bluuebunny/crossref_metadata_2025 huggingface dataset:
Platelet Adhesion to Native and Collagen Coupled Cuprophane: A Multispecies Comparative SEM Study
Authors: Lewis J.C. , Didisheim P. , Grabowski E.F. , Zollman P.E.
Published: June 2020

Platelet adhesion to intact endothelium seldom occurs, but platelet response to subendothelial structures has been extensively studied and is recognized as being focal to hemostasis and thrombosis. Platelet adhesion to subendothelium in-vivo is generally followed by platelet-platelet cohesion (aggregation) and the platelet release reaction (1). Since thrombus formation and the platelet release reaction have been implicated in several secondary pathologic responses, precise characterization of these events is essential to an understanding of many disease processes.

Previously we have demonstrated through the use of an in-vitro flow system that differences in platelet response to foreign surfaces exist among commonly studied animal species (2,3). The present report is an extension of these earlier studies and provides, through scanning microscopy, morphologic evidence for species related platelet responses.

Blood for these studies was obtained by venipuncture from sheep, calves, baboons, dogs, rabbits and human volunteers following informed consent.
Ultrastructural Study of the Cecal Epithelium in Normal Guinea Pig
Authors: Mora-Galindo Juan , Martínez-Palomo Adolfo
Published: June 2020

The use of guinea pig cecum in experimental amebiasis and other parasitic and microbial diseases, makes imperative to have a clear knowledge of its normal ultrastructure in order to evaluate properly the effects that the microorganisms may produce. Although several reports have appeared in the literature concerning the normal ultrastructure of the small and large intestines in various animal species including the human, it is difficult to extrapolate those descriptions to all animal species. The aim of the present work was to describe the ultrastructural characteristics of the normal guinea pig cecal mucosa, by means of transmission electron microscopy (TEM), scanning electron micro scopy (SEM), and freeze-fracture technics. Representative fragments of cecum from normal anesthetized guinea pigs were obtained through laparotomy and immediately fixed in 1% buffered osmium tetroxide and processed for TEM. Some samples were fixed in a 2.5% buffered glutaraldehyde solution and processed for SEM and freeze-fracture.
Scanning and Transmission Electron Microscopy of Endomentrium in Women Wearing TCu-380-A IUD 's
Authors: Gonzalez Angulo A. , Berlioz R. , Aznar R.
Published: June 2020

Recent ultrastructural studies on endometrial tissues from women wearing copper, wire intrauterine devices have disclosed morphological evidence of impaired glycogen degradation and secretion resulting in interference with the viability of blastocysts. Reduced microapocrine secretion observed with the scanning electron microscope supports this (1). In addition, organelle modifications have been observed in the epithelial cells of these women. The changes are seen in biopsies taken in the proliferative phase of the cycle and consist of mitochondrial vacuolation and myelin figure formation. These modifications disappear in the secretory phase and therefore have been regarded as reversible (2).

The aim of the present studies was to investigate surface epithelial changes as well as organelle modifications in relation to the site of contact with an IUD that releases greater amounts of copper. Endometrial tissue was obtained from the uterine cavity of four young women wearing TCu-380-A intrauterine contraceptive devices for 4-6 weeks.
A minimal does technique in 3d-electron microscopy
Authors: Knauer V. , Schramm H.J. , Hoppe W.
Published: June 2020

As discussed elsewhere a minimal dose technique for the exposure of a 3Dtilt series is very important. This can be achieved by choosing one area of the specimen for tilting and focussing and another for exposure. For 2D-microscopy this principle was already used by Williams and Fisher. We avoid mechanical movements by shifting the illumination spot by a magnetic deflection system in the microscope, and by simultaneously bringing back the image by an electrostatic field, generated by additional built-in deflection plates (which allow shifts up to 50μ). Electromagnetic shifts of the illumination spot (but without the synchronous shifts of the image) have already been applied for 2D-microscopy. Our method has the advantage that large shifts (which are independent of the magnification) can be used. In practice the shifts are of the order of 10μ. The deflection is orientated along the tilt axis of the microgoniometer.
Digital image enhancement for transmission electron microscopy
Authors: Goldfarb William , Frank Joachim
Published: June 2020

The term image enhancement refers to those techniques which make pictures more interpretable. This definition implies that the best choice between enhancement processing procedures is closely tied to the subject matter and application of the images. We report here on the use in transmission electron microscopy of some image processing methods that we well know from their use in other fields of science and engineering. These methods can be divided into the two general categories of point mappings and spatial processing.

In a point mapping process, density values of individual picture elements are transformed into new values by a function which is independent of the values of neighboring points. The most common motivation for point mapping is to use most effectively the available gray shades of the display device. Usually the density histogram of the image to be displayed is used to generate parameters for the mapping function.
Lead aspartate: a new en bloc stain for electron microscopy
Authors: Walton J.R.
Published: June 2020

In electron microscopy, lead is the metal most widely used for enhancing specimen contrast. Lead citrate requires a pH of 12 to stain thin sections of epoxy-embedded material rapidly and intensively. However, this high alkalinity tends to leach out enzyme reaction products, making lead citrate unsuitable for many cytochemical studies. Substitution of the chelator aspartate for citrate allows staining to be carried out at pH 6 or 7 without apparent effect on cytochemical products. Moreover, due to the low, controlled level of free lead ions, contamination-free staining can be carried out en bloc, prior to dehydration and embedding. En bloc use of lead aspartate permits the grid-staining step to be bypassed, allowing samples to be examined immediately after thin-sectioning.

