scholarly journals COLLOIDAL IRON USED AT pH'S LOWER THAN 1 AS ELECTRON STAIN FOR SURFACE PROTEINS

1974 ◽  
Vol 22 (5) ◽  
pp. 368-377 ◽  
Author(s):  
PIERRE R. BLANQUET ◽  
ANNIE LOIEZ
Keyword(s):  
Phospholipase C ◽  
Periodic Acid ◽  
Lipid Extraction ◽  
Surface Membrane ◽  
Ferric Hydroxide ◽  
Chloride Ions ◽  
Surface Proteins ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Colloidal Iron

The effects of hydrochloric acid on stability, charge and selective affinity of the colloidal ferric hydroxide were studied. The charge was tested electrophoretically. The stability was controlled by measuring the turbidity. The affinity was determined by applying colloid to gelled agarose sections containing hyaluronic acid, polyvinyl sulfate or polylysine. Affinity was also determined by applying the colloid to free tumor cells previously submitted to various types of chemical and enzymatic treatments (esterification, acetylation, periodic acid-hydroxylamine method; neuraminidase, phospholipase C, hyaluronidase) and to isolated rat liver surface membranes pretreated by lipid extraction or incubated with phospholipase C. It was found that the chloride ions at pH's lower than 1 bring about the recharging of the conventional positively charged colloid to a negative form. This negative colloid can be used as a new cytochemical method at the electron microscopic level, to visualize with relative specificity positively ionized groups such as the basic amino groups of protein side chains in the outer and inner hydrophilic leaflets of the cell surface membrane.

scholarly journals Colloidal alpha-stannic acid and negative iron colloid as differential electron stains for surface proteins.

1975 ◽  
Vol 23 (3) ◽  
pp. 174-186 ◽  
Author(s):  
P R Blanquet ◽  
E Puvion
Keyword(s):  
Colloidal Particles ◽  
Lipid Extraction ◽  
Ferric Hydroxide ◽  
Surface Protein ◽  
Rat Hepatocytes ◽  
Surface Proteins ◽  
Differential Staining ◽  
Stannic Acid ◽  
Wide Ph Range ◽  
Protein Components

Colloidal alpha-stannic acid and a negative iron colloid obtained from ferric hydroxide and potassium ferrocyanide, both negative sols being stable within a wide pH range, were refined as surface protein electron markers. Because of the relatively small size of its particles, colloidal alpha-stannic acid was used for staining all surface proteins. According to the pH at which the negative iron colloid was applied, it revealed either all surface proteins, or because of its large colloidal particles, stained basic proteins. This differential staining capability of the iron colloidal has been demonstrated previously on various control preparations (Puvion E, Blanquet PR: J Microsc 12:171, 1971). Controls on the affinity of the two colloids to surface amino groups were carried out on rat liver, mouse fibroblasts, HeLa and KB cells, Ehrlich and Zajdela ascites cells subjected to prior enzymatic and chemical treatments (incubation with neuraminidase or phospholipase C, esterification, acetylation or lipid extraction). At any pH below 9, the two sols stained proteins in the outer hydrophilic leaflet of esterified cells with relative selectivity, but the alpha-stannic acid showed them more accurately. The iron sol did reveal at high pH protein components of high isoionic point on the surfaces of rat hepatocytes and ascites cells which had only been treated with neuraminidase.

Bovine Myocardial Epithelial Inclusions

1993 ◽  
Vol 30 (1) ◽  
pp. 82-88 ◽  
Author(s):  
D. C. Baker ◽  
S. P. Schmidt ◽  
K. A. Langheinrich ◽  
L. Cannon ◽  
R. A. Smart
Keyword(s):  
Epithelial Cells ◽  
Periodic Acid ◽  
Intercellular Junctions ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Colloidal Iron ◽  
Periodic Acid Schiff ◽  
Holstein Calves ◽  
The Masses ◽  
Formalin Fixed

Light microscopic, histochemical, immunohistochemical, and ultrastructural methods were used to examine myocardial epithelial masses in the hearts of ten cattle. The tissues consisted of paraffin-embedded or formalin-fixed samples from eight hearts that were being inspected in slaughter houses and from two hearts from calves that died of septicemia. The ages of the cattle ranged from 4 days to 12 years; the breeds were unspecified for all but one Hereford female and the two Holstein calves; and there were three males, four females, and three steers. The masses in these cases were compared with similar appearing lesions found in other animal species. The lesions in the bovine hearts were single to multiple, well circumscribed, found in the left ventricle wall, and composed of squamous to cuboidal epithelial cells that formed tubular, ductular, and acinar structures with lumens that were void or filled with amorphous protein globules. Electron microscopic examination revealed epithelial cells that had sparse apical microvilli, tight apical intercellular junctions, perinuclear bundles of filaments, and rare cilia. Almost half of the bovine epithelial masses (4/9) had occasional diastase-resistant periodic acid-Schiff-positive granules in their cytoplasm, and few had hyaluronidase-resistant alcian blue-positive granules (2/9) or colloidal iron-positive granules (1/9). All myocardial masses had abundant collagen surrounding the tubular and acinar structures, and 2/9 had elastin fibers as well. None of the myocardial masses had Churukian-Schenk or Fontana Masson's silver staining granules in epithelial cells. Immunohistochemically, all bovine myocardial tumors stained positively for cytokeratin (8/8), and occasional masses stained positively for vimentin (3/8) or carcinoembryonic antigen (3/8). None of the masses stained positively for desmin. The myocardial epithelial tumors most likely represent endodermal rests of tissue misplaced during organogenesis.

