how presynaptic neurons secrete transmitters and how the postsynaptic neurons respond to their action. In the case of the CNS, it has been known since the advent of the electron microscope that specialized sites for vesicular discharge exist in nerve terminals; such regions, where the neighboring membranes which constitute the synapse appear thicker, were named by Palay “synaptic complexes”. Gradually, evidence has accumulated (ref. in 2) that they correspond to the so-called “active zones” of the neuromuscular junction, but whether the nature of transmission is similar at their level has remained a matter of controversy, primarily because of technical difficulties in the analysis of responses produced in i) deeply located and ii) polyinnervated neurons of the brain. Recently, electrophysiological and ultrastructural data obtained at the teleost's Mauthner cell have confirmed that indeed both types of junctions constitute “morphological synaptic units” which share common rules of operation.
Ultrastructure of neurons and astrocytes in cerebral cortex of fasted rats subjected to partial global ischemia
Authors: Dimlich R.V.W. , Biros M.H.
Published: June 2020

Evidence exists that the morphoglogical expression of ischemic cell change in the central nervous system not only depends on the nutritional state of the animal, the adequacy of reperfusion and length of ischemia, but whether or not this ischemia was complete or incomplete. (1) In addition, changes that occur in neurons, associated glial cells, and adjacent neuropil are selective as well as progressive and need to be investigated by area and location within a specific region of the brain as well as over time. (2) In a model of incomplete ischemia in the rat, other investigators have examined cortical tissue after 5 and 90 min of recirculation following 30 min of ischemia. (3) As a preliminary investigation into the time course of ischemic cell damage in this model, we have chosen an intermediate time of 30 min of recirculation.
Regenerating myofibers in tibialis anterior muscles of young ‘MDX’ mice
Authors: Geissinge H. D. , Rhodes L.D.
Published: June 2020

A recently discovered mouse model (‘mdx’) for muscular dystrophy in man may be of considerable interest, since the disease in ‘mdx’ mice is inherited by the same mode of inheritance (X-linked) as the human Duchenne (DMD) muscular dystrophy. Unlike DMD, which results in a situation in which the continual muscle destruction cannot keep up with abortive regenerative attempts of the musculature, and the sufferers of the disease die early, the disease in ‘mdx’ mice appears to be transient, and the mice do not die as a result of it. In fact, it has been reported that the severely damaged Tibialis anterior (TA) muscles of ‘mdx’ mice seem to display exceptionally good regenerative powers at 4-6 weeks, so much so, that these muscles are able to regenerate spontaneously up to their previous levels of physiological activity.
Effect of passive exercise on endplate architecture in rat skeletal muscle following nerve crush injury
Authors: Pachter B.R. , Eberstein A.
Published: June 2020

In Rehabilitation Medicine, passive exercise or stretch is often utilized as a therapeutic modality to decrease fibrosis, retard development of contractures, as well as enhancing blood flow to the muscle. In addition, we have recently shown that this modality can also retard muscle fiber atrophy following denervation in rat extensor digitorum longus (EDL) muscle. Following a nerve crush injury, the distal nerve segment degenerates while the proximal one regenerates along the empty neural pathway to reinnervate the muscle. This process of muscle reinnervation is influenced by several factors which regulate various stages such as nerve regrowth, axonal sprouting, reinnervation, and maintenance. It has been suggested that passive exercise may also stimulate muscle reinnervation. The present study was designed with the primary objective to determine if reinnervation is enhanced following passive exercise treatment.
Scanning Electron Microscopy of the glomerulus in diabetic BB/S wistar rats
Authors: Kaufmann R. C. , Khosho F. K. , Amankwah K. S.
Published: June 2020

Renal damage secondary to diabetes seems to be related to the severity and duration of the diabetes. In streptozotocin and alloxan-induced diabetic rats, renal disease is found only in those rats that have glycosuria and then only after the glycosuria has been present for many months. In these animals, the longer they have glycosuria, the more severe the renal damage. In our colony of BB/S Wistar rats, animals that are going. to become frankly diabetic demonstrate clinical diabetes before they begin spilling glucose in their urine. After glycosuria develops, the condition of the animals worsens; yet, the glucose tolerance tests(GTT) remain essentially unchanged. The purpose of this investigation was to study the animals' kidneys to discover if lesions are present at the onset of glycosuria and how severe the lesions are.