Procedures. To prevent precipitation of lead salts, double- or glass-distilled H20 used in the stain and rinses should be boiled to drive off carbon dioxide and glassware should be carefully rinsed to remove any persisting traces of calcium ion.
Ultrastructural localization of drug binding sites
Authors: Hoff S.F. , Macinnis A.J.
Published: June 2020

The need for a high resolution morphological method of identifying drug binding sites has been mentioned by several authors. Electron microscopic autoradiography may identify statisically an organelle as a drug binding site, but in practice has proven unreliable. This is primarily due to the tissue preparatory procedures used in conventional electron microscopy, which denature proteins and extract many proteins and lipid components from the tissue. As the drug binding sites are denatured and/or extracted, drugs are displaced from their “receptors.” Then the drugs are either extracted or attached to an anomolous binding site created by the preparatory procedure. Sjöstrand's minimal denaturation technique is based on a sound physical-chemical rationale and was modified slightly to accommodate the drug localization procedure used in this study.

It was found after many experiments that phosphotungstic acid (PTA) at pH7 had the highest specificity for the drugs used in this study and the lowest affinity for biological structures.
OsO4-tannin-OsO4 method for transmission and scanning electron microscopy of biological specimens
Authors: Takahashi Gen
Published: June 2020

Tissue blocks or slices about 1 mm in thickness were treated according to the following procedures both for transmission electron microscopy(SEM) and for scanning electron microscopy(SEM).

[A] Primary Osmication(Fixation): 2%0s04 in 0.1 M Na-cacodylate buffer or Millo- nig's phosphate buffer(pH7.4) for 2 hrs at 4°C.

[B] Postfixation and Mordanting: 2-4 % tannic acid—8(2-16)%glutaraldehyde in the buffer(pH6.8-7.0),for 1-40 hrs at 4°C.

[C] Secondary Osmication(Staining): 2 %0s04 in the buffer(pH7.4)for 2 hrs at 4°C. Specimens after step C were rapidly dehydrated and embedded in Epon. Ultrathin sections were cut with diamond knives. Although sections without heavy metal staining could be observed with moderate contrast, they were stained briefly with lead solution for high magnification, because of sublimation of osmium by intense electron bombardment.
Ultrathin frozen sections of yeast without aldehyde prefixation
Authors: Böhm K. J. , Unger a. E.
Published: June 2020

During the last years it was shown that also by means of cryo-ultra-microtomy a good preservation of substructural details of biological material was possible. However the specimen generally was prefixed in these cases with aldehydes.

Preparing ultrathin frozen sections of chemically non-prefixed material commonly was linked up to considerable technical and manual expense and the results were not always satisfying. Furthermore, it seems to be impossible to carry out cytochemical investigations by means of treating sections of unfixed biological material with aqueous solutions.

We therefore tried to overcome these difficulties by preparing yeast cells (S. cerevisiae) in the following manner:
Preparation, transfer and TEM observations of frozen hydrated muscle sections
Authors: Valdrè Ugo , Sjöström Michael , Edman Anne-Christine
Published: June 2020

Introduction

Ideally, biological specimens should be studied in their native state. Frozen hydrated sections of tissues, which are unfixed, unstained and unembedded, are closer to this state than material prepared with conventional techniques. Very recently some efforts have been devoted to the practical problems related to this new approach. The present work describes the methodology we have adopted, the techniques developed and some of the results obtained from frozen hydrated sections of muscle fibres.

Materials and methods

The major operations before observation in the electron microscope were the preparation and transfer of the specimens.

A. Specimen preparation. Fresh pieces of rabbit skeletal muscle were frozen by rapid immersion in liquid nitrogen chilled Freon 12. Cryo-sectioning was carried out in nitrogen atmosphere at -100°C, or lower, using an LKB CryoKit equipped with items from LKB Cryotools. The sections were manipulated onto pre-cooled grids which were then transferred to a press assembly. Teflon weights were used to ensure successful attachment of the sections to the grid.
Use of the STEM to obtain information from one level of a complex biological object
Authors: Furlong Deirdre
Published: June 2020

Since the beam in the STEM is conical, a specimen is in focus to different precision at different levels. This feature can be used to probe specific levels of complex objects. This property of the STEM was demonstrated using negatively stained, 1% uranyl acetate, Herpes simplex virus nucleocapsids. The capsid is a hollow icosadeltahedral protein shell, 1000A outer diameter, surrounding the DNA containing core. Capsids containing cores were used to prevent collapse of the capsid. The 750Å thick core kept the opposite sides separated by a known distance.

The grid was inserted into the microscope specimen side down. The microscope was focused on the carbon support film adjacent to the capsids. The beam was then shifted to the capsid and an image formed using the signal from the elastically scattered electrons was recorded. By using the signal from the elastically scattered electrons, which is predominantly due to the scattering from the negative stain, the scattering from the unstained core could be neglected. This method mainly gives an image of the negative stain surrounding the capsid.
Frozen ultrathin-sections for X-ray microanalysis of hard tissues
Authors: Ozawa H. , Yamada M. , Yamamoto T. , Takano Y. , Uchida T.
Published: June 2020

In order to clarify more completely the morphophysiology of hard tissues, it is of foremost importance to determine the precise location and the elements of the extracellular and cellular mineral constituents. In the present study we have avoided the potential artefacts which may be introduced by the routine preparation for electron microscopy by adapting both anhydrous and frozen ultrathin-sectioning techniques to our fine structural and elemental analytical studies on hard tissues.

