Use of Uranium-Labeled Antibodies for Electron Microscopic Localization of Various Antigens in Mammalian Cells

1967 ◽  
Vol 25 ◽  
pp. 136-137
Author(s):  
J. P. Petrali ◽  
E. J. Donati ◽  
L. A. Sternberger
Keyword(s):  
Endoplasmic Reticulum ◽  
Hela Cells ◽  
Mammalian Cells ◽  
Specific Antiserum ◽  
Electron Microscopic ◽  
E Coli ◽  
Cytoplasmic Membranes ◽  
Viral Progeny ◽  
Ultrathin Sections ◽  
Electron Microscopic Localization

Specific contrast is conferred to subcellular antigen by applying purified antibodies, exhaustively labeled with uranium under immunospecific protection, to ultrathin sections. Use of Seligman’s principle of bridging osmium to metal via thiocarbohydrazide (TCH) intensifies specific contrast. Ultrathin sections of osmium-fixed materials were stained on the grid by application of 1) thiosemicarbazide (TSC), 2) unlabeled specific antiserum, 3) uranium-labeled anti-antibody and 4) TCH followed by reosmication. Antigens to be localized consisted of vaccinia antigen in infected HeLa cells, lysozyme in monocytes of patients with monocytic or monomyelocytic leukemia, and fibrinogen in the platelets of these leukemic patients. Control sections were stained with non-specific antiserum (E. coli).In the vaccinia-HeLa system, antigen was localized from 1 to 3 hours following infection, and was confined to degrading virus, the inner walls of numerous organelles, and other structures in cytoplasmic foci. Surrounding architecture and cellular mitochondria were unstained. 8 to 14 hours after infection, antigen was localized on the outer walls of the viral progeny, on cytoplasmic membranes, and free in the cytoplasm. Staining of endoplasmic reticulum was intense and focal early, and weak and diffuse late in infection.

Temporal and spatial interrelationships of confronting cisternae to nuclear envelope

1992 ◽  
Vol 50 (1) ◽  
pp. 930-931
Author(s):  
John R. Palisano
Keyword(s):  
Endoplasmic Reticulum ◽  
Nuclear Envelope ◽  
Tumor Cells ◽  
Hela Cells ◽  
Microscopic Investigation ◽  
Electron Microscopic Investigation ◽  
Serial Sections ◽  
Electron Microscopic ◽  
Fetal Rat ◽  
Temporal And Spatial

Although confronting cistemae (CC) have been observed in a variety of tumor cells and normal fetal rat, mouse, and human epithelial tissues, little is known about their origin or role in mitotic cells. While several investigators have suggested that CC arise from nuclear envelope (NE) folding back on itself during prophase, others have suggested that CC arise when fragments of NE pair with endoplasmic reticulum. An electron microscopic investigation of 0.25 um thick serial sections was undertaken to examine the origin of CC in HeLa cells.

scholarly journals Decay-Accelerating Factor and Cytoskeleton Redistribution Pattern in HeLa Cells Infected with RecombinantEscherichia coli Strains Expressing Dr Family of Adhesins

1999 ◽  
Vol 67 (8) ◽  
pp. 3989-3997 ◽  
Author(s):  
Pawel Goluszko ◽  
Rangaraj Selvarangan ◽  
Vsevolod Popov ◽  
Tuan Pham ◽  
Julie W. Wen ◽  
...  
Keyword(s):  
Hela Cells ◽  
Regulatory Protein ◽  
Cytoskeletal Proteins ◽  
Electron Microscopic ◽  
Decay Accelerating Factor ◽  
E Coli ◽  
Recombinant Strains ◽  
Adhesion Sites ◽  
Receptor Aggregation ◽  
Associated Proteins

ABSTRACT Escherichia coli strains expressing Dr fimbriae are able to enter epithelial cells by interacting with a complement-regulatory protein, decay-accelerating factor. This model of bacterial internalization, with a well-characterized bacterial ligand and host receptor, provides a unique opportunity to investigate the early stages of invasion. We used immunofluorescence staining techniques to examine the distribution of receptor and cytoskeletal proteins in HeLa cells infected with E. coli recombinant strains that expressed Dr family of adhesins: Dr, Dr-II, F1845, AFA-I, and AFA-III. A major rearrangement of decay-accelerating factor was found at the adherence sites of recombinant strains expressing Dr, Dr-II, and F1845 adhesins. The changes in the distribution of receptor were significantly smaller on HeLa cells infected with E. coli bearing AFA-I or AFA-III afimbrial adhesins. Receptor aggregation was associated with the redistribution of cytoskeleton-associated proteins such as actin, α-actinin, ezrin, and occasionally tropomyosin. Purified Dr fimbriae coated on polystyrene beads were capable of triggering clustering of receptor and accumulating actin at the adhesion sites of beads to HeLa cells. Using scanning and transmission electron microscopic techniques, we have shown that beads coated with Dr fimbriae, as opposed to beads coated with bovine serum albumin, were enwrapped by cellular microvilli and ultimately internalized into HeLa cells. This indicates that interaction of Dr fimbriae with decay-accelerating factor is associated with redistribution of receptor and is sufficient to promote bacterial internalization.