Rats of our BB/S Wistar strain were used the day they developed glycosuria. Similarly aged non-diabetic animals were used as controls.
I. The morphology of interleukin-2 stimulated human peripheral blood mononuclear cells killing tumor Cellsin vitro
Authors: Hook G. R. , Greenwood M. A. , Barba D. , Ikejiri B. , Chen S. N. , Oldfield E. , Muul L. M.
Published: June 2020

Culture of peripheral blood mononuclear cells (PBM) with recombinant interleukin-2 (rIL-2) results in lymphokine activated killer cells (LAK) that can kill fresh non-cultured, natural killer cell resistant tumor cells. Clinically, human LAK and rIL-2 immunotherapy can cause regression of some tumors in humans and preliminary trials using LAK cells to treat glioma patients are beginning . LAK effector cells are a subset of the LAK population that must physically contact target cells in order to kill them. In this and the following abstract, we report the morphology of LAK cells killing human glioma derived cellsin vitro.

PBM cells were obtained from normal adult volunteers and stimulated in culture for 4 days with rIL-2 (Cetus, Emeryville, CA). Glioma-derived cells were obtained at surgery from a recurrent glioma patient and established as a cell line (SNB-92). SNB-92 cells were positive for glial fibrillary acidic protein and were used at passage 8.
II. The morphology of interleukin-2 stimulated human peripheral blood mononuclear cells killing tumor cells In Vitro
Authors: Greenwood M. A. , Hook G. R. , Barba D. , Ikejiri B. , Chen S. N. , Oldfield E. , Muul L. M.
Published: June 2020

In the previous abstract, we presented data on the cytolysis of glioma-derived cells (SNB-92) by lymphokine activated mononuclear cells (LAK). In this abstract, we describe our transmission electron microscopy (TEM) study of the interaction of LAK cells and SNB-92 in vitro during a time sequence of 5 minutes to 4 hours. Co-cultures were fixed in situ by the direct addition to the culture medium of 2.5% glutaraldehyde and prepared in a routine manner for TEM.

Within 30 minutes, some cells within the LAK population make contact with glioma cells (Fig 1). These LAK cells assume an elongated shape when they approach their target. The nucleus is asymmetrically located distal to the point of contact. The cellular organelles and microtubules orient towards the target cell. The plasma membranes of both cells bind closely together and form numerous interdigitations.
The structure of sickle hemoglobin fibers within fascicles
Authors: Carragher Bridget , Bluemke David A. , Becker Murray , McDade William , Potel Michael J. , Josephs Robert
Published: June 2020

The structure of HbS fibers has been analyzed from electron micrographs of negatively stained particles using three-dimensional reconstruction techniques. Fibers are composed of the same structural unit as crystals, namely the Wishner-Love double strand, but the arrangement of the double strands in fibers and crystals is quite different. This raises questions regarding the manner in which such structures might fuse to form crystals. The crystallization must be accompanied by a considerable rearrangement of the double strands and the fibers, which have net polarity, must combine in such a way as to form a non-polar crystal. It is of interest in this regard to consider whether the structure of fibers associated into ordered bundles, or fascicles, differs from the structure of individual fibers. The structure and interaction of fibers within large arrays may have clinical significance as it is the formation of bundles of fibers within the red blood cell which leads to the clinical manifestations of sickle cell disease.
Improvement of an artificial test substrate technique for evaluation of em immunocytochemical labeling
Authors: Gagne G. D. , Miller M. F.
Published: June 2020

We recently described an artificial substrate system which could be used to optimize labeling parameters in EM immunocytochemistry (ICC). The system utilizes blocks of glutaraldehyde polymerized bovine serum albumin (BSA) into which an antigen is incorporated by a soaking procedure. The resulting antigen impregnated blocks can then be fixed and embedded as if they are pieces of tissue and the effects of fixation, embedding and other parameters on the ability of incorporated antigen to be immunocyto-chemically labeled can then be assessed. In developing this system further, we discovered that the BSA substrate can also be dried and then sectioned for immunolabeling with or without prior chemical fixation and without exposing the antigen to embedding reagents. The effects of fixation and embedding protocols can thus be evaluated separately.
IGM antibodies can be used as direct immunoelectron markers
Authors: Keene Douglas R. , Sakai Lynn Y. , Burgeson Robert E. , Bächinger Hans Peter
Published: June 2020