Silver Staining of Pancreatic Islets as Revealed by Electron Microscopy

1967 ◽  
Vol 25 ◽  
pp. 44-45
Author(s):  
Kazuaki Misugi ◽  
Nobuko Misugi ◽  
Hiroshi Yamada
Keyword(s):  
Pancreatic Islet ◽  
Silver Staining ◽  
Islet Cells ◽  
Severe Hypoglycemia ◽  
Granular Cell ◽  
Electron Microscopic Level ◽  
Staining Reaction ◽  
Electron Microscopic ◽  
Pancreatic Islet Cell ◽  
D Cell

The authors had described the fine structure of a type of pancreatic islet cell, which appeared different from typical alpha and beta cells, and tentatively considered that this third type of granular cell probably represents the D cell (Figure 1).Since silver staining has been widely used to differentiate different types of pancreatic islet cells by light microscopy, an attempt to examine this staining reaction at the electron microscopic level was made.Material and Method: Surgically removed specimens from three infants who suffered from severe hypoglycemia were used. The specimens were fixed and preserved in 20% neutral formalin. Frozen sections, 30 to 40 micron thick, were prepared and they were stained by Bielschowsky's method as modified by Suzuki (2). The stained sections were examined under a microscope and islet tissues were isolated. They were fixed in 1% osmium tetroxide in phosphate buffer for one hour and embedded in Epon 812 following dehydration through a series of alcohols and propylene oxide.

A simplified method of emulsion coating and development for quantitative electron microscopic radioautography

1978 ◽  
Vol 36 (2) ◽  
pp. 64-65
Author(s):  
K. Yoshida ◽  
F. Murata ◽  
S. Ohno ◽  
T. Nagata
Keyword(s):  
Mass Production ◽  
Identical Condition ◽  
Microscopic Level ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Simplified Method ◽  
Complicated Process ◽  
Critical Problems ◽  
Electron Microscopic Radioautography ◽  
Quantitative Radioautography

IntroductionSeveral methods of mounting emulsion for radioautography at the electron microscopic level have been reported. From the viewpoint of quantitative radioautography, however, there are many critical problems in the procedure to produce radioautographs. For example, it is necessary to apply and develop emulsions in several experimental groups under an identical condition. Moreover, it is necessary to treat a lot of grids at the same time in the dark room for statistical analysis. Since the complicated process and technical difficulties in these procedures are inadequate to conduct a quantitative analysis of many radioautographs at once, many factors may bring about unexpected results. In order to improve these complicated procedures, a simplified dropping method for mass production of radioautographs under an identical condition was previously reported. However, this procedure was not completely satisfactory from the viewpoint of emulsion homogeneity. This paper reports another improved procedure employing wire loops.

A New Technique for the Light and Electron Microscopic Study of Individual Cerebro-Spinal Fluid Cells in a Mona Plane Layer

1976 ◽  
Vol 34 ◽  
pp. 362-363
Author(s):  
L.A. Dell
Keyword(s):  
Microscopic Study ◽  
Electron Microscopic Study ◽  
Plane Layer ◽  
Ultrastructural Level ◽  
Spinal Fluid ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Easy Method ◽  
Cell Pellet ◽  
A New Technique

A new method has been developed which readily offers the microscopist a possibility for both light and electron microscopic study of selected cells from the cerebrospinal fluid. Previous attempts to examine these cells in the spinal fluid at the ultrastructural level were based on modifications of cell pellet techniques developed for peripheral blood. These earlier methods were limited in application by the number of cells in spinal fluid required to obtain a sufficient size pellet and by the lack of an easy method of cellular identification between the light and electron microscopic level. The newly developed method routinely employs microscope slides coated with Siliclad and tungsten oxide for duplicate cytocentrifuge preparations of diagnostic spinal fluid specimens. Work done by Kushida and Suzuki provided a basis for our use of the metal oxide.

