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At least 163 records · Page 9Linked to original sources
PubMed1988

A simple two-dye basic stain facilitating recognition of mitosis in plastic embedded tissue sections.

Semithin sections of buccal and palatal mucosa fixed in 2.5% glutaraldehyde followed by 1% osmium and embedded in Durcupan (an araldite-based resin) were stained with 2% malachite green in 50% ethanol at 80 C and poststained in 0.05% crystal violet in Sorensen's phosphate buffer (pH 6.4) at 45 C. Nuclear envelopes and chromatin stain vivid purple in contrast to the surrounding green cytoplasm and cell borders. Chromosomes of dividing cells stain bluish violet. Nucleoli, depending on their level in the epithelium, stain differing shades of greenish blue. The distinct and differential staining of each of these components facilitates recognition of mitoses in oral epithelium, where the small size and crowding of cells in the proliferative compartment renders more conventional stains for plastic sections inadequate.

Animals↗
PubMed1990

Optimization of differential immunogold-silver and peroxidase labeling with maintenance of ultrastructure in brain sections before plastic embedding.

The limited success of immunogold labeling for pre-embedding immunocytochemistry of neuronal antigens is largely attributed to poor penetration of large (5-20 nm) colloidal gold particles. We examined the applicability of using silver intensification of 1 nm colloidal gold particles non-covalently bound to goat anti-rabbit immunoglobulin (1) for single labeling of a rabbit antiserum against the catecholamine synthesizing enzyme, tyrosine hydroxylase (TH), and (2) for immunogold localization of rabbit anti-TH simultaneously with immunoperoxidase labeling of a mouse monoclonal antibody against the opiate peptide, leucine-enkephalin (LE). Vibratome sections were collected from acrolein fixed brains of adult rats. These sections were immunolabeled without use of freeze-thawing or other methods that enhance penetration, but damage ultrastructure. By light microscopy, incubations in the silver intensifier (Intense M, Janssen) for less than 10 min at room temperature resulted in a brownish-red reaction product for TH. This product was virtually indistinguishable from that seen using diaminobenzidine reaction for detection of peroxidase immunoreactivity. Longer incubations produced intense black silver deposits that were more clearly distinguishable from the brown immunoperoxidase labeling. However, by light microscopy, the gold particles seen by electron microscopy were most readily distinguished from peroxidase reaction product with shorter silver intensification periods. The smaller size of gold particles with shorter periods of silver intensification also facilitated evaluation of labeling with respect to subcellular organelles. Detection of the silver product did not appear to be appreciably changed by duration of post-fixation in osmium tetroxide. In dual-labeled sections, perikarya and terminals exhibiting immunogold-silver labeling for TH were distinct from those containing immunoperoxidase labeling for LE. These results (1) define the conditions needed for optimal immunogold-silver labeling of antigens while maintaining the ultrastructural morphology in brain, and (2) establish the necessity for controlled silver intensification for light or electron microscopic differentiation of immunogold-silver and peroxidase reaction products and for optimal subcellular resolution.

Animals↗
PubMed1990

Bone lining (endosteal) cells and hematopoiesis: a light microscopic study of normal and pathologic human bone marrow in plastic-embedded sections.

Human trabecular bone that encloses the bone marrow (BM) is covered by a single layer of thin, sometimes inconspicuous, flat, elongated (spindle-shaped) endothelium-like cells with a round or oval nucleus. These "bone lining" cells, or endosteal cells (EC), form a continuous membrane (endosteum) over the trabecular bone surfaces. In most cases, the composition and thickness of these cells do not vary unless the cells are in intimate contact with hematopoietic tissue. In that instance, they are seen as a single layer adjacent to hematopoietic tissue or as a zone of tightly packed or loosely arranged mononuclear (hematopoietic) cells, some apparently originating from the endosteum. In a reparative process, such as following BM harvest, during which bony trabeculae (BT) are mechanically fractured, these cells are seen giving rise to osteoprogenitor (osteoblasts and osteoclasts) cells. Occasionally, the EC appear similar to BM stromal cells (morphologically and by their association with collagen/reticulin fibers) and are best seen at or near the BT that are cut tangentially. Short processes extending from the EC towards the underlying osteocytes have also been observed, suggesting that a channel of communication exists between them and osteocytes. Our observations, coupled with the experimental findings of others (i.e., that hematopoietic stem cells are concentrated near the endosteum, that cells responsible for BM and stroma regeneration are derived from the endosteal layer, and that high concentrations of hematopoietic colony-stimulating factors are produced there), indicate that, in addition to functioning as a simple membranous covering layer for BT, the endosteum helps to support osteocytes and maintains mineral homeostasis.(ABSTRACT TRUNCATED AT 250 WORDS)

Bone Marrow↗
PubMed1992

Plastic embedding procedure for 15 microns sections of large undecalcified tissue blocks.

A method is described that produces sections (15 microns) of large specimens (with dimensions up to 140 x 100 x 200 mm). They are suitable for comparison to CT images of that specimen and can be used as a basis for three-dimensional reconstructions. Frozen unfixed parts of the human body are embedded in polyurethane and scanned with CT. Undecalcified slices, 6 mm thick, of these specimens are cut with a bandsaw. These slices are successfully embedded in polyester resin by means of fixation, dehydration, and subsequent impregnation with this resin. Sections of 15 microns are obtained by cutting the trimmed and sandpapered polyester blocks with an LKB multirange microtome. They are collected on adhesive tape and stained according to Weigert-Azan, sealed between adhesive acetate sheets and photographed on colour slide film. As an example photographs of sections of the human elbow and lumbar spine region are presented.

