Three-dimensional ultrastructure of a unicellular cyanobacterium.

The first complete three-dimensional ultrastructural reconstruction of a cyanobacterium was accomplished with high-voltage electron microscopy and computer-aided assembly of serial sections. The precise arrangement of subcellular features within the cell body was very consistent from one cell to another. Specialized inclusion bodies always occupied specific intracellular locations. The photosynthetic thylakoid membranes entirely surrounded the central portion of the cytoplasm, thereby compartmentalizing it from the rest of the cell. The thylakoid membranes formed an interconnecting network of concentric shells, merging only at the inner surface of the cytoplasmic membrane. The thylakoids were in contact with the cytoplasmic membrane at several locations, apparently to maintain the overall configuration of the thylakoid system. These results clarified several unresolved issues regarding structure-function relationships in cyanobacteria.

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Staining Serial Thick Sections For High-Voltage Electron Microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100120230 ◽

1985 ◽

Vol 43 ◽

pp. 708-709

Author(s):

John C. Kinnamon ◽

Terri A. Sherman

Keyword(s):

Electron Microscopy ◽

High Voltage ◽

Carbon Coating ◽

Three Dimensional ◽

Taste Bud ◽

Serial Sections ◽

High Voltage Electron Microscopy ◽

Ultrastructural Studies ◽

Voltage Electron ◽

High Voltage Electron

Ultrastructural studies often require three-dimensional reconstructions from serial electron micrographs. The use of thick sections and high voltage electron microscopy can reduce the numbers of serial sections needed, but we have found that over 300 serial sections (0.5 μm thick) are still needed for our studies of taste bud ultrastructure. Serial reconstructions are almost always done with sections mounted onto formvar-coated slot grids, because they provide an unobscured view of the specimen. Staining and carbon coating several hundred formvar-coated slot grids often results in the loss of serial sections because the formvar films break due to handling during the staining and carbon-coating procedures. In addition, the staining process itself can be tedious and time-consuming when staining thick sections because staining times of hours are often required. Such lengthy staining times may result in staining artifacts due to exposure of the stain to the atmosphere, especially with lead stains.

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Structure of Drosophila polytene chromosomes. Evidence for a toroidal organization of the bands

Journal of Cell Science ◽

10.1242/jcs.57.1.73 ◽

1982 ◽

Vol 57 (1) ◽

pp. 73-113

Author(s):

L.I. Mortin ◽

J.W. Sedat

Keyword(s):

Electron Microscopy ◽

Fluorescent Microscopy ◽

Polytene Chromosomes ◽

Three Dimensional ◽

Structural Features ◽

Serial Sections ◽

High Voltage Electron Microscopy ◽

Voltage Electron ◽

Chromosome Bands ◽

Whole Mounts

Avoiding acid fixation or squashing, the structure of Drosophila salivary gland polytene chromosomes has been examined in detail in nuclei, with special emphasis on the organization of the DNA in the chromosome bands. Cut serial sections, optical serial sections, scanning electron microscopy (SEM) on whole mounts, high-voltage electron microscopy (HVEM) on whole mounts, and pancreatic DNase I digestion monitored by fluorescent microscopy have been used to complement one another in this analysis. With all five of these techniques, stereo pairs were used to aid in the three-dimensional reconstruction of chromosomal structures. Evidence is presented that most, if not all, of the polytene chromosome bands are torus-shaped. The DNA of these bands is largely confined to the rim, with the interior essentially DNA-free. The chromatin in each polytene band is also seen to have an extremely regular and highly ordered substructure. This substructural organization is largely radially symmetric in the bands and generally parallel to the chromosome axis. In addition, each band appears to be a distinct architectural entity with regard to its exact structural features and dimensions. A model is presented that follows these organizational boundary conditions.

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Three-Dimensional Visualization of the T-System of Frog Muscle Using High Voltage Electron Microscopy and a Lanthanum Stain

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100080286 ◽

1977 ◽

Vol 35 ◽

pp. 570-571 ◽

Cited By ~ 2

Author(s):

Lee D. Peachey ◽

Clara Franzini-Armstrong

Keyword(s):

Electron Microscopy ◽

High Voltage ◽

Three Dimensional ◽

Biological Tissues ◽

High Voltage Electron Microscopy ◽

Transverse Tubular System ◽

Peroxidase Reaction ◽

Voltage Electron ◽

High Voltage Electron ◽

T System

The effective study of biological tissues in thick slices of embedded material by high voltage electron microscopy (HVEM) requires highly selective staining of those structures to be visualized so that they are not hidden or obscured by other structures in the image. A tilt pair of micrographs with subsequent stereoscopic viewing can be an important aid in three-dimensional visualization of these images, once an appropriate stain has been found. The peroxidase reaction has been used for this purpose in visualizing the T-system (transverse tubular system) of frog skeletal muscle by HVEM (1). We have found infiltration with lanthanum hydroxide to be particularly useful for three-dimensional visualization of certain aspects of the structure of the T- system in skeletal muscles of the frog. Specifically, lanthanum more completely fills the lumen of the tubules and is denser than the peroxidase reaction product.