Electron Microscopic Analysis of Myonecrosis Induced by a Pure Component Isolated From Rattlesnake Venom

1976 ◽  
Vol 34 ◽  
pp. 292-293
Author(s):  
Charlotte L. Ownby ◽  
David Cameron ◽  
Anthony T. Tu
Keyword(s):  
Gel Filtration ◽  
Microscopic Analysis ◽  
The United States ◽  
Exchange Chromatography ◽  
Pure Component ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Mouse Muscle ◽  
Major Health Problem ◽  
Rattlesnake Venom

In the United States the major health problem resulting from snakebite poisoning is local tissue damage, i.e. hemorrhage and myonecrosis. Since commercial antivenin does not usually prevent such damage to tissue, a more effective treatment of snakebite-induced myonecrosis is needed. To aid in the development of such a treatment the pathogenesis of myonecrosis induced by a pure component of rattlesnake venom was studied at the electron microscopic level.The pure component, a small (4,300 mol. wt.), basic (isoelectric point of 9.6) protein, was isolated from crude prairie rattlesnake (Crotalus viridis viridis) venom by gel filtration (Sephadex G-50) followed by cation exchange chromatography (Sephadex C-25), and shown to be pure by electrophoresis. Selection of the myotoxic component was based on light microscopic observations of injected mouse muscle.

X-Ray Microanalysis of Nickel and Cobalt Intensified Peroxidase- Antiperoxidase For Verification of Reaction Sites

1985 ◽  
Vol 43 ◽  
pp. 468-469
Author(s):  
A. Angel ◽  
K. Miller ◽  
V. Seybold ◽  
R. Kriebel
Keyword(s):  
Reaction Mechanisms ◽  
Visual Discrimination ◽  
Osmium Tetroxide ◽  
Ultrastructural Level ◽  
Microscopic Level ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
X Ray ◽  
Insoluble Polymer ◽  
Cytochemical Reaction

Localization of specific substances at the ultrastructural level is dependent on the introduction of chemicals which will complex and impart an electron density at specific reaction sites. Peroxidase-antiperoxidase(PAP) methods have been successfully applied at the electron microscopic level. The PAP complex is localized by addition of its substrate, hydrogen peroxide and an electron donor, usually diaminobenzidine(DAB). On oxidation, DAB forms an insoluble polymer which is able to chelate with osmium tetroxide becoming electron dense. Since verification of reactivity is visual, discrimination of reaction product from osmiophillic structures may be difficult. Recently, x-ray microanalysis has been applied to examine cytochemical reaction precipitates, their distribution in tissues, and to study cytochemical reaction mechanisms. For example, immunoreactive sites labelled with gold have been ascertained by means of x-ray microanalysis.

scholarly journals Vascular Mineralization in the Monkey Brain

1994 ◽  
Vol 31 (5) ◽  
pp. 546-552 ◽  
Author(s):  
T. Yanai ◽  
T. Masegi ◽  
K. Ueda ◽  
J. Manabe ◽  
M. Teranishi ◽  
...  
Keyword(s):  
Periodic Acid ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Periodic Acid Schiff ◽  
X Ray ◽  
Vascular Walls ◽  
Dense Deposits ◽  
Calcium Phosphorus ◽  
Phosphorus Iron ◽  
The Media

Mineralization of various degrees was found in the brains of 79 (59%) of 134 cynomolgus monkeys ( Macaca fascicularis). There was no age dependency in the incidence or severity, nor were there any abnormalities in growth, weight gain, or neurologic signs, although a slight sex difference was observed. The lesions, which were basophilic and intensely positive for periodic acid-Schiff or von Kossa stain, occurred in the vascular walls of the globus pallidus in two types: globoid bodies with prominent concentric lamellar structures in and around the arteriolar and venular wall (type A) and fine granules in the media of small or medium-sized arteries (type B). Electron microscopic examination revealed dense deposits in the degenerated media of small or medium-sized arteries or the thickened walls of the arterioles. X-ray microanalysis demonstrated the presence of calcium, phosphorus, iron, zine, magnesium, and aluminum.

scholarly journals Murine endodermal cytokeratins Endo A and Endo B are localized in the same intermediate filament.

1986 ◽  
Vol 34 (6) ◽  
pp. 785-793 ◽  
Author(s):  
W E Howe ◽  
F G Klier ◽  
R G Oshima
Keyword(s):  
Immunoelectron Microscopy ◽  
Intermediate Filament ◽  
Microscopic Level ◽  
Cytoskeletal Proteins ◽  
Electron Microscopic Level ◽  
Electron Microscopic ◽  
Double Label ◽  
Secondary Antibodies ◽  
Endodermal Cells ◽  
20 Nm

The intracellular distribution of extra-embryonic endodermal, cytoskeletal proteins A (Endo A) and B (Endo B) was investigated by double-label immunofluorescent microscopy and double-label immunoelectron microscopy. In parietal endodermal cells, the immunofluorescent distribution of Endo B was always coincident with that of Endo A and could be distinguished from vimentin, particularly at the periphery of the cell. At the electron microscopic level, antibodies against both Endo A and Endo B recognized both bundles and individual intermediate filaments. Double-label immunoelectron microscopy was achieved by use of two sizes of colloidal gold particles (5 nm and 20 nm) that were stabilized with secondary antibodies. These results show that Endo A and B are found in the same intermediate filament and probably co-polymerize to form such structures.

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