Cryopreservation↗
PubMed1987

Simultaneous demonstration of bone alkaline and acid phosphatase activities in plastic-embedded sections and differential inhibition of the activities.

Bone alkaline (AlP) and acid phosphatase (AcP) activities were simultaneously demonstrated in tissue sections obtained from mice, rats, and humans. The method involved tissue fixation in ethanol, embedding in glycol methacrylate (GMA), and demonstration of AlP and AcP activities employing a simultaneous coupling azo dye technique using substituted naphthol phosphate as a substrate. AlP activity was demonstrated first followed by AcP activity. Both enzyme activities were demonstrated in tissue sections from bones fixed and/or stored in acetone or 70% ethanol for up to 14 days or stored in GMA for 2 months. AlP activity in tissue sections from bones fixed in 10% formalin, 2% glutaraldehyde, or formal-calcium, however, was markedly inhibited after 3-7 days and was no longer detectable after 14 days of fixation. Moreover, AlP activity was diminished in tissue sections from bones fixed in 70% ethanol or 10% formalin and subsequently demineralized in 10% EDTA (pH 7) for 2 days, and the activity was completely abolished in tissue sections from bones subsequently demineralized in 5% formic acid: 20% sodium citrate (1:1, pH 4.2) for 2 days. Methyl methacrylate (MMA) embedding at concentrations above 66% completely inhibited AlP activity. AcP activity, however, was only partially inhibited by formalin, glutaraldehyde, or formal-calcium after 7 or 14 days of fixation or by MMA embedding and was unaffected by the demineralizing agent formic acid-citrate for 2 days. While AcP activity was preserved in bones fixed in formalin and subsequently demineralized in EDTA, the activity was completely abolished when EDTA demineralization was carried out on bones previously fixed in 70% ethanol.(ABSTRACT TRUNCATED AT 250 WORDS)

Acid Phosphatase↗
PubMed1981

Postembedding immunohistochemical demonstration of antigen in experimental polyarthritis using plastic embedded whole joints.

A method is presented for the immunohistochemical demonstration of antigens in whole undecalcified joints of small laboratory animals. With this method of tissue preparation, involving embedding in a medium mainly based on 2-hydroxyethyl methacrylate, preservation of antigenicity is satisfactory. Antigens can be demonstrated in 2 micron sections by either immunofluorescence or immunoperoxidase and an indirect technique. Therefore in addition to the morphological analysis of joint alterations in experimental polyarthritis, there is now an opportunity to trace the inciting antigen and to study in parallel the enzymatic equipment of the cells involved, using consecutive sections from a single block of tissue.

Animals↗
PubMed1994

Histology of the metal-bone interface: interpretation of plastic embedded slides.

The interference of processing and preparation of histological slides for the study of morphology and morphometry of bone-implant interfaces was investigated in an experimental model, in which a titanium plate was inserted through the cortical bone into the medullary cavity of rat tibiae. The thickness of the sections, burr and notching of the cut border, and staining properties of the embedding resin were found to significantly influence the appearance of the bone-implant interface and, when morphometry was applied, the extent of direct bone-metal contact. The model of the interface resulting from this study is that of some bony processes abutting on the metal surface, while most of the contact is between metal and connective tissue or vascular spaces.

Animals↗
PubMed1989

Tissue distortion in three-dimensional reconstruction of wax or plastic embedded microscopic structures.

Three-dimensional reconstruction at the light microscopic level depends on obtaining reliable serial sections without "distortion" i.e., expansion and compression during section preparation. We have studied the extent of such distortion in serial sections from paraffin and resin embedded blocks of brain, kidney, liver and lung, using an IBAS 2000 Image Analyser. We found that, taking the uncut block as 100%, the section area, perimeter and minimum diameter varied by no more than 8%, except for the lung sections which varied up to 14%. There was no progressive compression due to knife bluntening. Resin sections also varied up to 8% (16% for lungs) but in addition creasing was a problem. We conclude that, provided the serial sectioning is carefully standardised for block shape and orientation, floating out temperature and time, serial paraffin sections are more suitable for three dimensional reconstruction than resin sections.

Acrylic Resins↗
PubMed1988

Proteoglycan in fast-frozen, freeze-dried, plastic-embedded rabbit arteries.

In contrast to glutaraldehyde-fixed vascular tissue with or without staining with cationic dye, the nonfibrous extracellular matrix of fast-frozen, freeze-dried rabbit aorta and renal artery contained a continuous reticulum of fine filaments, closely associated with collagen, elastin, and smooth muscle cells. Three morphologically distinct types of filament were distinguished; one type was selectively sensitive to chondroitinase ABC degradation, and therefore contains chondroitin and/or dermatan sulfate. The remaining filaments of the reticulum may represent the protein core of the proteoglycan monomer, and the hyaluronic acid backbone of the aggregate. Filaments associated with the surface of smooth muscle cells were usually linked to a continuous filament parallel to the cell surface, which was degraded by heparitinase and therefore contains heparan sulfate. The filaments linked directly to the cell surface were not degraded by either enzyme. The preservation of PG in fast-frozen material provides a significant improvement over that obtained by any presently available technique.

Actin Cytoskeleton↗