Freshly dissected developing epiphyseal cartilage, bone and dentin of tooth germs from young postnatal rats have been prepared for the analytical electron microscopy(Hitachi H500H Total System equipped with Kevex 7000 X-ray Energy Spectrometry) by the methods of the ultracryo- microtomy(Christensen,1971) and the inert dehydration technique(Pease, 1966.,Landis et al, 1977).

Nuclei, ribosomes, endoplasmic reticulum and a large number of dense intramitochondrial granules, 20 to 80nm in diameter, within the chondrocytes(Fig. 1a,b), osteoblasts(Fig.2) and odontoblasts(Fig.3) were observed in unfixed, unstained frozen ultrathin-sections, obtained with dry knives in the temperature range of -60 to -80°C, and also in unstained sections treated anhydrously by ethylene glycol.
Increase in the intracellular concentration of Na and CI as hepatocytes transform from the normal to the neoplastic state: an electron probe microanalysis study
Authors: Cameron Ivan L. , Smith Nancy R. , Pool Thomas B.
Published: June 2020

We have tested the hypothesis that the intracellular concentrations of various elements are involved in control of cell proliferation. Electron probe microanalysis using energy dispersive X-ray spectroscopy was used to test this hypothesis.

Briefly, unfixed tissues were rapidly frozen in liquified propane, then sectioned at 4ym and cryosorbed. Ribbons of dry sections were transferred to a polished carbon planchet with a 2mm hole drilled in the center and the sections to be analyzed were suspended over the hole. The tissues were electron probed in a scanning microscope. Suspending the tissue over the hole eliminated continuum counts due to substrate and allowed measurement of Na and Mg. The standard conditions during analysis were: accelerating voltage = 15 KV, count time = 100 sec, working distance = 39 mm, detector distance = 1.5 cm, raster size = 1.5 or 2 cm2 (selected area mode), magnification = 8000X, specimen current = 1.0-1.5 x 10-10 amps.
X-ray microanalysis of calcium around the membrane of platelets
Authors: Daimon T. , Mizuhira V.
Published: June 2020

Since the function of platelets is linked to an increase in the cytoplasmic calcium level, it would be of great interest to establish its subcellular distribution. Recently we found the existence of the surface connected membrane system (SCS) in the chicken thrombocyte(1,2) and succeeded in detecting calcium on the membrane of the SCS using ultrahistochemical methods in combination with an electron probe X-ray microanalyzer(1). This study is undertaken to demonstrate calcium accumulation around the membrane of the SCS of mammalian platelets.

Ultrahistochemistry: Human platelet rich buffy coat was fixed in the potassium antimonate-OsO4. method or in the potassium oxalate-glutaraldehyde method without post-fixation with OsO4. After incubation, specimens were dehydrated with ascending concentrations of ethyl alcohol and embedded in Epon 812.
Freeze-fracture, freeze-etch complementary pairs of virus-infected cells reveal value of etching even when relatively high concentrations of cryoprotectants are used
Authors: Steere Russell L. , Erbe Eric F.
Published: June 2020

It has been assumed by many involved in freeze-etch or freeze-fracture studies that it would be useless to etch specimens which were cryoprotected by more than 15% glycerol. We presumed that the amount of cryoprotective material exposed at the surface would serve as a contaminating layer and prevent the visualization of fine details. Recent unexpected freeze-etch results indicated that it would be useful to compare complementary replicas in which one-half of the frozen-fractured specimen would be shadowed and replicated immediately after fracturing whereas the complement would be etched at -98°C for 1 to 10 minutes before being shadowed and replicated.

Standard complementary replica holders (Steere, 1973) with hinges removed were used for this study. Specimens consisting of unfixed virus-infected plant tissue infiltrated with 0.05 M phosphate buffer or distilled water were used without cryoprotectant. Some were permitted to settle through gradients to the desired concentrations of different cryoprotectants.
Lipidic intramembraneous particles
Authors: Ververgaert P.H.J.Th. , Verkleij A.J.
Published: June 2020

Freeze-fracturing splits membranes into 2 halves, thus allowing an examination of the membranes interior. The 5-10 nm particles visible on both monolayers are widely assumed to be proteinaceous in nature.

Most membranes do not reveal impressions complementary to particles on the opposite face. Even when it is considered that shadowing, contamination or fracturing itself might obscure complementary pits, there is no satisfactory explanation why under similar physical circumstances matching halves of other membranes can be visualized. A prominent example of uncomplementarity is found in the erythrocyte membrane. It is well established that band III protein and possibly glycophorln represent these uncomplementary structures.