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High-voltage electron microscopy of maxillary gland of spirochete-infected brine shrimp: lesion of efferent tubule

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100103875 ◽

1988 ◽

Vol 46 ◽

pp. 362-363

Author(s):

G. E. Tyson ◽

M. J. Song

Keyword(s):

Electron Microscopy ◽

High Voltage ◽

Brine Shrimp ◽

Natural Populations ◽

Three Dimensional ◽

Dimensional Structure ◽

High Voltage Electron Microscopy ◽

Voltage Electron ◽

High Voltage Electron ◽

Infected Adult

Natural populations of the brine shrimp, Artemia, may possess spirochete- infected animals in low numbers. The ultrastructure of Artemia's spirochete has been described by conventional transmission electron microscopy. In infected shrimp, spirochetal cells were abundant in the blood and also occurred intra- and extracellularly in the three organs examined, i.e. the maxillary gland (segmental excretory organ), the integument, and certain muscles The efferent-tubule region of the maxillary gland possessed a distinctive lesion comprised of a group of spirochetes, together with numerous small vesicles, situated in a cave-like indentation of the base of the tubule epithelium. in some instances the basal lamina at a lesion site was clearly discontinuous. High-voltage electron microscopy has now been used to study lesions of the efferent tubule, with the aim of understanding better their three-dimensional structure.Tissue from one maxillary gland of an infected, adult, female brine shrimp was used for HVEM study.

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High-voltage cytochemistry for three-dimensional observation of nuclei and calculation of total numbers of nuclear pores in retinal Mueller glia

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010016234x ◽

1990 ◽

Vol 48 (3) ◽

pp. 956-957

Author(s):

H. Ishigooka ◽

S. Ueno ◽

L.M. Hjelmeland ◽

M.B. Landers ◽

K. Ogawa

Keyword(s):

Nuclear Envelope ◽

High Voltage ◽

Three Dimensional ◽

Reaction Medium ◽

Nuclear Pores ◽

High Voltage Electron Microscopy ◽

Voltage Electron ◽

High Voltage Electron ◽

G6pase Activity ◽

Mueller Glia

Introduction: We have demonstrated that Glucose-6-phosphatase (G6Pase) activity is localized to the endoplasmic reticulum and nuclear envelope of Mueller glia in the normal and pathological guinea pig retina. Using a combination of this cytochemical technique and high voltage electron microscopy, the distribution of nuclear pores could be clearly observed on the nuclear envelope of Mueller glia because of their anatomical lack of reaction products. This technique was developed to study the three-dimensional structure of nuclei and to calculate total numbers of nuclear pores utilizing a computer graphic analysis system in the normal and pathological retina.Materials and methods: Normal and photocoagulated retina of pigmented adult guinea pigs were perfused with a cold mixture of 0.25% glutaraldehyde and 2% paraformaldehyde in 0.1M cacodylate buffer, and the enucleated globes were hemisected and immersed in the same fixative for 30 min. After sectioning and incubation in the reaction medium for the detection of G6Pase activity by the method of Wachstein-Meisel, the sections were postfixed, dehydrated and embedded in Spurr’s epoxy resin. Serial thick sections (1.0um) were prepared for the observation by a Hitachi high voltage electron microscope (H 1250-M) with an accelerating voltage of 1000 Kv. and pictures were analyzed and three-dimensionally reconstructed by TRI (RATOC Co., Ltd.).

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High Voltage Electron Microscopy of Viral Inclusion Bodies

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002117 ◽

1980 ◽

Vol 38 ◽

pp. 486-487 ◽

Cited By ~ 1

Author(s):

H.M. Mazzone ◽

W.F. Engler ◽

G. Wray ◽

A. Szirmae ◽

J. Conroy ◽

...

Keyword(s):

New York ◽

Environmental Protection Agency ◽

Inclusion Bodies ◽

Host Cells ◽

High Voltage Electron Microscopy ◽

Voltage Electron ◽

Viral Particles ◽

Pest Insects ◽

Viral Inclusion ◽

Insect Gut

Viral inclusion bodies isolated from infected pest insects are being evaluated by the U.S. Dept. of Agriculture as biological insecticides against their hosts. Our research on these inclusion bodies constitutes part of an effort to support their approval by the Environmental Protection Agency as insect control agents. The inclusion bodies in this study are polyhedral in shape and contain rod-shaped viral particles. When ingested by pest insects, the inclusion bodies are broken down in the insect gut and release the viral particles which infect and multiply in the nuclei of host cells. These viruses are termed nucleopolyhedrosis viruses (NPV) and are representatives of the baculoviruses (Wildy, P. 1971 IN J.L. Melnick, ed., Monographs in Virology, vol. 5, S.Karger, New York).

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