scholarly journals ULTRASTRUCTURE OF DYADS IN MUSCLE FIBERS OF ASCARIS LUMBRICOIDES

1969 ◽  
Vol 42 (3) ◽  
pp. 817-825 ◽  
Author(s):  
Jack Rosenbluth
Keyword(s):  
Membrane Structure ◽  
Osmium Tetroxide ◽  
Phosphotungstic Acid ◽  
Uranyl Acetate ◽  
Double Staining ◽  
Glutaraldehyde Fixation ◽  
Negative Images ◽  
Lead Hydroxide ◽  
T Tubules ◽  
Conventional Methods

The dyads of Ascaris body muscle cells consist of flattened intracellular cisternae applied to the sarcolemma at the cell surface and along the length of T-tubules. In specimens prepared by conventional methods (glutaraldehyde fixation, osmium tetroxide postfixation, double staining of sections with uranyl acetate and lead hydroxide), both the sarcolemma and the limiting membrane of the cisterna exhibit unit membrane structure and the space between them is occupied by a layer of peg-shaped densities which is referred to as the subsarcolemmal lamina. The lumen of the cisterna contains a serrated layer of dense material referred to as the intracisternal lamina. In specimens fixed in glutaraldehyde, dehydrated, and then postfixed in phosphotungstic acid, with no exposure to osmium tetroxide or heavy metal stains, the membranous components of the dyads appear only as negative images, but the subsarcolemmal and intracisternal laminae still appear dense. Except for the lack of density in membranes and in glycogen deposits, the picture produced by the latter method is very much like that of tissue prepared by conventional methods.

The Ultrastructure of Metastatic Human Mammary Carcinoma After Prolonged Estrogen Therapy

1967 ◽  
Vol 25 ◽  
pp. 180-181
Author(s):  
John H.L. Watson ◽  
John L. Swedo ◽  
R.W. Talley
Keyword(s):  
Mammary Carcinoma ◽  
Osmium Tetroxide ◽  
Estrogen Therapy ◽  
Ultrastructural Level ◽  
Uranyl Acetate ◽  
Human Mammary Carcinoma ◽  
Surgical Biopsy ◽  
Lead Hydroxide ◽  
Preliminary Study ◽  
Hormonal Therapies

A preliminary study of human mammary carcinoma on the ultrastructural level is reported for a metastatic, subcutaneous nodule, obtained as a surgical biopsy. The patient's tumor had responded favorably to a series of hormonal therapies, including androgens, estrogens, progestins, and corticoids for recurring nodules over eight years. The pertinent nodule was removed from the region of the gluteal maximus, two weeks following stilbestrol therapy. It was about 1.5 cms in diameter, and was located within the dermis. Pieces from it were fixed immediately in cold fixatives: phosphate buffered osmium tetroxide, glutaraldehyde, and paraformaldehyde. Embedment in each case was in Vestopal W. Contrasting was done with combinations of uranyl acetate and lead hydroxide.

Electron Microscopy of Wound Healing in Patients with Hernia

1975 ◽  
Vol 33 ◽  
pp. 352-353
Author(s):  
C.N. Sun ◽  
H.J. White ◽  
R.C. Read
Keyword(s):  
Electron Microscopy ◽  
Wound Healing ◽  
Phosphate Buffer ◽  
Osmium Tetroxide ◽  
Phosphotungstic Acid ◽  
Rectus Sheath ◽  
Uranyl Acetate ◽  
Collagen Fibrils ◽  
Lead Citrate ◽  
Native Collagen

Previously we have reported the defect of collagen fibrils from herniated rectus sheath. This presentation includes additional sections from postsurgical incisions (10 days) from both control and hernia patients. Small pieces of rectus sheath were fixed in 3% glutaraldehyde in phosphate buffer (pH 7.2) and post fixed with buffered 2% osmium tetroxide. The tissues were then dehydrated in serially increasing concentrations of alcohol and embedded in Epon 812. Sections were stained with 2.5% phosphotungstic acid or uranyl acetate and lead citrate.Previously we found that collagen fibrils from "non-herniated" rectus sheath have uniform diameters and 640 Å periodicity with seven or more intraperiodic bands resembling typical native collagen fibrils, while the fibrils from fascia obtained from patients with direct herniation show considerable variation in diameter. These variations are often found in the same individual fibers with a range from 300 Å to 3000 Å.