On the other hand a number of experiments has made us aware that particles of the Eeoheriohia ooli outer membrane are basically constructed of lipid micels (i.e., lipo- polysaccharlde). These particles have complementary Impressions on the opposite face (Fig. 1).
Immune electron microscopy of haemocyanins from two southern african whelks
Authors: Stannard Linda M. , Lennon Margaret
Published: June 2020

Burnupena cincta and Fusus verruculatus are two whelks which inhabit the intertidal zones of the Cape Peninsula shore. Their respiratory pigments, or haemocyanins, are morphologically similar in structure (Figs. 1 and 2) and appear in the electron microscope as short cylindrical rods about 34 nm in diameter and 36 nm high. Viewed side-on the molecules show regular banding suggesting a structure composed of six equidistant rings of sub-units. Occasionally the particles have the appearance of possessing a central “belt” in the position of the 3rd and 4th rows of sub-units. End-on views of the haemocyanin molecules show a circular contour with a dense outer ring and a less dense inner ring in which 10 definite sub-units may frequently be distinguished. A number of molecules display an extra central inner component which appears either as a diffuse plug or as a discrete ring-shaped core ± 8 nm in diameter.
Structural rearrangements studied by topographical mapping of antigenic sites by electron microscopy
Authors: Kistler J. , Aebi U. , Ten Heggeler B. , Onorato L. , Showe M.
Published: June 2020

The maturation of complex viruses like bacteriophages P22, λ and T4 is often regarded as a model system for biological regulatory processes at the molecular level. The visualisation and characterisation of structural rearrangements which occur on the capsid or its related protein assemblies provide information on such processes. In the case of T4, a 16% linear expansion of the capsid is associated with the limited proteolysis of the major capsid protein P23 (MW=58000) to P23* (MW=47000). Image processing on micrographs of negatively stained T4 polyheads (Fig. 1a,d), which are aberrant capsid related cylinders, revealed an altered stain excluding pattern after the expansion (Fig. 1b,e) (Steven et al., 1976; Laemmli et al., 1976).

In order to learn more about the extent and the nature of this structural rearrangement we have followed two procedures: Freeze-drying and shadowing to look at the surface topographies rather than only projection densities obtained from specimen prepared by negative staining (Kistler et al., 1977).
Troponin in embryonic chick skeletal muscle cell in vitro: an immunoelectron microscopic study
Authors: Shimada Yutaka , Obinata Takashi
Published: June 2020

In developing skeletal muscle, myosin and actin are synthesized and polymerized into filamentous forms, thick and thin filaments, respectively. These myofilaments of two varieties have been shown, with the use of "decoration with heavy meromyosin" technique, to be arranged at the right polarity and spatial position as those seen in mature myofibrils from the initial phases of myofibrillogenesis. The question arises as to whether regulatory proteins are distributed along embryonic thin filaments from such early stages of development. In order to clarify this problem, the fine structural localization of troponin was investigated by the use of immunoelectron microscopy.

Troponin and its components (troponin C [TN-C], I [TN-I] and T [TN-T]) were prepared from adult chicken breast muscles. Rabbit antisera against each of these troponin components were prepared. The serum was fractioned by ethanol or ammonium sulphate fractionation. Myogenic cells from 12-day chick embryonic thigh muscles were grown in monolayer and, after 2-8 days in vitro, the cultures were treated as follows: (i) They were immersed in 50% glycerol. The suspension of separated thin filaments was obtained by gentle homogenizing the cells.
Quaternary structure of Limulus polyphemus hemocyanin
Authors: Wichertjes T. , Kwak E.J. , Van Bruggen E.F.J.
Published: June 2020

Hemocyanin of the horseshoe crab (Limulus polyphemus) has been studied in nany ways. Recently the structure, dissociation and reassembly was studied using electron microscopy of negatively stained specimens as the method of investigation. Crystallization of the protein proved to be possible and X-ray crystallographic analysis was started. Also fluorescence properties of the hemocyanin after dialysis against Tris-glycine buffer + 0.01 M EDTA pH 8.9 (so called “stripped” hemocyanin) and its fractions II and V were studied, as well as functional properties of the fractions by NMR. Finally the temperature-jump method was used for assaying the oxygen binding of the dissociating molecule and of preparations of isolated subunits. Nevertheless very little is known about the structure of the intact molecule. Schutter et al. suggested that the molecule possibly consists of two halves, combined in a staggered way, the halves themselves consisting of four subunits arranged in a square.
Electron microscopy of a dna helix destabilizing protein crystal (gp32*i)
Authors: Chiu Wah , Hosoda Junko
Published: June 2020

Gp 32 is a protein coded by gene 32 in bacteriophage T4. It is essential for DNA replication, recombination and repair. Gp32*I with molecular weight of 27,000 was obtained by proteolytic removal of 8000-dalton peptide from COOH-terminal of gp32 (Hosoda et al. to be published). It is a stronger helix destabilizer than gp32, and has been found to be able to replace gp32 in constructing an in vitro DNA replication apparatus, which is active in the leading strand synthesis.

Thin gp32*I crystal was formed under a low salt condition in the absence of glycerol. Figure 1 shows a typical image of the crystal embedded in 1% glucose taken at low magnification. The optical diffractogram in the insert confirms the crystallinity, though no visible feature is seen in the micrograph due to the low magnification power. By using the tungsten shadowing technique, the thickness of the crystal was measured, along with polystyrene latex spheres as a standard, to be between 85 to 400 Å.
Surface structure of the golgi complex and identification of concanavalin-A binding sites
Authors: Sturgess J.M. , Teitelman M. , Moscarello M.A.
Published: June 2020