A mouse monoclonal IgM antibody has been used in this study as a direct ultrastructural marker to immunolocalize human Type III collagen at the level of the transmission and scanning electron microscopes. The method utilized involves the use of none of the electron dense conjugates such as colloidal gold, ferritin, or peroxidase: Hence ultrastructural immuno- localization relies exclusively on the size and electron density of the bound IgM complex after routine staining. As compared to similar immuno- localization studies utilizing the same monoclonal anti-Type III collagen IgM but with additional secondary antibody-gold conjugates, the direct “naked” IgM technique consistently gave results of higher specificity and greater sensitivity.
Ultrastructure of rat alveolar macrophages activated in situ by poly:IC
Authors: Klinkner A. M. , Weiss R. A. , Kelley A. , Bugelski P. J.
Published: June 2020

Polyinosinic:polycytidylic acid is an inducer of interferon and a macrophage activator. We have found that intratracheal instillation of polyI:C (IT-pI:C) activates rat bronchoalveolar lavage cells (BAL) for a variety of functions. Examination of Giemsa stained, cytocentrifuge preparations showed that IT-pI:C induced a population of BAL not seen in resident BAL. The morphology of these cells suggested that they might be derived from blood monocytes. To test this hypothesis we have examined several populations of macrophages that had been stained for endogenous peroxidase activity as a marker of cells derived from the monocyte-macrophage lineage.

Macrophages were obtained from Fischer 344 rats. Peritoneal exudate cells (PEC) were collected by lavage 4 days after i.p. injection of 20 ml 3% thioglycolate. Buffy coat monocytes were separated from venous blood from naive rats.
Immunogold labeling of protein kinase in muscle cells
Authors: Philpott D. E. , Hand A. , Mednieks M. , Kato K. , Stevenson J.
Published: June 2020

Muscle loss during spaceflight remains a medical concern in prolonged space travel. We have studied changes in cyclic AMP-dependent protein kinase (ATP: protein phosphotransferase, EC 2.7.1.37; cAPK) levels in heart muscle after simulated weightlessness and after actual microgravity on Space Lab-3. Biochemical measurements were made in the soluble and the particulate fraction of disrupted muscle tissue, but did not identify the exact subcellular location of this enzyme. In order to establish localization of the protein kinase in the muscle cells of the rat, monoclonal antibodies to the regulatory (R) subunits of the type II isoenzyme were prepared and employed in a protein A-gold immunolabeling procedure.

Both rat striated and heart muscle were fixed in 2.5% glutaraldehyde (GA), cacodylate buffer, for 1 hour. Similar samples were fixed in Triple Fix (T.F.) in cacodylate buffer for one week. Osmium was not used.
Immunoelectron microscopic localization of endoglucanase in the frozen thin sections of a hypercellulolytic mutant of Trichoderma reesei
Authors: Ganguly Tapan , Ghosh Bijan K.
Published: June 2020

Endoglucanase (EG) is an enzyme of cellulase complex depolymerizing crystalline cellulose to cellulose microfibril fragments. T. reesei Rut-C30 is a catabolite repression resistant mutant which enhances endoplasmic reticulum (ER) and produces 15-20 times more endoglycanase than the wild type QM6a. About 90% of this enzyme is secreted.

Purified secreted EG from three day old culture was used for polyclonal antibody production. The IgG was labeled with 15-18nm colloidal gold. EG secreting mycellia, grown in Vogel's medium with 1% Avicel PH101, were collected by centrifugation. This pellate was washed in sodium cacodylate buffer, fixed in tannic acid-glutaraldehyde mixture and cryoprotected with 30% glycerol. This material was placed on the specimen holders and frozen in liquid freon followed by liquid nitrogen. Frozen thin sections were prepared in a LKB Cryonova microtome using glass knives and collected on sucrose drops.
Disruption of collagen crosslinking during sea urchin morphogenesis
Authors: Hardin Jeffrey D.
Published: June 2020

The extracellular matrix (ECM) is important for many events during embryonic development. An important component of the ECM in sea urchin embryos is collagen. Collagen has been identified in the basal lamina of sea urchin embryos by morphological and immunological criteria. Recently, the need for a crosslinked collagenous matrix in the basal lamina during sea urchin embryogenesis has been demonstrated using β–aminoproprionitrile (BAPN). BAPN is a specific inhibitor of lysyl oxidase, an enzyme involved in collagen crosslinking. BAPN treatment causes completely reversible developmental arrest at the early gastrula stage, or supression of mesenchyme cell motility and archenteron elongation, depending on the time of addition of the drug. BAPN also significantly decreases the extent of collagen crosslinking, based on solubility assays. This study examines fine structural changes in cellular morphology and motility and the three-dimensional structure of the ECM induced by BAPN treatment during gastrulation and larva formation in the sea urchin embryo.
What's where in intermediate filaments?
Authors: Steven A.C. , Trus B.L. , Steinert P.M.
Published: June 2020