scholarly journals PREFERENTIAL STAINING OF NUCLEIC ACID-CONTAINING STRUCTURES FOR ELECTRON MICROSCOPY

1961 ◽  
Vol 11 (2) ◽  
pp. 273-296 ◽  
Author(s):  
H. E. Huxley ◽  
G. Zubay
Keyword(s):  
Nucleic Acid ◽  
Osmium Tetroxide ◽  
Dry Weight ◽  
Uranyl Acetate ◽  
Test Material ◽  
Intense Staining ◽  
Osmium Tetroxide Solution ◽  
Thymus Tissue ◽  
Lead Hydroxide ◽  
Staining Methods

Oriented fibres of extracted nucleohistone were employed as test material in a study of satisfactory fixation, embedding, and staining methods for structures containing a high proportion of nucleic acid. Fixation in buffered osmium tetroxide solution at pH 6, containing 10-2 M Ca++, and embedding in Araldite enabled sections of the fibres to be cut in which the orientation was well preserved. These could be strongly stained in 2 per cent aqueous uranyl acetate, and showed considerable fine structure. Certain regions in the nuclei of whole thymus tissue could also be strongly stained by the same procedure, and were identical with the regions stained by the Feulgen procedure in adjacent sections. Moreover, purified DNA was found to take up almost its own dry weight of uranyl acetate from 2 per cent aqueous solution. Strongest staining of whole tissue was obtained with very short fixation times-5 minutes or so at 0°C. Particularly intense staining was obtained when such tissue stained in uranyl acetate was further stained with lead hydroxide. Although the patterns of staining by lead hydroxide alone and by uranyl acetate were similar in tissues fixed for longer times (½ hour to 2 hours, at 0°C or 20°C), in briefly fixed material the DNA-containing regions appeared relatively unstained by lead hydroxide alone, whilst often there was appreciable staining of RNA-containing structures. Observations on the staining of some viruses by similar techniques are also described.

Uranyl Acetate as a Fixative—from pH 2.0 to 8.0

1971 ◽  
Vol 29 ◽  
pp. 490-491
Author(s):  
V. R. Mumaw ◽  
B. L. Munger
Keyword(s):  
Electron Microscopy ◽  
Oxalic Acid ◽  
Osmium Tetroxide ◽  
Uranyl Acetate ◽  
En Bloc ◽  
Tissue Sections ◽  
Normal Rat ◽  
Lead Hydroxide ◽  
Ultrathin Sections ◽  
Rat Tissue

Numerous applications utilizing uranyl acetate as an electron stain for electron microscopy have been described. Uranyl acetate has become a routine stain used in conjunction with lead hydroxide for staining ultrathin sections. En bloc staining with uranyl acetate following osmium tetroxide post-fixation produces undesirable effects on some cytoplasmic components, especially glycogen. Recent studies using uranyl acetate as a fixative and en bloc stain at pH 7.2 before osmification has shown uranyl acetate to have desirable fixation and staining qualities. Tissues treated with uranyl acetate at a pH of 2.0-8.0 were studied. Normal rat tissue was fixed in Karnovsky's paraformaldehyde-glutaraldehyde fixative. The tissue was post-fixed in 0.5% uranyl acetate in water at pH 2.0 and 0.5% uranyl acetate in 0.1M s-collidine with 0.01M oxalic acid at pH 4, pH 6.0, pH 7.2, and pH 8.0 for 1 hour at 4°C. Following several rinses of 0.1M s-collidine buffer, the tissues were treated with 1.33% osmium tetroxide 1 hour at 4°C followed by rapid dehydration in ethanol and embedded in Durcupan ACM. Tissue sections were stained with lead hydroxide.