Scanning electron microscopy has been applied to study the surface ultrastructure of the Golgi complex and labelling techniques have been developed to investigate the distribution of lectin-binding sites on the membrane surfaces. The study is based on the examination of Golgi-rich fractions, isolated by homogenisation and differential centrifugation of rat liver. The membranes are fixed in suspension with 1% glutaraldehyde in 0.1 M sodium phosphate buffer, pH 7.4 for 60 mins and then rinsed in distilled water. For scanning electron microscopy, a thin film of membrane is frozen rapidly on coverglasses using liquid Freon 22, cooled by liquid nitrogen and dried in vacuo at -60°C. Membranes are coated with approximately 100 Å gold in a sputter coater and examined at 20 kV in a JEOL JSM-35U scanning electron microscope. For transmission electron microscopy, membranes are processed as described previously. For examination of lectin binding sites, isolated Golgi membranes are washed in sodium bicarbonate buffer, fixed in glutaraldehyde, incubated with concanavalin A (Con A), rinsed in buffer and then incubated with hemocyanin1.
Ultracytochemical studies on effects of cortisone acetate on the lysosomal system of Yoshida ascites sarcoma cells
Authors: Abe Shyogo , Ogawa Kazuo
Published: June 2020

Effects of cortisone acetate (CA) on the lysosomal system were investigated in Yoshida ascites sarcoma cells maintained by serial intraperitoneal passage at 6-day intervals in the peritoneal cavity of adult male Donryu rats both in vivo and in vitro under the light and the regular (75-100KV) as well as the high voltage (1,000KV) electron microscopes (EM).

In in vivo experiments ascites sarcoma cells taken from CA-treated rats half an hour to 8 hours after intravenous injection of CA (24mg/kg b. w.) suspended in 0. 5ml of saline were investigated. In control rats only saline was injected. Routine morphological observation as well as the demonstration of acid phosphatase (ACPase) activity was performed. Light microscopic observation revealed an obvious increase in number of lysosomes having strongly positive ACPase activity in cells collected 2 to 4 hours after CA injection, but 8 hours after no marked differences were observed between cells taken from the control and CA-treated rats.
An attempt to modify the configurational states of mitochondria from the ependyma of the choroid plexus in vivo.
Authors: Rydzynski Konrad
Published: June 2020

It was shown in vitro-on isolated mitochondria as well as on mitochondria from Ehrlich Ascites Tumor Cells-that adding of 2,4-dinitrophe- nol /DNP/ to the incubation medium leads to characteristic structural changes coinciding with adequate respiratory states. The aim of this study is to investigate, at the ultrastructural level, to what extent it would be possible to modify the configurational states in vivo on the model of the choroid plexus of rats' brain lateral ventricles. It produces the cerebrospinal fluid /CSP/, and is responsible for its composition through the active secretion of certain ions. ATP is indispensable in the process. It has been proved that DNP penetrates the plexus' epithelium and probably inhibits the active processes there by means of uncoupling the oxidative phosphorylation in the mitochondria.

Material and Methods

Wistar rats, weighed 280-330 g,were anesthetized with Nembutal /30 mg/kg i.p./ and fortal /1 mg/kg i.m./, then placed in stereotactic apparatus for ventriculo-cisternal perfusion.
Structure and development of cortical microtubule arrays in plant cells
Authors: Hardham A.R. , Gunning B.E.S.
Published: June 2020

Microtubules in the plant cell cortex are usually aligned parallel to microfibrils of cellulose that are being deposited in the cell wall, and are considered to function in guiding or orienting cellulose synthetase complexes that lie in or on the plasma membrane. The cellulose component is largely responsible for the mechanical reaction of the wall to turgor forces, thereby determining cell size and shape, and therefore the role of the cortical microtubules is a fundamental part of the overall morphogenetic process in plants. It is important to determine the structure of cortical arrays of microtubules and to learn how the cell regulates their development, neither of these aspects having been investigated adequately since the original description likened the microtubules to “hundreds of hoops around the cell”.
Microtubule initiating proteins compose rootlet MTOCs ofpolytomella
Authors: Stearns Mark E. , Brown David L.
Published: June 2020

Based on electron microscopic studies the control of microtubule assembly was originally postulated to reside with electron dense amorphous material localized at the ends of microtubules. These centers which are now more generally recognized as microtubule organizing centers (MTOCs) appear to function in determining the spatial and temporal distribution of microtubules in many developing cell systems. In the alga Polytomella, amorphous material is found coating 8 flagella rootlets to form MTOCs which serve to initiate assembly of a highly patterned cytoskeletal network of microtubules fig. 1. The discovery of conditions for assembly of tubulin in vitro has made possible the reconstitution of an MTOC controlled microtubule assembly system in vitro. We have developed procedures for isolating intact rootlet MTOCs from Polytomella and more recently we have investigated the composition of rootlet MTOCs and the role of these MTOCs in the control of microtubule assembly using phosphocellulose purified tubulin protein.
The effect of nocodazole on microtubule formation and histamine release from mast cells
Authors: Holm Nielsen Ellen , Johansen Torben
Published: June 2020

In exocrine and endocrine gland cells and in nerve cells microtubules (MT) seem to be associated with intracellular transport or secretory activity, although this association has been questioned by contradictory results. A role of MT in secretion of histamine from mast cells is not established. Experiments with drugs, which inhibit either polymerization or depolymerization of MT as colchicine and D2O so far have led to contradictory conclusions. The reason for this may be that the effect of colchicine and D2O in the intact cell is not well defined. This study compares the number and distribution of MT in untreated and in compound 48/80 treated mast cells, and examines the effect of a new synthetical MT inhibitor, Nocodazole, on histamine release and on MT in mast cells.