Electron Microscopic Characterization of Insect Hemocytes

1968 ◽  
Vol 26 ◽  
pp. 68-69 ◽  
Author(s):  
Gertraude Wittig
Keyword(s):  
Endoplasmic Reticulum ◽  
Amorphous Material ◽  
Cell Type ◽  
Electron Microscopic ◽  
Ultrathin Sections ◽  
The Subject ◽  
Insect Hemocytes ◽  
Spherical Cells ◽  
Army Worm

The fine structure of insect hemocytes has been the subject of very few investigations. In particular, the hemocytes of Lepidoptera have received almost no attention. The study presented here was carried out on the armyworm, Pseudaletia unipuncta. Hemocytes of the larva were fixed 2 to 4 days after molt to the sixth instar and studied in ultrathin sections.Microplasmatocytes (Fig. 1) were the most important phagocytes of army-worm hemolymph. They were relatively small, spherical cells with a small, round or lobed nucleus. Distensions of the perinuclear cisterna (p) were frequent and sometimes continuous with the rough endoplasmic reticulum (e). The latter formed greatly distended cisternae which almost filled the whole cytoplasm. The cisternae contained an amorphous material which appeared to be condensed in certain sacs (at e). Mitochondria (m) were rare, and they had tubular cristae. Up to four Golgi complexes (g) were identified in a microplasmatocyte section. Structured granules (sg) were specific for this cell type. Microfibrils (f) traversed the whole cytoplasm but were most frequent around the nucleus (N) and under the cell membrane.

The Function and Metabolism of Certain Insect Muscles in Relation to their Structure

1955 ◽  
Vol s3-96 (34) ◽  
pp. 151-159
Author(s):  
GEORGE A. EDWARDS ◽  
HELMUT RUSKA
Keyword(s):  
Endoplasmic Reticulum ◽  
Oxygen Consumption ◽  
Dehydrogenase Activity ◽  
White Muscle ◽  
Muscle Fibres ◽  
Electron Microscopic ◽  
Leg Muscles ◽  
Red Muscle ◽  
Ultrathin Sections

Electron microscopic observations on ultrathin sections of the red thoracic flightmuscles and white leg muscles of Hydrophilus and Dytiscus are reported. In red muscle-fibres with high values in frequency of contraction, oxygen consumption, and dehydrogenase activity, the single fibrils are completely surrounded by huge mitochondria. Tracheoles penetrate the sarcolemma and supply the mitochondria with oxygen by intracellular branches. In the less active white muscle fibres, mitochondria are found irregularly scattered between the fibrils or along the I band. The intracellular tracheolization is sparse but an endoplasmic reticulum is widely spread between the synfibrillar contractile material. The same muscles of the two insects differ considerably in detail.

scholarly journals Immunogold probes for light and electron microscopic localization of Ole e I in several Oleaceae pollens.

1996 ◽  
Vol 44 (2) ◽  
pp. 151-158 ◽  
Author(s):  
M C Fernandez ◽  
A Olmedilla ◽  
J D Alche ◽  
P Palomino ◽  
C Lahoz ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Major Allergen ◽  
Rapid Screening ◽  
Silver Enhancement ◽  
Electron Microscopic ◽  
Enhancement Technique ◽  
Transmission Electron ◽  
Forsythia Suspensa ◽  
Olive Pollen ◽  
Electron Microscopic Localization

We investigated the immunolocalization of the olive major allergen Ole e I and Ole e I-like proteins in pollen from several Oleaceae species [olive (Olea europaea), ash (Fraxinus excelsior), privet (Ligustrum vulgaris), lilac (Syringa vulgare), and forsythia (Forsythia suspensa)]. Crossreactions among different pollens were found in enzyme immunoassays. For immunolocalization with light microscopy we used the silver enhancement technique with three monoclonal antibodies (1D8, 10H1, and 16G2) that recognize three different epitopes of the allergen Ole e I. Our findings show that the silver enhancement technique is very useful when several antibodies are to be used for rapid screening of different materials. MAb 10H1 gave the most precise results and was selected for further immunolocalization studies with transmission electron microscopy. The epitope recognized by this MAb was localized exclusively in the endoplasmic reticulum in olive pollen. In lilac, privet, and ash pollen, most of the reactivity was also seen in the endoplasmic reticulum; however, the 10H1 epitope was not detected in forsythia pollen.