Of the major cytoskeletal proteins of eukaryotic cells, intermediate filament (IF) proteins form a uniquely diverse family whose molecular weights range from ∼44kDa for the smallest keratin subunit to ∼140kDa for the largest neurofilament subunit. From the recent deluge of amino-acid sequence information (e.g. 1-3), it has become clear that this variability is confined to the two terminal regions of each sequence: they flank a partially homologous domain, the so-called ‘rod’ domain, that is of uniform size (∼311 residues) and predicted secondary structure (a segmented α-helix). The expected α-helicity is supported experimentally by circular dichroism of IF molecules in solution, and by high-angle X-ray fiber diffraction from intact IF (e.g. 5-7). IF molecules are all dimers bonded through an inter-chain coiled-coil formed by the two rod domains. Of the end-domains of IF subunits, on the other hand, relatively little is known.
Ultrastructure of the epithelium of the upper ejaculatory duct of the mealworm beetle, Tenebrio molitor
Authors: Bricker S. , Happ G. M.
Published: June 2020

The male mealworm, Tenebrio molitor produces a spermatophore to facilitate transfer of sperm to the female. The wall of the spermatophore is largely produced from the secretions of the paired bean-shaped accessory glands (BAGs). As the cottony pre-spermatophoric mass from the BAGs comes together in the ejaculatory duct where it is molded into the spermatophore, it becomes tougher and more elastic. The mechanisms involved in this stabilization of the wall of the spermatophore were unknown. Mechanisms of stabilization of other acellular structures assembled in extracellular space include quinone-tanning and β-sclerotization in cuticle, shear forces in silk, and pH changes in the spermatophore of Rhodnius. The cells found in the epithelium of the upper ejaculatory duct of the mealworm beetle were examined by transmission electron microscopy for ultrastructural evidence of a role in the stabilization of the spermatophore wall.
Vesicular system associated with spermiogenesis of bullfrog
Authors: Liu K. C. , Tsay S. F.
Published: June 2020

In the histologic and electron microscopic study of the male reproductive system of bullfrog, Rana catesbeiana, a vesicular system associated with spermiogenesis was observed. It appeared in the lumenal space of the seminiferous tubule (Fig. 1), in the heads of spermatids (Fig. 2), associated with the chromatins of the spermatid (Fig. 4). As deduced from sections, this vesicular system consisted of vesicles of various size or a large group of waving and twisted tubules (Fig. 3), After routine procedure of treatment for electron microscopy, the lumens of both of the vesicles and tubules were electron lucent.

In human, vesicles and vesicular system associated with reproductive cell and tissue were reported. In abnormal spermiogenesis, flower-like body, actually vesicles, and giant vesicle associated with the head of spermatid were observed. In both cases the number of vesicle was limited from a single one to a few.
Growth factors and the primary cilium in BALB/c 3T3 cells
Authors: Tucker R. W. , More N. S. , Jayaraman S.
Published: June 2020

The mechanisms by which polypeptide growth factors Induce DNA synthesis in cultured cells is not understood, but morphological changes Induced by growth factors have been used as clues to Intracellular messengers responsible for growth stimulation. One such morphological change has been the transient disappearance of the primary cilium, a “9 + 0” cilium formed by the perinuclear centriole in interphase cells. Since calcium ionophore A23187 also produced both mitogenesis and ciliary changes, microtubule depolymerization might explain ciliary disappearance monitored by indirect immunofluorescence with anti-tubulin antibody. However, complete resorption and subsequent reformation of the primary cilium occurs at mitosis, and might also account for ciliary disappearance induced by growth factors. To settle this issue, we investigated the ultrastructure of the primary cilium using serial thin-section electron microscopy of quiescent BALB/c 3T3 cells before and after stimulation with serum.
Iron particles isolated from the dura of pygmy sperm whalesKogia breviceps
Authors: Credle V. R.
Published: June 2020

Magnetite, a highly magnetic iron oxide, is a biomineralization product found in organisms sensitive to the earth's magnetic field. Biogenically produced magnetite differs from magnetite-bearing rocks of geological origin in crystalline morphology, size, and elemental composition. Standard paleomagnetic methods indicate that cetacean brain dura may also contain magnetite. Correllations have been documented between the distribution of cetacean live stranding events and anomalously minimal magnetic field intensities. This would suggest magnetoreceptive abilities in whales. Pygmy sperm whales are one of the most common cetaceans to strand alive in Florida. Therefore, it is of interest to determine whether pygmy sperm whale brain contains magnetite, as this will establish sensitivity to variations in the earth's magnetic field. Reported here are the results of isolation experiments to ascertain whether iron-rich particles are present in pygmy sperm whales,Kogia breviceps.
Ultrastructure of the malpighian tubules in adult female Dermacentor variabilis
Authors: Coons Lewis B. , Rosell-Davis Rosemarie , Rose Lisa , Lamoreaux William
Published: June 2020