Mixed peritoneal cells from male Sprague-Dawley rats, 400-500 g, were obtained as previously described and used for the TEM experiments.
A scanning electron microscope study of lectin receptors on neuroblastoma cells
Authors: Maher P. , Molday R.S.
Published: June 2020

The organization and redistribution of concanavalin A (Con A), Ricinus communis agglutinin I (RCA I) and wheat germ agglutinin (WGA) receptors on mouse neuroblastoma cells was analyzed using lectins conjugated to fluorescent polymeric microspheres as visual markers for scanning electron microscopy (SEM) and fluorescent light microscopys. Results indicate that all three lectins are internalized by these cells but the patterns of redistribution for Con A differ from those of RCA I and WGA.

Labeling of prefixed undifferentiated and differentiated neuroblastoma cells indicated that Con A, WGA and RCA I receptors were densely and uniformly distributed over the cell surface and neurites. Cells treated with lectin for 5-10 minutes at 37°C, washed free of excess reagent and maintained for up to 15 minutes in buffer exhibited uniform labeling patterns.
The significance of SEM in the diagnosis of haematopoietic malignancies
Authors: de Harven Etienne , Lampen Nina
Published: June 2020

Samples of heparinized blood, or bone marrow aspirates, or cell suspensions prepared from biopsied tissues (nodes, spleen, etc. ) are routinely prepared, after Ficoll-Hypaque concentration of the mononuclear leucocytes, for scanning electron microscopy. One drop of the cell suspension is placed in a moist chamber on a poly-l-lysine pretreated plastic coverslip (Mazia et al., J. Cell Biol. 66:198-199, 1975) and fifteen minutes allowed for cell attachment. Fixation, started in 2. 5% glutaraldehyde in culture medium at room temperature for 30 minutes, is continued in the same fixative at 4°C overnight or longer. Ethanol dehydration is immediately followed by drying at the critical point of CO2 or of Freon 13. An efficient alternative method for ethanol dehydrated cells is to dry the cells at low temperature (-75°C) under vacuum (10-2 Torr) for 30 minutes in an Edwards-Pearse freeze-dryer (de Harven et al., SEM/IITRI/1977, 519-524). This is preceded by fast quenching in supercooled ethanol (between -90 and -100°C).
Ultrastructure of immunoblastic sarcoma as observed by freeze-fracturing
Authors: Nakai Ei-Ichi , Asamoto Hitoshi , Furuta Mutsuhiro , Nishiura Mitsugu
Published: June 2020

Malignant tumor cells have an enhanced rate of cell division and of metabolic activities of cells. We studied IgG positive immunoblastic sarcoma cells by means of freeze-fracturing and found significant structural differences between these tumor cells and control B-cells.

Lymphnodes infiltrated with sarcoma cells were dissected from the cervical areas of two patients. The sarcoma cell suspension was obtained from these lymphnodes. Sarcoma cells had fine granules stained with PAS and mothyl green pyronin in cytoplasma. Most of sarcoma cells were positive fluorescent (SIgG bearing). Alpha-naphthyl-acetate-esterase test was negative or false positive. Single cell suspension of lymphnodes was obtained from patients of non-lymphatic diseases. EAC rosette forming cells and adherent cells to nylon column fiber were used as control B-cells. The pellet of sarcoma cells and control B-cells were fixed with 2. 5% glutaraldehyde solution. Freeze-fracture was done as described previously. Under the electron microscope, EAC rosette forming cells were selected as control B-cells.
High resolution imaging of the rigid layer of bacterial cell walls at liquid helium temperatures
Authors: Fox F. , Knapek E. , Weyl R.
Published: June 2020

Construction and working capability of a superconducting lens system installed in a 400 kV electron microscope have already been described in. The main features of the imaging possibilities are shown in Fig. 1 As a consequence of cooling the object to liquid helium temperature and the ideal cryovacuum the lens system is especially suited for investigating biological specimens (absence of contamination and ion etching)

We tried to obtain information on the structure of murein or peptido- glycan, the rigid layer of bacterial cell walls. Details on the preparation of the specimens, which are decorated with heavy atoms (3 Hg or Pt) are given elsewhere in these proceedings. Graphite oxide was normally used as a carrier foil with little contrast. The images taken from the murein in most cases showed a large number of dark spots which probably are contaminations dating from the preparation of the specimens (Fig. 2).
The application of ultracryotomy in the study of the fine structure of intracellular virus particles: mouse mammary tumor virus (MMTV)
Authors: Hanaichi T. , Hoshino M. , Nakamura S. , Nishi Y. , Imai M. , Takatani-Yamada C.
Published: June 2020

For the electron microscopy of intracellular virus particles, two techniques have mainly been used; thin sections of plastic- embedded, virus-infected cells and negative staining of the virus particles isolated by disrupting the infected cells. For the observation of minute structures of the particles at high magnification, the latter is often far superior to the former. However, the latter requires the destruction of relations of virus particles to the surrounding cell organelles and may result in the observation of the particles in a distorted state due to the loss of certain materials related to the virus during the isolation process. Ultracryotomy, developed for the past decade, takes advantage of the process of negative staining for the detection of complex intracellular structures at a high resolution. This led us to apply technique to the study of the virus-infected cells. This paper will report the detailed ultrastructures of MMTV at different stages of evolution in the infected cells as revealed by the ultracryotomy technique.
Cytochemical localization of alkaline phosphatase in the cyanobacterium Anabaena cylindrica
Authors: Doonan Barbara B. , Jensen Thomas E.
Published: June 2020

