scholarly journals ELECTRON MICROSCOPIC LOCALIZATION OF PHYTOCHROME IN PLANTS USING AN INDIRECT ANTIBODY-LABELING METHOD

1974 ◽  
Vol 22 (11) ◽  
pp. 1039-1047 ◽  
Author(s):  
RICHARD A. COLEMAN ◽  
LEE H. PRATT
Keyword(s):  
Electron Microscope ◽  
Polyethylene Glycol ◽  
Avena Sativa ◽  
Subcellular Distribution ◽  
Electron Microscopic ◽  
Electron Microscope Level ◽  
Antibody Labeling ◽  
Labeling Method ◽  
Specific Rabbit ◽  
Electron Microscopic Localization

The use of polyethylene glycol as a preembedding medium for immunocytochemistry at the electron microscope level has been adapted to the localization of phytochrome in etiolated oat (Avena sativa L., cv. Garry) seedlings. Phytochrome was indirectly labeled in 3-µm sections with rabbit antiperoxidase-peroxidase complex using sheep antirabbit serum and a specific rabbit antiphytochrome serum. Following localization the 3-µm sections were reembedded for ultrathin sectioning. In the absence of information regarding the subcellular distribution of phytochrome, it was necessary to develop a control which would demonstrate that all organelles and areas of the tissue being localized were penetrated by all reagents. Such a control is described. In those cells which contained phytochrome it was found to be generally distributed throughout the cytoplasm and to be associated with both amyloplasts and mitochondria. No activity was observed in nuclei.

Pollen-Wall Proteins: Electron-Microscopic Localization of Acid Phosphatase in the Intine of Crocus Vernus

1971 ◽  
Vol 8 (3) ◽  
pp. 727-733
Author(s):  
R. B. KNOX ◽  
J. HESLOP-HARRISON
Keyword(s):  
Electron Microscope ◽  
Acid Phosphatase ◽  
Coupling Reaction ◽  
Pollen Grain ◽  
Microscope Level ◽  
Electron Microscopic ◽  
Electron Microscope Level ◽  
Microscopic Localization ◽  
Filamentous Inclusions ◽  
Electron Microscopic Localization

Acid phosphatase has been localized in the wall of the pollen grain of Crocus vernus Wulf at the electron-microscope level by a method using 2-naphthyl thiol phosphate as substrate in a simultaneous coupling reaction with fast blue BBN at pH 5.0, the product being given electron opacity by osmication. Activity was found to be concentrated mainly in the intine, and to be associated with ribbon-like or filamentous inclusions believed to be proteinaceous on the basis of other criteria. Some activity was also detectable in the interstices of the exine. The observations confirm the general interpretation of the distribution of wall-held enzyme based upon light-microscopic cytochemistry, and provide the resolution necessary to establish unambiguously that they are associated with protein layers inserted during intine growth.

scholarly journals SOLUBLE HEMIN COMPOUNDS AS ULTRASTRUCTURAL TRACERS

1973 ◽  
Vol 21 (12) ◽  
pp. 1047-1052 ◽  
Author(s):  
J. F. ARONSON ◽  
A. P. FISHMAN ◽  
G. G. PIETRA
Keyword(s):  
Electron Microscope ◽  
Horseradish Peroxidase ◽  
Peroxidase Activity ◽  
Glucuronic Acid ◽  
Prosthetic Group ◽  
Electron Microscopic ◽  
Size Classes ◽  
Ultrastructural Studies ◽  
Microscopic Localization ◽  
Electron Microscopic Localization

We have demonstrated that soluble hemin derivatives have electron microscope-detectable peroxidase activity. Hemin was coupled to several different size classes of dextran, to sucrose and to glucuronic acid. The peroxidase activity of the attached hemin was detected with the electron microscope using the method described by Graham and Karnovsky for horseradish peroxidase. These observations indicate that hemin may be used as a prosthetic group to label macromolecules for electron microscopic localization and raise the prospect of creating groups of inexpensive tracer substances of varying size, charge and specificity for histologic and ultrastructural studies.

scholarly journals LIGHT AND ELECTRON MICROSCOPIC LOCALIZATION OF ANTIGENS IN TISSUES EMBEDDED IN POLYETHYLENE GLYCOL WITH A PEROXIDASE-LABELED ANTIBODY METHOD

1972 ◽  
Vol 20 (12) ◽  
pp. 969-974 ◽  
Author(s):  
JOSEPH E. MAZURKIEWICZ ◽  
PAUL K. NAKANE
Keyword(s):  
Polyethylene Glycol ◽  
Anterior Pituitary ◽  
Room Temperature ◽  
Microscopic Level ◽  
Water Soluble ◽  
Electron Microscopic Level ◽  
Immunocytochemical Localization ◽  
Electron Microscopic ◽  
Antibody Method ◽  
Electron Microscopic Localization

Tissues embedded in polyethylene glycol (PEG) were used for the immunocytochemical localization of cellular antigens at the light and electron microscopic level. For demonstration of the method, growth hormone and luteinizing hormone were localized in the anterior pituitary gland of the rat embedded in PEG, using peroxidase-labeled antibodies. PEG is water-soluble, has a low melting temperature yet is firm enough to be sectioned at room temperature. Preservation of cellular ultrastructure is good. The use of PEG-embedded tissue permits precise correlation of light and electron microscopic observations of the same tissue section.