The excretory system of the American dog tick Dermacentor variabilis (Say) consists of a pair of Malpighian tubules that end in a blind ampulla and insert into the rectal sac which in turn leads to the anus. Visceral muscle envelopes the tubules. The single layer of epithelial cells in the tubule wall have an ultrastructure characteristic of cells that transport fluids and ions, e.g. an infolded basal plasma membrane, relatively large numbers of mitochondria, and an extensive development of microvilli on the apical luminal surface. Ixodid ticks have a cycle of excretion during their life span as an adult. Most excretory activity occurs during and after feeding. The purpose of this study was to describe the ultrastructure of the Malpighian tubules in an unfed tick, a fed tick and an ovipositing tick, in an attempt to correlate ultrastructure with excretory activity. We subdivided the tubules into distal, middle and proximal regions in which each area was about 1/3 of a tubule. The proximal region was closest to the rectal sac.
Intracisternal microtubules in proximal tubule cells of duck kidney: A serendipitous finding
Authors: Bhatnagar M.K. , Snelgrove-Hobson S.M. , Prasada Rao P.V.V.
Published: June 2020

Cytpplasmic microtubules, ubiquitous cell organelles, lie free in the cytosol without being compartmented off by membranes. In rare instances, however, intracisternal microtubules (ICM) have been reported in ganglionic neurons of aging dogs and in the proximal tubule cells (PTC) of rabbit kidneys. We report here the presence of ICM in the PTC of the kidneys of Peking ducks (Anas platyrhynhos).

A total of 106 Peking ducks (24 males and 24 females of 9 months, 48 females of 15 months, 6 males of 30 months and 2 males and 2 females of 48 months of age) were anesthetized and exsanguinated. Pieces of kidney cortex were fixed in 4% glutaraldehyde in 0.5M phosphate buffer (pH 7.3) at 29°C for four hours. Samples were post fixed in 2% osmium tetroxide for two hours. One micron sections of Epon-embedded cortices were stained with toluidine blue and thin sections (70-70 nm) contrasted with uranyl acetate and lead citrate were examined with a JEOL 100-S electron microscope at 80 kV.
Deep etching of rapidly frozen cytoskeletons of motile fibroblasts
Authors: Heath Julian P. , Bard Frédérique
Published: June 2020

The cytoskeleton of motile cultured cells is a complex three dimensional network of actin microfilaments, microtubules and intermediate filaments. We are using rapid freezing and deep etching to visualise this network in human and chick fibroblasts after detergent extraction. This method offers high resolution, high contrast images that retain three dimensionality with minimal artefact, and gives views of the cytoskeleton that differ from those obtained with more conventional techniques such as HVEM of thick plastic sections, or CTEM of critical point dried whole mounts.

Human skin fibroblasts and chick embryo heart fibroblasts were grown on No. 1 glass coverslips and fixed and extracted with 0.1 M PIPES buffer pH 7 containing 0.25% glutaraldehyde, 0.5% Triton X-100, 1 mM MgCl2, 1 mM EGTA, 1 μg/ml phalloidin and 5 μg/ml taxol, for 30 s to 10 min at RT; further fixed in 2.5% glutaraldehyde in PIPES for 10 min, postfixed in 1% osmium in PIPES for 5 min and dehydrated in 70% ethanol.
Detection of viruses by electron microscopy: Increased sensitivity by direct micro-ultracentrifugation of samples
Authors: Trudel Alain R. , Trudel M.
Published: June 2020

AirfugeR (Beckman) direct ultracentrifugation of viral samples on electron microscopy grids offers a rapid way to concentrate viral particles or subunits and facilitate their detection and study. Using the A-100 fixed angle rotor (30°) with a K factor of 19 at maximum speed (95 000 rpm), samples up to 240 μl can be prepared for electron microscopy observation in a few minutes: observation time is decreased and structural details are highlighted. Using latex spheres to calculate the increase in sensitivity compared to the inverted drop procedure, we obtained a 10 to 40 fold increase in sensitivity depending on the size of particles. This technique also permits quantification of viral particles in samples if an aliquot is mixed with latex spheres of known concentration.