scholarly journals INTRAMITOCHONDRIAL FIBERS WITH DNA CHARACTERISTICS

1963 ◽  
Vol 19 (3) ◽  
pp. 593-611 ◽  
Author(s):  
Margit M. K. Nass ◽  
Sylvan Nass
Keyword(s):  
Acetic Acid ◽  
Nucleic Acid ◽  
Chick Embryo ◽  
Potassium Permanganate ◽  
Fibrous Material ◽  
Osmium Tetroxide ◽  
Uranyl Acetate ◽  
Blue Green Algae ◽  
Lead Hydroxide ◽  
Dna Characteristics

Chick embryo mitochondria, studied with the electron microscope, show crista-free areas of low electron opacity. These areas are observable after fixation with osmium tetroxide, calcium permanganate, potassium permanganate, formaldehyde, acrolein, acrolein followed by osmium tetroxide, uranyl acetate followed by calcium permanganate, and acetic acid-alcohol. Staining of sections with lead hydroxide or uranyl acetate, or with both, resulted in an increased density of a fibrous material within these areas. The appearance of the fibrous structures varied with the fixative employed; after fixation with osmium tetroxide the material was clumped and bar-like (up to 400 A in diameter), whereas after treatment of osmium tetroxide-fixed tissues with uranyl acetate before dehydration the fibrous structures could be visualized as 15 to 30 A fibrils. Treatment with ethylenediaminetetraacetate (EDTA) in place of uranyl acetate coarsened the mitochondrial fibrils. After fixation with calcium permanganate or potassium permanganate, or a double fixation by uranyl acetate followed by calcium permanganate, the fibers appeared to have a pattern and ultrastructure similar to that observed after the osmium tetroxide-uranyl acetate technique, except that some of them had a slightly greater diameter (up to 50 A). Other fixatives did not preserve the fibers so well. The fibers appeared strongly clumped by formaldehyde fixation, and were difficult to identify after fixation with acrolein or acetic acid-alcohol. The staining of nucleic acid-containing structures by uranyl acetate and lead hydroxide was improved by treatment of osmium tetroxide-fixed sections with hydrogen peroxide, and the mitochondrial fibers also had an increased density in the electron beam after this procedure. The staining characteristics suggest the fibrous material of chick embryo mitochondria to be a nucleic acid-containing structure, and its variable appearance after different fixations parallels that previously reported, or described in this paper, for the nucleoplasm of bacteria and blue-green algae. The results, in addition to those described in the accompanying communication, indicate that these mitochondria contain DNA.

scholarly journals The Striated Musculature of Blood Vessels

1959 ◽  
Vol 6 (3) ◽  
pp. 383-392 ◽  
Author(s):  
H. E. Karrer
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Electron Microscope ◽  
Osmium Tetroxide ◽  
Inferior Vena ◽  
Phosphotungstic Acid ◽  
Vena Cava ◽  
Uranyl Acetate ◽  
Transverse Tubules ◽  
Thin Sections

The musculature of small lung veins, of the thoracic portion of the inferior vena cava, and of other thoracic veins of the mouse have been studied in the electron microscope. Tissues were fixed in 1 per cent osmium tetroxide buffered with veronal, to which either sodium chloride or sucrose had been added. Methacrylate or araldite served as embedding matrices. Phosphotungstic acid or uranyl acetate was used to stain some of the preparations. Thin sections were examined in a Siemens and Halske Elmiskop Ib electron microscope. The entire musculature of the veins examined was of the striated type. It represents a variety of cardiac muscle, characterized by centrally located nuclei, typical mitochondria, and narrow I bands. Many I bands cannot be recognized at all. H and M bands are likewise indistinct. There is a double array of primary and secondary myofilaments. Mitochondria are large and numerous and contain many cristae. The endoplasmic reticulum consists of longitudinal tubules which run through the whole sarcomeres and bypass Z bands, and of transverse tubules which accompany Z bands. Some "triads," located at Z levels, consist of flattened vacuoles flanked by such transverse tubules. Small vesicles located at Z bands, close to the nucleus, and beneath the plasma membrane may represent still other portions of the reticulum.