Electron-microscopic autoradiography of tritiated testosterone in rat testis

1977 ◽  
Vol 26 (1) ◽  
pp. 339-346
Author(s):  
P.M. Frederick ◽  
H.J. van der Molen ◽  
D. Klepper ◽  
H. Galjaard
Keyword(s):  
Electron Microscope ◽  
Seminiferous Tubules ◽  
Cell Cytoplasm ◽  
Electron Microscopic ◽  
Electron Microscope Level ◽  
Tissue Samples ◽  
Electron Microscopic Autoradiography ◽  
Thin Sections ◽  
Freeze Dried ◽  
Lipid Inclusions

The feasibility of a technique for autoradiography of diffusible substances has been further tested by analysing the localization of steroids in rat testes with the light-and electron-microscope. Testes of rats were perfused with tritiated testosterone (3 min) followed by 15-min perfusion with buffer containing a 100-fold of unlabelled testosterone. Tissue samples were frozen, freeze dried, fixed in osmium vapour and embedded in Epon. To exclude extraction of steroids, contact with water and other solvents was prevented during cutting of thin sections on an ultracryotome and further treatments for autoradiography. Light- and electron-microscopic observations indicate that the highest concentration of labelled testosterone was present within the basal parts of the Sertoli cell cytoplasm and in lipid inclusions of Sertoli cells within the seminiferous tubules. This is the first account of autoradiography of steroids at the electron-microscope level.

A Reproducible Selective Stain for Cardiac Sarcoplasmic Reticulum

1974 ◽  
Vol 32 ◽  
pp. 214-215
Author(s):  
R. A. Waugh ◽  
J. R. Sommer
Keyword(s):  
Complex System ◽  
Electron Microscope ◽  
Sarcoplasmic Reticulum ◽  
Transmission Electron Microscope ◽  
Electron Microscopic ◽  
Cardiac Sarcoplasmic Reticulum ◽  
Transmission Electron

Cardiac sarcoplasmic reticulum (SR) is a complex system of intracellular tubules that, due to their small size and juxtaposition to such electron-dense structures as mitochondria and myofibrils, are often inconspicuous in conventionally prepared electron microscopic material. This study reports a method with which the SR is selectively “stained” which facilitates visualizationwith the transmission electron microscope.

Observation of biological macromolecules using various electron microscopic methods

1978 ◽  
Vol 36 (2) ◽  
pp. 170-171
Author(s):  
Mitsuo Ohtsuki ◽  
Michael Sogard
Keyword(s):  
Electron Microscope ◽  
Phase Contrast ◽  
Image Interpretation ◽  
Density Lipoprotein ◽  
Biological Macromolecules ◽  
Contrast Effects ◽  
Biological Structure ◽  
Electron Microscopic ◽  
Structural Investigations ◽  
Staining Techniques

Structural investigations of biological macromolecules commonly employ CTEM with negative staining techniques. Difficulties in valid image interpretation arise, however, due to problems such as variability in thickness and degree of penetration of the staining agent, noise from the supporting film, and artifacts from defocus phase contrast effects. In order to determine the effects of these variables on biological structure, as seen by the electron microscope, negative stained macromolecules of high density lipoprotein-3 (HDL3) from human serum were analyzed with both CTEM and STEM, and results were then compared with CTEM micrographs of freeze-etched HDL3. In addition, we altered the structure of this molecule by digesting away its phospholipid component with phospholipase A2 and look for consistent changes in structure.

Preparatory Procedures for Electron Microscopic Analysis at the Molecular Level of Resolution

1978 ◽  
Vol 36 (3) ◽  
pp. 516-520
Author(s):  
F.J. Sjostrand
Keyword(s):  
Electron Microscope ◽  
Molecular Level ◽  
Microscopic Analysis ◽  
Resolving Power ◽  
Cellular Structures ◽  
Electron Microscopic ◽  
Systematic Analysis ◽  
Technical Developments ◽  
Thin Sectioning ◽  
Obvious Reason

In the 1940's and 1950's electron microscopy conferences were attended with everybody interested in learning about the latest technical developments for one very obvious reason. There was the electron microscope with its outstanding performance but nobody could make very much use of it because we were lacking proper techniques to prepare biological specimens. The development of the thin sectioning technique with its perfectioning in 1952 changed the situation and systematic analysis of the structure of cells could now be pursued. Since then electron microscopists have in general become satisfied with the level of resolution at which cellular structures can be analyzed when applying this technique. There has been little interest in trying to push the limit of resolution closer to that determined by the resolving power of the electron microscope.

