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Biomedical subjects

E de Harven

Publications and source records attributed to E de Harven.

At least 37 records · Page 2Linked to original sources
PubMed1983

Cytochemical methods for the backscattered electron imaging mode of scanning electron microscopy: further applications to the study of human leukemic cells.

The Backscattered Electron Imaging (BEI) mode of Scanning Electron Microscopy (SEM) has been applied to the study of cells stained with various heavy metals in cytochemical reactions. Improvements or modifications of some of these methods and their application to the study of normal and leukemic leukocytes have been evaluated in this report. The results obtained after staining peroxidase-positive granules with osmium, copper, cobalt-nickel and gold-cobalt are compared. Granules containing non-specific esterase activity were demonstrated in the BEI mode after incubation of the cells in Hanker medium and staining with osmium. While sites of acid phosphatase activity were easily localized with a conventional lead method, alkaline phosphatase activity was demonstrated only in the phagocytic vacuoles of cells previously incubated with latex particles and subsequently stained with lead. Cell nuclei were identified in the BEI mode after silver methenamine or bismuth staining. Combining their cell surface and cytochemical characteristics a more accurate identification of the different blood cell types with the SEM becomes possible.

Acid Phosphatase↗
PubMed1982

Immuno-electron microscopic tracing of lactoferrin, a regulator of myelopoiesis, into a subpopulation of human peripheral blood monocytes.

Lactoferrin (LF), an iron binding glycoprotein present in the specific granules of mature granulocytes, is capable of suppressing the production of granulocyte-macrophage colony stimulatory factors (GM-CSF) by monocytes and macrophages in vitro and rebound myelopoiesis in vivo. By an immuno-electron microscopic technique (peroxidase-antiperoxidase procedure) in combination with a three-step fixation and detergent treatment of mononuclear blood cells pulsed with LF, intracellular localization of LF in a sub-population of human monocytes with a well-preserved subcellular morphology was demonstrated. LF was found in the monocytes after a 30 min pulse with concentrations of LF as low as 10(-14) M. Under the electron microscope, deposits of approximately 110 nm diameter appeared scattered in the area of the endoplasmic reticulum. Large numbers of smaller deposits (20-70 nm) were found in the euchromatin. The heterochromatin and other cytoplasmic organelles were free of LF. These results suggest that the euchromatin might be the functional site for LF inhibition of the production of GM-CSF by a subpopulation of human monocytes.

Cytoplasmic Granules↗
PubMed1982

Solid-phase immune electron microscopy-double-antibody technique for rapid detection of papovaviruses.

The solid-phase immune electron microscopy-double-antibody technique, which takes less than 1 h to perform, was applied as a rapid, sensitive, and specific diagnostic tool in the demonstration of papovavirus particles. BK virus propagated in 82C human skin fibroblasts and a monospecific high-titer immune serum to BK virus were used to establish the test procedure. When Formvar-carbon-coated grids were treated with appropriately diluted antibody, a 28-fold increase of virus particles per square micrometer was observed. Viewing of the virus particles was facilitated by the addition of a second "decorator" antibody. BK virus preparations at concentrations of 10(2) to 10(3) PFU/ml could be detected by this technique. There was no cross-reaction with mouse polyomavirus.

Antibodies, Viral↗
PubMed1982

Bone infection caused by debaryomyces hansenii in a normal host: a case report.

Debaryomyces hansenii was isolated from a cystic lesion in the distal tibia of a healthy 23-year-old woman. Ascospores were demonstrated when the organism was incubated at 25 but not at 30 degrees C. Electron microscopy was necessary to demonstrate the warty surface of the ascospore wall. Growth was inhibited in vitro by low concentrations of amphotericin B, 5-fluorocytosine, miconazole, and ketoconazole. Four previous cases of infection caused by Debaryomyces spp. and one caused by the related fungus Torulopsis candida were reviewed.

Adult↗
PubMed1982

Iron carbonyl, a tracer for phagocytosis in scanning electron microscopy.