Direct ultracentrifugation for electron microscopy can be performed on laboratory samples such as gradient or column fractions, infected cell supernatant, or on clinical samples such as urine, tears, cephalo-rachidian liquid, etc..
Protein-induced conformational changes in 16S rRNA during assembly of the Escherichia Coli 30S ribosomal subunit: A STEM study
Authors: Boublik M. , Mandiyan V. , Hainfeld J.F. , Wall J.S.
Published: June 2020

The aim of this study is to understand the mechanism of 16S rRNA folding into the compact structure of the small 30S subunit of E. coli ribosome. The assembly of the 30S E. coli ribosomal subunit is a sequence of specific interactions of 16S rRNA with 21 ribosomal proteins (S1-S21). Using dedicated high resolution STEM we have monitored structural changes induced in 16S rRNA by the proteins S4, S8, S15 and S20 which are involved in the initial steps of 30S subunit assembly. S4 is the first protein to bind directly and stoichiometrically to 16S rRNA. Direct binding also occurs individually between 16S RNA and S8 and S15. However, binding of S20 requires the presence of S4 and S8. The RNA-protein complexes are prepared by the standard reconstitution procedure, dialyzed against 60 mM KCl, 2 mM Mg(OAc)2, 10 mM-Hepes-KOH pH 7.5 (Buffer A), freeze-dried and observed unstained in dark field at -160°.
Scanning Electron Microscopy of the vagina in diabetic BB/S Wistar rats
Authors: Khosho F. K. , Kaufmann R. C. , Amankwah K. S.
Published: June 2020

The influence of diabetes on reproductive organs in humans and various other mammalian species, including the laboratory rat, is not well documented. We have previously shown that 13% of our BB/S Wistar rat colony demonstrated persistent estrous on vaginal smears. Spontaneously-diabetic BB Wistar rats of our colony demonstrate persistent diestrous on vaginal smears long before showing glucose in urine. In this report, we describe our findings on the ultrastructure of the vaginal epithelium of the frankly diabetic(FD) rat. The rats which started our colony were obtained from the Animal Resource Division of the Government of Canada.

Scanning electron microscopy was used to examine specimens of vaginal surface epithelium with comparisons being made with specimens from nondiabetic controls. Cardiovascular perfusion with a buffered glutaraldehyde solution was used forin vivofixation of the cervix and vagina.
Modified gach technique vs critical point drying: Shrinkage analysis for SEM
Authors: Thompson B. , Sculov N. , Crang R.E.
Published: June 2020

The use of co-polymerized glutaraldehyde-carbohydrazide (GACH) was proposed for specimen preparation in scanning electron microscopy (SEM) as a means of avoiding dehydration in organic solvents, and to provide dimensionally stable biological specimens through a process of air-drying. It has been assumed that shrinkage of specimens prepared by the GACH technique should be less than that of conventionally-prepared material by critical point drying (CPD). In a previous study, Bell has reported significant shrinkage of whole cells for SEM. This report compares cell shrinkage in GACH and CPD preparations.

Fibroblasts from newborn rats were grown on collagen-coated glass cover-slips (with alpha numeric grids etched onto the surface of the coverslips) in Eagle's minimum essential medium + 10% fetal calf serum for 7 d. (3). Using an inverted microscope with phase-contrast optics, micrographs were taken of the cultures in their live state and 1 h. after fixation with 2.5% glutaraldehyde in Dulbecco's phosphate buffered saline (Figs. 1 and 3).
Relationship between various types of inclusions in the broncho-alveolar space of axolotl lung
Authors: Keyhani E.
Published: June 2020

The surfactant system of the mexican axolotl Ambystoma mexicanum exhibited the same basic structures as those described for mammalian surfactant. Here we report subdivision of the surfactant inclusions, their interconversion and their frequency in the lung of the axolotl.