The Redox Dysregulation Visualized by Electron Microscopic Localization of Cerium Perhydroxide in The Kidney of (Nzb X Nzw)F1 Mice.

1997 ◽  
Vol 3 (S2) ◽  
pp. 59-60
Author(s):  
Megumi Nishikawa ◽  
Li-Li Chen ◽  
Rie Igarashi ◽  
Tomoko Nakazawa ◽  
Eizo Aikawa
Keyword(s):  
Electron Microscopy ◽  
Renal Involvement ◽  
Radical Reactions ◽  
Oxygen Species ◽  
Rt Pcr ◽  
Standard Medium ◽  
Electron Microscopic ◽  
Ultrathin Sections ◽  
Pcr Method ◽  
Electron Microscopic Localization

Redox dysregulation is known as pathogenesis of renal involvement in autoimmune lupus mice [1,2]. However, the behavior of reactive oxygen species (ROS) is still unclear [3]. The purpose of this study is to make clear the localization of hydrogen perhydroxide which is generated as intermediate substances of radical reactions.Modified Brigg's method [4] was used to determine the generation of ROS in renal tissues of (NZB x NZW)F1 mice. The renal specimens were incubated with a standard medium, which consisted of 0. IM tris-malate buffer (pH7.4) with 7% sucrose, lmM CeCl3 and lOmM aminotriazole at 37° C for 30 min and ultrathin sections were prepared and examined by electron microscopy. Lecithinized-recombinant-superoxide dismutase (SOD) was used as redox regulator, which is ROS inhibitor. SOD (1000 U/kg) was injected 3 times a week, tail-intravenously to Fl mice from 4 to 60 weeks of age. The mRNA of inflammatory cytokines and IkB was detected by RT-PCR method.

scholarly journals Electron Microscopic Localization of Russell Bodies in the Human Plasma Cell

Blood ◽  
1960 ◽  
Vol 16 (3) ◽  
pp. 1307-1312 ◽  
Author(s):  
RONALD A. WELSH
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Human Plasma ◽  
Plasma Cell ◽  
Plasma Cells ◽  
Electron Microscopic ◽  
Degenerative Process ◽  
Microscopic Localization ◽  
Electron Microscopic Localization

Abstract The location of Russell bodies in the human plasma cell was shown by electron microscopy to be within the intracisternal space of the endoplasmic reticulum. The significance of this finding was discussed from the standpoint of possible intracellular function of the endoplasmic reticulum. The appearance of the affected plasma cells tended to negate a degenerative process, and the suggestion was offered that the Russell body results from a condensation of intracisternal secretion.

Overexpression of calreticulin fails to abolish its induction by perturbation of normal ER function

1998 ◽  
Vol 76 (5) ◽  
pp. 875-880 ◽  
Author(s):  
David H Llewellyn ◽  
H Llewelyn Roderick
Keyword(s):  
Endoplasmic Reticulum ◽  
Stress Response ◽  
Endoplasmic Reticulum Stress ◽  
Fusion Protein ◽  
Er Stress ◽  
Hela Cells ◽  
Mammalian Cells ◽  
Stress Signaling ◽  
Glucose Response ◽  
The Stability

Along with other endoplasmic reticulum (ER) Ca2+-binding proteins, notably the glucose-response proteins grp78 and grp94, expression of calreticulin is induced in response to perturbation of normal ER function. It has yet to be clearly defined how this stress is signaled from the ER to the nucleus in mammalian cells, particularly with regard to its initiation. Using a GFP-calreticulin fusion protein, we have generated and selected stably transfected HeLa cells that overexpress calreticulin to investigate whether the protein might be involved in signaling its own induction. Basal levels of endogenous calreticulin mRNA and protein were unaffected in these cells, indicating that overexpression alone does not induce a stress response. ER stress induced calreticulin expression in response to either thapsigargin or tunicamycin was equivalent in these cells to that seen in control, nontransfected cells, leading us to conclude that calreticulin is unlikely be involved in its own induction. Levels of the mRNA encoding the fusion protein were also increased by tunicamycin, but not thapsigargin, suggesting that, in agreement with our previous observations, inhibition of N-linked glycosylation may increase the stability of calreticulin mRNA. This indicates that in mammalian cells, there is more than one signaling pathway for the ER stress response.Key words: calreticulin, endoplasmic reticulum stress, signaling.

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