Freeze-Fracture Replication in the Ultrastructure of Blue-Green Algae

1967 ◽  
Vol 25 ◽  
pp. 74-75
Author(s):  
L. V. Leak
Keyword(s):  
Cell Wall ◽  
Electron Microscope ◽  
Green Algae ◽  
Three Dimensional ◽  
Freeze Fracture ◽  
Electron Microscopic ◽  
Photosynthetic Membranes ◽  
Blue Green Algae ◽  
Cell Biol ◽  
Cellular Components

Electron microscopic observations of freeze-fracture replicas of Anabaena cells obtained by the procedures described by Bullivant and Ames (J. Cell Biol., 1966) indicate that the frozen cells are fractured in many different planes. This fracturing or cleaving along various planes allows one to gain a three dimensional relation of the cellular components as a result of such a manipulation. When replicas that are obtained by the freeze-fracture method are observed in the electron microscope, cross fractures of the cell wall and membranes that comprise the photosynthetic lamellae are apparent as demonstrated in Figures 1 & 2.A large portion of the Anabaena cell is composed of undulating layers of cytoplasm that are bounded by unit membranes that comprise the photosynthetic membranes. The adjoining layers of cytoplasm are closely apposed to each other to form the photosynthetic lamellae. Occassionally the adjacent layers of cytoplasm are separated by an interspace that may vary in widths of up to several 100 mu to form intralamellar vesicles.

A scanning electron microscopic study on dissolving process of cholesterol gallstones by chenodeoxycholic acid (CDCA)

1978 ◽  
Vol 36 (2) ◽  
pp. 522-523
Author(s):  
T. Kanetaka ◽  
M. Cho ◽  
S. Kawamura ◽  
T. Sado ◽  
K. Hara
Keyword(s):  
Electron Microscope ◽  
Chenodeoxycholic Acid ◽  
Outer Surface ◽  
Electron Microscopic ◽  
Cholesterol Gallstones ◽  
The Core ◽  
Scanning Electron Microscopic Study ◽  
Scanning Electron Microscopic ◽  
Acceleration Voltage ◽  
Scanning Electron

The authors have investigated the dissolution process of human cholesterol gallstones using a scanning electron microscope(SEM). This study was carried out by comparing control gallstones incubated in beagle bile with gallstones obtained from patients who were treated with chenodeoxycholic acid(CDCA).The cholesterol gallstones for this study were obtained from 14 patients. Three control patients were treated without CDCA and eleven patients were treated with CDCA 300-600 mg/day for periods ranging from four to twenty five months. It was confirmed through chemical analysis that these gallstones contained more than 80% cholesterol in both the outer surface and the core.The specimen were obtained from the outer surface and the core of the gallstones. Each specimen was attached to alminum sheet and coated with carbon to 100Å thickness. The SEM observation was made by Hitachi S-550 with 20 kV acceleration voltage and with 60-20, 000X magnification.

Further Observations on the Virus of Epizootic Diarrhea of Infant Mice. An Electron Microscopic Study

1967 ◽  
Vol 25 ◽  
pp. 110-111
Author(s):  
W. G. Banfield ◽  
G. Kasnic ◽  
J. H. Blackwell
Keyword(s):  
Electron Microscope ◽  
Infected Cell ◽  
Microscopic Study ◽  
Intestinal Epithelium ◽  
Ultrastructural Study ◽  
Osmium Tetroxide ◽  
Lead Citrate ◽  
Electron Microscopic ◽  
Virus Particles ◽  
Infected Cells

An ultrastructural study of the intestinal epithelium of mice infected with the agent of epizootic diarrhea of infant mice (EDIM virus) was first performed by Adams and Kraft. We have extended their observations and have found developmental forms of the virus and associated structures not reported by them.Three-day-old NLM strain mice were infected with EDIM virus and killed 48 to 168 hours later. Specimens of bowel were fixed in glutaraldehyde, post fixed in osmium tetroxide and embedded in epon. Sections were stained with uranyl magnesium acetate followed by lead citrate and examined in an updated RCA EMU-3F electron microscope.The cells containing virus particles (infected) are at the tips of the villi and occur throughout the intestine from duodenum through colon. All developmental forms of the virus are present from 48 to 168 hours after infection. Figure 1 is of cells without virus particles and figure 2 is of an infected cell. The nucleus and cytoplasm of the infected cells appear clearer than the cells without virus particles.

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