1)

Type I inclusion or tubular myelin represented 8% of the total inclusions. It consisted of square to rectangular grid patterns (Figs. 1, 4). Each square or rectangle shared its walls with four neighboring units. The lattice appeared to be composed of membranous elements of 6 nm, crossing in two directions perpendicular to each other. A centrally located dense line of 2 nm diameter with a fuzzy contour crossed the middle section of each rectangle in its long axis (Fig. 4). Each rectangle measured about 17 x 27 nm for internal perimeter and 27 x 36 nm for center to center spacing of its membrane element. The average size of type I inclusions was about 0.8 μm.
Immunoelectron microscopy in diagnostic pathology: The myths and realities of applicability and methodology
Authors: Lewis Jon C.
Published: June 2020

Immunohistochemical techniques since introduction in the 1940's have held promise as ancillary methods in situations when confident diagnostic decisions could not be made using standardized histologic or histochemical methods. Recently as a result of monoclonal antibody technology and availability of reliable commercial reagents, the potential for immunohistochemistry as a diagnostic aid has been realized. Although immunologic techniques are applicable to a variety of pathological questions, particularly those related to pathogenesis and etiology, from a practical standpoint widespread use has not occurred, and diagnostic use primarily has been in the area of tumor pathology for differentiation of malignant from non-malignant tissues and for identifying antigenic differences between cancers of similar histology. At the ultrastructural level, immunocytochemistry has also received attention for use in pathology; but, as in the case of histologic use, diagnostic application at the EM level has been restricted to select well defined situations with virus identification and characterization of poorly differentiated leukemias being the major areas.
Comparative strengths and weaknesses of peroxidase and colloidal gold in pre- and post-embedding immunolabeling techniques
Authors: Abrahamson D. R. , St.John P. L. , Perry E. W.
Published: June 2020

Antibodies coupled to tracers for electron microscopy have been instrumental in the ultrastructural localization of antigens within cells and tissues. Among the most popular tracers are horseradish peroxidase (HRP), an enzyme that yields an osmiophilic reaction product, and colloidal gold, an electron dense suspension of particles. Some advantages of IgG-HRP conjugates are that they are readily synthesized, relatively small, and the immunolabeling obtained in a given experiment can be evaluated in the light microscope. In contrast, colloidal gold conjugates are available in different size ranges and multiple labeling as well as quantitative studies can therefore be undertaken through particle counting. On the other hand, gold conjugates are generally larger than those of HRP but usually can not be visualized with light microscopy. Concern has been raised, however, that HRP reaction product, which is exquisitely sensitive when generated properly, may in some cases distribute to sites distant from the original binding of the conjugate and therefore result in spurious antigen localization.
Backscattered electron signals of coated specimens at low and high detector take-off angles
Authors: Hejna Jan
Published: June 2020

Backscattered electron (BSE) signals of differently coated specimens are studied at two ranges of BSE take-off angles. Specimens are coated with one layer of a high atomic number (high Z) material or with two layers: first of a high Z material and second of a low Z material. Experiments are carried out in a scanning electron microscope equipped with a tungsten filament.

Fig. 1 shows an arrangement of detectors. A ring detector subtending angles from 0° to 20° in respect to the specimen surface and a disc detector subtending angles from 60° to 80° are used. Detectors contain plastic scintillators (102A, NE Technology). The first specimen consists of an Au layer on a Cu substrate and a part of it is covered with an additional Cu layer. Fig. 2 shows a dependence of BSE signals on the thickness of the Au layer and Fig. 3 on the thickness of the Cu layer.
Acoustic microscopy techniques for the inspection of integrated circuit devices and packages
Authors: Moore Thomas M.
Published: June 2020

In the last decade, a variety of characterization techniques based on acoustic phenomena have come into widespread use. Characteristics of matter waves such as their ability to penetrate optically opaque solids and produce image contrast based on acoustic impedance differences have made these techniques attractive to semiconductor and integrated circuit (IC) packaging researchers.

These techniques can be divided into two groups. The first group includes techniques primarily applied to IC package inspection which take advantage of the ability of ultrasound to penetrate deeply and nondestructively through optically opaque solids. C-mode Acoustic Microscopy (C-AM) is a recently developed hybrid technique which combines the narrow-band pulse-echo piezotransducers of conventional C-scan recording with the precision scanning and sophisticated signal analysis capabilities normally associated with the high frequency Scanning Acoustic Microscope (SAM). A single piezotransducer is scanned over the sample and both transmits acoustic pulses into the sample and receives acoustic echo signals from the sample.
Thin-section photomicrography
Authors: Holm Jack , Goss Rachel
Published: June 2020

A fundamental problem in conventional light microscopy has long been the lack of image quality brought about by the use of sections which are thicker than the depth of field (DOF) of the microscope. These sections are used because “paraffin & steel” microtomy techniques preclude sections thinner than a few microns. The transmission electron microscope, however, requires section thicknesses of less than a micron. This requirement has resulted in the development of “epoxy & glass” microtomy techniques, and the ability to cut sections as thin as a few hundred Ångstroms. These thin sections are not routinely used for light microscopy because of the difficulty of preparation, the need for large section areas, and because of photomicrographic problems. The first two reasons may inhibit the use of thin sections in some situations, but the photomicrographic problems are surmountable.