scholarly journals ULTRATHIN FROZEN SECTIONS

1967 ◽  
Vol 34 (3) ◽  
pp. 757-771 ◽  
Author(s):  
W. Bernhard ◽  
Elizabeth H. Leduc
Keyword(s):  
Electron Microscopy ◽  
Room Temperature ◽  
Osmium Tetroxide ◽  
Phosphotungstic Acid ◽  
Cultured Cells ◽  
Uranyl Acetate ◽  
Distilled Water ◽  
Frozen Sections ◽  
Simple Method ◽  
Ultrathin Frozen Sections

A relatively simple method for obtaining ultrathin, frozen sections for electron microscopy has been developed. Tissues, cultured cells, and bacteria may be employed. They are fixed in 1.25–4% glutaraldehyde for 1–4 hr, are washed overnight in buffer at 3°C, and are embedded in 20% thiolated gelatin or pure gelatin. Before sectioning they are partially dehydrated in 50% glycerol, frozen in liquid nitrogen on a modified tissue holder, and subsequently maintained at -70°C with dry ice. Finally, they are sectioned very rapidly with glass knives on a slightly modified Porter-Blum MT-1 microtome in a commercial deep-freeze maintained at -35°C and are floated in the trough of the knife on a 40% solution of dimethylsulfoxide (DMSO). The sections are picked up in plastic loops and transferred to distilled water at room temperature for thawing and removal of the DMSO, placed on grids coated with Formvar and carbon, air-dried, and stained with phosphotungstic acid, sodium silicotungstate, or a triple stain of osmium tetroxide, uranyl acetate, and lead. Large flat sections are obtained in which ultrastructural preservation is good. They are particularly useful for cytochemical studies.

Uranyl Acetate as a Stain and a Fixative for Heart Tissue

1969 ◽  
Vol 27 ◽  
pp. 412-413
Author(s):  
M. A. Hayat
Keyword(s):  
Nucleic Acids ◽  
Osmium Tetroxide ◽  
Heart Tissue ◽  
Uranyl Acetate ◽  
Cellular Structures ◽  
Mitochondrial Matrix ◽  
Double Staining ◽  
Lead Salts ◽  
Wide Range ◽  
Cell Components

Lead salts show affinity for a wide range of cellular structures and they increase general contrast much more intensely than any other known electron stain. However, lead stains do not show a strong affinity for nucleic acids, especially DNA. Since uranyl salts exhibit special affinity for nucleic acids, double staining of sections with uranyl acetate followed by lead acetate has become quite popular. Uranyl acetate is also an excellent fixative provided the tissue is treated with this reagent prior to dehydration. After double fixation with glutaraldehyde and osmium tetroxide, uranyl acetate treatment prior to dehydration markedly improves the preservation of all cell components especially that of nucleic acids, ground proteins, myofibril including Z bands, and mitochondrial matrix. Thus, when employed prior to dehydration, uranyl acetate acts not only as a stain but also as a general fixative for both nucleic acids and proteins (Hayat, 1969). Uranyl acetate is so easy to apply to both the tissue block and the sections that there seems no reason for not routinely using it for general staining. In the present study, tissues were exposed to uranyl acetate twice. The details of the method used are given below.

Cellular Contacts Within the Interhemal Membrane of the Rat Placenta at Term

1974 ◽  
Vol 32 ◽  
pp. 146-147
Author(s):  
William P. Jollie
Keyword(s):  
Osmium Tetroxide ◽  
Uranyl Acetate ◽  
Capillary Endothelium ◽  
En Bloc ◽  
Aldehyde Fixation ◽  
Chorioallantoic Placenta

By routine EM preparative techniques, the tissues which, collectively, separate maternal and fetal bloods in the fully formed chorioallantoic placenta of the rat have been shown to consist of three chorionic layers, or trophoblast, and a layer of allantoic capillary endothelium [Fig. 1]. Relationships between these layers are best demonstrated by special techniques, viz., cacodylate-buffered aldehyde fixation, collidine-buffered osmium tetroxide postfixation, and en bloc staining with uranyl acetate. By using this method on placentas at term, the cells of the outermost chorionic layer (Trophoblast 1) appear to be attached to each other by means of maculae adherentes which sometimes occur in clusters [Fig. 2].

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