Human peripheral blood mononuclear cells and polymorphonuclear (PMN) leukocytes, separated by Ficoll-Hypaque have been incubated with a preparation containing iron carbonyl, and studied by scanning electron microscopy (SEM) in the backscattered electron imaging (BEI) mode. Iron carbonyl particles were visible only on the cell surface when studied in the conventional secondary electron (SE) mode of the scanning electron microscope. However, ingested iron carbonyl particles were observed in high contrast when the backscattered electrons were selectively collected. The BEI mode of the scanning electron microscope appeared very useful in identifying phagocytic activity in mixed cell populations and in establishing direct correlations, at the level of each individual cell, between surface morphology and phagocytic activity.

Humans↗
PubMed1981

Ultrastructural cytochemical localizations by back-scattered electron imaging of white blood cells.

White blood cells have been studied in the back-scattered electron imaging (BEI) mode of scanning electron microscopy (SEM) with cytochemical methods for endogenous peroxidase, acid phosphatase, and a silver-staining method for nuclei. Peroxidase-positive granules were seen with good contrast and resolution in myeloid precursor cells and acid phosphatase activity was easily detected in macrophages and monoblasts. Silver staining permitted recognition of the shapes and location of the nuclei. In spite of the cytochemical procedures, cell surface structures were reasonably well-preserved in all methods, making direct correlation of BEI and secondary electron imaging (SEI) images an attractive feature in cell research with the scanning electron microscope.

Acid Phosphatase↗
PubMed1981

Backscattered electron imaging of human leukocytes.

Methods of backscattered electron imaging (BEI) have been applied to the study of normal and malignant human leukocytes. Four different elements, principally of high atomic number, have been used: iron, silver, lead and osmium. Iron, in the form of iron carbonyl, has been found to be an excellent marker for the phagocytic activity of granulocytes and monocytes. Silver staining permitted a clear observation of the shape of the nuclei and of the nucleo-cytoplasmic ratio and made the identification of polymorphonuclear cells and of certain normoblasts possible. Lead phosphate precipitates, deposited as a result of a classic acid phosphatase reaction, were detected in several cases of myelocytic and monocytic leukemias. Precipitates of osmium/diaminobenzidine complex on peroxidase-containing granules permitted the unambiguous identification of several classes of myeloid precursors. In all these techniques, the surface morphology of the cells was adequately preserved and could be correlated with the presence and/or the distribution of the various markers.

Acid Phosphatase↗
PubMed1980

The acute monocytic leukemias: multidisciplinary studies in 45 patients.

The clinical and laboratory features of 37 patients with variants of acute monocytic leukemia are described. Three of these 37 patients who had extensive extramedullary leukemic tissue infiltration are examples of true histiocytic "lymphomas." Three additional patients with undifferentiated leukemias, one patient with refractory anemia with excess of blasts, one patient with chronic myelomonocytic leukemia, one patient with B-lymphocyte diffuse "histiocytic" lymphoma and one patient with "null" cell, terminal deoxynucleotidyl transferase-positive lymphoblastic lymphoma had bone marrow cells with monocytic features. Another patient had dual populations of lymphoid and monocytoid leukemic cells. The true monocytic leukemias, acute monocytic leukemia (AMOL) and acute myelomonocytic leukemia (AMMOL), are closely related to acute myelocytic leukemia (AML) morphologically and by their response to chemotherapy. like AML, the leukemic cells from the AMMOL and AMOL patients form leukemic clusters in semisolid media. Cytochemical staining of leukemic cells for nonspecific esterases, presence of Fc receptor on the cell surface, phagocytic ability, low TdT activity, presence of surface "ruffles" and "ridges" on scanning EM, elevations of serum lysozyme, and clinical manifestations of leukemic tissue infiltration are features which accompanied monocytic differentiation in these cases.

Adolescent↗
PubMed1979

Human epidermal cell cultures: growth and differentiation in the absence of differentiation in the absence of dermal components or medium supplements.

Human epidermal cells grew and differentiated in vitro, provided that the pH of the culture medium was at 5.6-5.8, the seeding density was optimal (approximately 2.5 x 10(5) cells per cm2), and the incubation temperature was maintained at 35-37 degrees C. Under these conditions, epidermal cells from many different skin locations grew to confluency within 15-20 days and formed multi-layered sheets whose differentiated structure resembled that of the full depth of skin epidermis. Cell proliferation and differentiation did not require a feeder layer, a collagen substrate, a high concentration of fetal bovine serum, or added hormones. The sheets of differentiated epidermal cells could be dissociated from the plastic surfaces of the tissue culture flasks. The use of such cultured cells for wound dressing is proposed.