Studies have shown that significant quantities of the enzyme alkaline phosphatase are synthesized by a number of microorganisms only when grown in a phosphate deficient or phosphate free environment. In gram-negative organisms, certain degradative enzymes, including alkaline phosphatase are located within the cell wall. Compartmenta1ization of alkaline phosphatase within the cell wall allows cells efficient and economic functioning by permitting usage of organic substrates when inorganic phosphate is unavailable. As only the phosphate is taken into the cell, substrates which are unable to penetrate the cell membrane can be used by the cells as a result of the alkaline phosphatase placement in the cell wall. The most thoroughly investigated procaryotic alkaline phosphatase is that of Escherichia coli. The purpose of this investigation was to determine the location of alkaline phosphatase activity in the cyanobacterium Anabaena cylindrica (UTEX 629).
Sensitivity of scanning electron microscopy for the detection of mycoplasma contamination
Authors: Quinn Patricia A. , Ho Tai Y.
Published: June 2020

Mycoplasma contamination of cell cultures causes a serious problem in the interpretation of experimental data. These organisms interfere with cell metabolism, induce heritable chromosome abnormalities, alter the antigenicity and function of the cell membrane, contaminate virus preparations and make the interpretation of cell surface structures difficult. Mycoplasmas can be isolated by direct culture. However certain strains will not grow in cell-free media. Indirect methods based on the staining of cultures and the assay of uridine phosphorylase enzyme activity require large numbers of organisms to yield positive results. With SEM, individual particles or clusters on only a few cells in the culture can be visualized.

The actual sensitivity of SEM as a detection method is difficult to evaluate since many mycoplasmas grow in aggregates and reproduce by budding or the breaking apart of filaments as in Fig. 1.
Use of detergents to facilitate penetration of fixatives into thick-walled fungal spores
Authors: Edwards Mercedes R.
Published: June 2020

Fungi are difficult to prepare for good ultra-thin sectioning. Aerial hyphae and certain types of spores have rigid layered walls with outer coats relatively impermeable to common fixatives such as glutaraldehyde and osmium. The slow penetration of these reagents into the cell apparently leads to the formation of artifacts (e. g., mycelin figures and vesicular systems) indicating bad fixation. Poor plastic embedding ensues and many holes are found in the sections, around cell walls or within the cells. Obtaining suitable thin sections from such materials is almost impossible. This problem has been overcome in part by the use of potassium permanganate, which penetrates fungal cells readily and produces good outline of membranes. However, this fixative can destroy most of the cytoplasmic and nuclear matrices and thus is not adequate for the preservation of ribosomes and other cell structures.
Structural aspects of osmoregulation in porphyra
Authors: Wiencke C. , Lauchli A.
Published: June 2020

Osmoregulatory mechanisms in algae were investigated mainly from a physiological point of view (KAUSS 1977, HELLEBUST 1976). In Porphyra two osmotic agents, i. e. floridoside/isofloridoside (KAUSS 1968) and certain ions, such as K+ and Na+(EPPLEY et al. 1960) are considered for osmotic balance. Accumulations of ions (particularly Na+) in the cytoplasm during osmotic adaptation is improbable, because the activity of enzymes is generally inhibited by high ionic concentrations (FLOWERS et al. 1977).

The cellular organization of Porphyra was studied with special emphasis on the development of the vacuolar system under different hyperosmotic conditions. Porphyra was cultivated at various strengths of the culture medium ASP 12 (PROVASOLI 1961) ranging from normal to 6 times concentrated (6x) culture medium. Por electron microscopy freeze fracturing was used (specimens fixed in 2% glutaraldehyde and incubated in 30% glycerol, preparation in a BALZERS BA 360 M apparatus), because chemical fixation gave poor results.
Ultrastructure of mimosa leaflet and pulvinule
Authors: Stein-Margolina V. A.
Published: June 2020

The ultrastructure of leaflet and tertiary pulvini (pulvinule) of the sensitive plant, Mimosa pudica L., as well as the distribution of ATPase were studied before and after stimulation.

There are two zones in the majority of upper epidermis cells of the leaflet (Fig. 1). The upper zone contains a thin layer of cytoplasma, a large central vacuole and much tannin. The lower zone of these cells is electron-transparent and has a lens-like shape. It is filled with fibrillar-foamy material (Fig. 2). “The foam” fibrills are 30-100 Å in diameter. “The foam” is stained with ruthenium red and is supposed to be a mucilaginous inner (lower) wall of the epidermal cell. The cells of the lower epidermis of the leaflet and the pulvinule epidermis do not contain “the foam”. The lens-shaped foamy material might focus light, promote its reception and transfer the light stimuli within the leaflets and pulvini, thus regulating the heliotropic and nyctinastic position of the leaf itself.
Fine structure of the vitteline cells in the cestode Proteocephalus longicollis (proteocephalidea)
Authors: Swiderski Z. , Eklu-natey R. D. , Subilia L. , Huggel H.
Published: June 2020

The mature vitelline cells of Proteocephalus longicollis (Zeder, 1800) are ovoid or spherical and measure about 30μm in diameter. They represent the holocrine type of secretory cells.

The spherical nuclei, about 10μm in diameter, are localized in the central part of the cell. They contain more or less prominent nucleoli enclosed in the moderately electrondense nucleoplasm (Fig. 1).