There is some disagreement in the literature regarding the actual DOF in photomicrographic situations.
New computer-controlled dual stage SEM—model DS-700
Authors: Ueda T. , Nakatsuka H. , Kimura T. , Buchanan R.
Published: June 2020

The dual stage electron-optical column design of the DS-700 provides a unique combination of high resolution and flexible sample handling. The top stage accomodates a sample between the polepieces of a low aberration immersion objective lens, while the bottom stage accepts large samples below a conical second objective lens. This column configuration has been described elsewhere, and is therefore not the subject of this paper.

A completely new digital electronic system with computer control of all operating parameters has been developed. A combined keyboard and menue control system has been adopted. Commonly used functions are assigned to a simple operating panel, with discrete functions which may be set and easily changed. Secondary functions are addressed using a menue display. In this manner, rapid simple operation is realized, while still providing extreme flexibility.

A single high quality TV monitor displays real time and processed images. It is also used to display menues, various operating parameters, and vacuum system status.
The CM10-bioTWIN: a new CM microscope optimised for high contrast and analytical performance
Authors: de Jong Marc J.C. , Busing Wim M. , Otten Max T.
Published: June 2020

In order to improve our understanding of life processes, morphological aspects of a specimen must be related to its chemistry. More and more morphological observations are combined with biochemistry, immunocytochemistry and analysis, demanding a different approach to specimen preparation than the one employed for morphological observation. In the case of immunocytochemistry the fixation is mild, while for X-ray analysis vapour fixation must be used to avoid loss of water-soluble substances. Heavy-element stains such as lead citrate or uranyl acetate normally cannot be used without introducing undesired and serious artefacts. The special specimen preparation procedures result in low TEM contrast, making it difficult to observe the morphology and hamper focussing especially if low beam intensities must be used as for beam-sensitive materials.

Analytical TEMs typically are optimised for materials science applications, where low contrast is rarely a problem. These microscopes have low objective-lens aberrations and short focal lengths, giving low point resolution and nm-sized smallest spot sizes but at a expense of image contrast.
Correlation of cryo-electron microscopic and x-ray data and compensation of the contrast transfer function
Authors: Cheng R. Holland
Published: June 2020

Cryo-electron microscopy (cryoEM) along with image reconstruction techniques can produce vivid images of biological macromolecules in their “native” state, although objective interpretation of these images is influenced by the fact that the contribution of phase contrast greatly exceeds that of amplitude contrast in such weakly scattering objects. The microscope contrast transfer function (CTF), which is strongly dependent on the defocus level of objective lens, modulates images of the object density distribution as a function of spatial frequency. Compensation for the effects of phase contrast transfer is important because underweighting of the low spatial frequencies usually causes difficulties in evaluating absolute mass distributions in objects.

Correct compensation for the CTF is difficult to achieve. This is due, in part, to ambiguities in measuring the exact defocus level in noisy micrographs, and in knowing the relative contributions of amplitude and phase contrast, beam coherence, and inelastic scattering. The availability of atomic resolution determinations for a few viruses allows one to determine empirically how to correct the cryoEM images to best fit the x-ray data.
Correlation averaging of ordered arrays of Photosystem II
Authors: Furcinitti Paul S. , Marr Kathleen M. , Lyon Mary K.
Published: June 2020

Photosystem II (PS II) is one of two reaction centers found in the photosynthetic membranes of eukaryotic cells. PS II converts solar to chemical energy and can split water, giving off H+ and O2. Neither bacterial reaction centers nor PS I can evolve O2, making PS II unique. Oxygen evolution involves three polypeptides (17, 24 and 33 kDa) that are exposed on the inner surface of the membrane. The remainder of PS II consists of intrinsic membrane proteins with molecular masses of 47, 43, 34, 33 and 10 kDa and multiple low molecular mass polypeptides (for review, see Hansson and Wydrzynski, 1990). In order to obtain structural information regarding PS II, and particularly the oxygen evolving-polypeptides (OEPs), we have induced the complexes to form two-dimensional crystals. The crystals are formed by sequential treatment of whole membranes with low concentrations of Triton X-100. The two-dimensional crystals are tubular, with the complexes arranged in a helical manner around the tube (Fig. 1). The tubes are being characterized by gel electrophoresis and immunoblotting, using antibodies directed against the OEPs.