Cell Differentiation↗
PubMed1979

Labeling of lymphocyte surface immunoglobulins with T4 as marker for scanning electron microscopy.

The technique originally described by Kumon et al. for the labeling of cell surface antigens under the SEM has been adapted to the study of surface immunoglobulins on human lymphocytes. Briefly, the peripheral blood lymphocytes are: 1) separated in a Ficoll Hypaque gradient, 2) rinsed with culture medium, 3) allowed to attach to plastic coverslips, 4) prefixed with 1% glutaraldehyde for 10 minutes, 5) incubated with goat anti-human immunoglobulins for 30 minutes at 24 degrees C, 6) rinsed, 7) incubated with a rabbit anti-goat IgG coupled with purified bacteriophage T4, 8) rinsed, 9) postfixed in glutaraldehyde for several hours, and 10) dried from ethanol at -75 degrees C and under 10(-2) Torr. Results from normal human lymphocytes and cells from cases of Chronic Lymphoblastic Leukemia (CLL) indicate that the method makes it possible to recognize cells with surface immunoglobulins and permits some correlation between the surface architecture of the cells and the presence of surface Ig. It also permits the study under the SEM of T-derived lymphocytes and of null cells, the unlabeled cell population, without resorting to techniques for lymphocyte subclass separation which may alter surface morphology.

Humans↗
PubMed1978

The sizes of RNA subunits isolated from high and low leukaemogenic Friend virus.

The sizes of non-covalently linked RNA subunits isolated from highly leukaemogenic Friend virus derived from the plasma (PV, plasma virus) of leukaemic mice were compared to the RNA subunits isolated from low leukaemogenic Friend virus grown in tissue culture (TCV, tissue culture virus). Histograms derived from electron microscope measurements showed that about one-half of the plasma virus RNA was 1.4 to 2.5 micron in length, corresponding to a mol. wt. range from 1.8 X 10(6) to 3.2 X 10(6) and the other half less than 1.4 micron. In contrast, approx. 50% of the TCV RNA was only 0.7 to 1.6 micron in length (mol. wt. 0.9 X 10(6) to 2.0 X 10(6)) and the remainder less than 0.7 micron in length regardless of whether the virus RNA was isolated from 3, 9, 36 or 72 h cultures. The histograms suggest size classes for both TCV and PV derived RNA subunits. Data obtained from sucrose gradient sedimentation of heat-denatured FLV RNA agreed with the electron microscope length measurements. The smaller sizes of the TCV RNA subunits are hypothetically related to the limited biological activity of Friend leukaemia virus produced from leukaemic cells in culture. Comparable results were obtained using two different RNA extraction procedures. Contamination of TCV nucleic acid preparations by cellular DNA was observed even when the virions were harvested from short term cultures and purified by isopycnic sucrose gradient centrifugation. Consequently, preparations of intact virus were treated with DNase prior to sucrose gradient purification of the virions.

Animals↗
PubMed1978

Herpes simplex virus and human cytomegalovirus replication in WI-38 cells. III. Cytochemical localization of lysosomal enzymes in infected cells.

Cytochemical localization of the lysosomal enzymes acid phosphatase and arylsulfatase in cells infected by herpes simplex virus (HSV) or human cytomegalovirus (CMV) showed the following interactions between viruses and host cell lysosomes: (i) many enveloped progeny viruses were located within cytoplasmic vacuoles containing lysosomal enzyme activity; (ii) naked cytoplasmic capsids appeared to acquire an envelope by budding directly into lysosomes; and (iii) many of the cytoplasmic dense bodies that are characteristic of CMV-infected cells and are thought to represent noninfectious aggregates of CMV structural proteins (I. Sarov and I. Abady, Virology 66:464-473, 1975) also acquired a limiting membrane by budding into lysosomes. Autophagy of other cytoplasmic elements was not observed, suggesting that there is some specificity involved in the association of viral particles and CMV dense bodies with lysosomes. Despite the presence of potentially destructive hydrolases, there was little evidence of significant morphological damage to intralysosomal viruses, and high titers of infectious particles were released into the medium. It would therefore appear that significant levels of HSV and CMV infectivity normally persist even though many progeny particles are directly exposed to lysosomal enzymes.

Acid Phosphatase↗