The cytoplasm (Fig. 2, 3, 4) is packed with numerous shell globules of heterogenous type, large lipid droplets, and a few glycogen islands composed mainly of α-glycogen rosettes.

The remaining granular cytoplasmic matrix (Fig. 3, 4) contains: (a) numerous polysomes, (b) cysternae of granular endoplasmic reticulum, (c) several mitochondria, and (d) extended Golgi regions, composed of vesicles of different sizes and density.

The extensive development of granular endoplasmic reticulum (GER) and Golgi complexes indicates high secretory activity of these cells. Both Golgi and GER are evidently engaged in shell globule formation and are considered therefore to function as the shell-protein producing units
A dense cell in the epithelium of the gill lamellae of the fresh-water brook trout, salvelinus fontinalis
Authors: Morrison C. M.
Published: June 2020

Introduction Unspecialized, dark, chloride and mucous cells have been described in the epithelium of the gill lamellae of Salmo gairdneri and the chloride cell of fresh-water Salvelinus fontinalis has been described. In the course of other studies we found another cell type in the epithelium of fresh-water Salvelinus fontinalis, which is described in the following account.

Methods Six fresh-water brook trout, and two adapted to salt-water were killed by a blow on the head, and gill filaments were removed, fixed in Karnovsky's fixative then osmium tetroxide, dehydrated in acetone and embedded in epoxy resin. Thin sections for electron microscopy were stained with 25% uranyl acetate in methanol and lead citrate.

Observations The epithelial cells described in Salmo gairdneri were also found in Salvelinus fontinalis, but another type was seen in fresh-water Salvelinus fontinalis. These cells were often as large as chloride cells and had a similar shape, but the cytoplasm and basal nucleus were usually darker, round vesicles were present in the apical cytoplasm, and large membrane-bound bodies were often present in the basal cytoplasm (Fig. 1).
Electron microscopy of endocardial cell populations
Authors: Sarphie T. G. , Comer C. R. , Allen D. J.
Published: June 2020

Previous ultrastructural studies have characterized surface morphology during norma cell cycles in an attempt to associate specific changes with specific metabolic processes occurring within the cell. It is now known that during the synthetic ("S") stage of the cycle, when DNA and other nuclear components are synthesized, a cel undergoes a doubling in volume that is accompanied by an increase in surface area whereby its plasma membrane is elaborated into a variety of processes originally referred to as microvilli. In addition, changes in the normal distribution of glycoproteins and polysaccharides derived from cell surfaces have been reported as depreciating after cellular transformation by RNA or DNA viruses and have been associated with the state of growth, irregardless of the rate of proliferation. More specifically, examination of the surface carbohydrate content of synchronous KB cells were shown to be markedly reduced as the cell population approached division Comparison of hamster kidney fibroblasts inhibited by vinblastin sulfate while in metaphase with those not in metaphase demonstrated an appreciable decrease in surface carbohydrate in the former.
Association of microtubules and microfilaments with mucin granules in tracheal goblet cells—their possible role in exocytosis
Authors: Adler Kenneth B. , Brody Arnold R. , Craighead John E.
Published: June 2020

Accumulating experimental evidence suggests that microtubules (MT) and microfilaments (MF) play a major role in the translocation of intracellular organelles. MF are composed of the contractile protein actin; several groups of investigators (Bikle, D. et al: Protoplasma 61:322, 1966; Van Oberghen, E. et al: Diabetes 24:892, 1975) have speculated that these structures provide the force which propels intracellular granules along a pathway delineated by a network of MT. We carried out an ultrastructural investigation of the possible role of MT and MF in the intracellular transport and exocytosis of membrane-bound mucin granules by goblet cells.

Tracheal tissue was excised from newborn female piglets and immersed in 4% gluteraldehyde in a buffer at pH 6. 9 of 1mM GTP, 1mM MgSO4, l00mM PIPES, and 2mM EGTA, a fixative designed to improve visualization of MT (Luftig, R. B. et al: J. Histochem. Cytochem. 25:l75, 1977).
A simple method for ultrastructural visualization of cytoplasmic microfilament bundles (mfb) in rat hepatocytes
Authors: Yokota S. , Fahimi H.D.
Published: June 2020

Microfilaments (MF) are present in a variety of animal and plant cells, and there is substantial evidence that they are involved in cellular motility and contractility (1). Although MF's were first discovered by electron microscopy (EM), they are not distinctly visible in routine EM preparations of whole tissue. In partially disrupted hepatocytes and in isolated bile canaliculi, however, the MF's are prominently contrasted with the uranyl acetate en bloc treatment (2). We now report our observations on the treatment of glutaraldehyde- (GA) fixed rat liver with various detergents. This procedure, followed by uranyl staining, increases markedly the electron density and the contrast of MFB's in tissue sections.

Materials and Methods: Livers of adult rats were fixed by perfusion with 2.5% purified GA in 0.1M Na-cacodylate buffer, pH 7.2, and 0.05% CaCl2 for 10 min. 50-μm chopper sections were treated with 0.1-1% Triton X-100 or sodium deoxycholate (DOC) for 1 h at 25°C, rinsed briefly with the buffer, and then postfixed in 2% aqueous OsO4 for 60 min. at 4°C. This was followed by en bloc staining for 60 min. with 1% uranyl acetate in 0.2M Tris-maleate buffer, pH 5.2, and embedding in Epon. Ultrathin sections were stained with uranyl acetate and lead citrate. In control preparations the detergent treatment was omitted, but all other procedures were identical.