scholarly journals A monoclonal antibody against the nucleus reveals the presence of a common protein in the nuclear envelope, the perichromosomal region, and cytoplasmic vesicles.

1987 ◽  
Vol 104 (1) ◽  
pp. 1-7 ◽  
Author(s):  
M Wataya-Kaneda ◽  
Y Kaneda ◽  
T Sakurai ◽  
H Sugawa ◽  
T Uchida
Keyword(s):  
Monoclonal Antibody ◽  
Nuclear Envelope ◽  
Mammalian Cells ◽  
Immunoblot Analysis ◽  
Antigenic Determinants ◽  
Reactive Site ◽  
Premature Chromosome Condensation ◽  
Cytoplasmic Vesicles ◽  
Interphase Cells ◽  
Immunofluorescence Studies

A monoclonal antibody that recognizes antigenic determinants on the nucleus of cultured mammalian cells was isolated. Immunofluorescence studies using this antibody showed that the recognized antigen was present not only on the nucleus but also in cytoplasmic vesicles of interphase cells and in the perichromosomal region of mitotic cells. Premature chromosome condensation analysis showed that the reactive site for this monoclonal antibody could be detected in the perichromosomal region during the G2 and M phases, but not during the G1 and S phases. Finally, immunoblot analysis showed that this monoclonal antibody prepared against the nucleus recognized a protein of approximately 40 kD both in the cytoplasm and in the perichromosomal regions.

The analysis of 40 kDa nuclear protein, p40, in interphase cells and mitotic cells

1993 ◽  
Vol 106 (3) ◽  
pp. 741-748 ◽  
Author(s):  
Y. Kaneda ◽  
K. Kinoshita ◽  
M. Sato ◽  
K. Tanaka ◽  
Y. Kaneda
Keyword(s):  
Monoclonal Antibody ◽  
Nuclear Envelope ◽  
Nuclear Protein ◽  
Dnase I ◽  
High Salt ◽  
The Other ◽  
Interphase Nuclei ◽  
Interphase Cells ◽  
Detergent Treatment ◽  
Sequential Treatments

We previously reported that the monoclonal antibody M108 recognized a 40 kDa protein both in the nucleus and the cytoplasm. This nuclear 40 kDa antigen was located in the nuclear envelope in interphase cells and in the perichromosomal region during mitosis. Now, we have analyzed this nuclear 40 kDa protein (p40) further, through morphological and biochemical approaches. At the beginning of mitosis, the perinuclear p40 detached from the nuclear envelope and moved to surround the condensing chromatin, while in the late stage of mitosis, the perichromosomal p40 moved back to the reassembled nuclear envelope. Most of the perichromosomal p40 on the metaphase chromosome was solubilized only by DNase I treatment, not by either high salt or detergent treatment. On the other hand, the perinuclear p40 was not solubilized by DNase1 alone, or high salt detergent alone. Sequential treatments with DNase I and high salt detergent were required to extract p40 in interphase nuclei. These results suggest that p40 was associated both with the nuclear envelope and chromatin DNA in interphase nuclei, while it bound only to chromatin DNA in mitosis.

Identification of a spindle-associated protein in ciliate micronuclei

1989 ◽  
Vol 93 (2) ◽  
pp. 287-298
Author(s):  
GUY KERYER ◽  
NICOLE GARREAU DE LOUBRESSE ◽  
NICOLE BORDES ◽  
MICHEL BORNENS
Keyword(s):  
Cell Cycle ◽  
Monoclonal Antibody ◽  
Nuclear Envelope ◽  
Mammalian Cells ◽  
Fibrous Material ◽  
Cortical Microtubule ◽  
Ciliated Protozoa ◽  
Paramecium Tetraurelia ◽  
Electron Microscope Level ◽  
Microtubule Organizing Centres

Ciliated protozoa display a nuclear dualism, with germinal micronuciei and a somatic macronucleus. During mitosis, which proceeds without disruption of the nuclear envelope, a spindle is organized within the micronucleus from, presumably, intranuclear microtubule-organizing centres (MTOCs). In order to characterize these MTOCs, monoclonal antibodies generated against human centrosomes were screened on several ciliates and particularly on Paramecium tetraurelia. In this ciliate, the monoclonal antibody CTR 532, which decorates centrosomal and spindle-associated components in mammalian cells, specifically labelled the micronuclei during interphase. At the electron-microscope level, it stained a fibrous material surrounding microtubules localized on the inner face of the nuclear envelope. During mitosis this decoration extended all over the metaphase spindle. At all stages of the cell cycle, the decoration remained specific to the micronucleus and was absent not only from all of the various cytoplasmic and cortical microtubule arrays but also from the macronuclei, even at early stages of their development from the zygotic nucleus. CTR 532 recognizes a single 170x103 Mr polypeptide in the cytoskeletal fraction that contains micronuclei and this polypeptide is absent in the cytoskeletal fraction of amicronucleate cells.

A monoclonal antibody, raised against mammalian centrosomes and screened by recognition of plant microtubule organizing centers, identifies a pericentriolar component in different cell types

1992 ◽  
Vol 101 (4) ◽  
pp. 823-835 ◽  
Author(s):  
V. Chevrier ◽  
S. Komesli ◽  
A.C. Schmit ◽  
M. Vantard ◽  
A.M. Lambert ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Monoclonal Antibody ◽  
Mammalian Cells ◽  
Cell Types ◽  
Immunoblot Analysis ◽  
Dependent Manner ◽  
Specific Staining ◽  
Microtubule Organizing Centers ◽  
Pericentriolar Material ◽  
Cycle Dependent

We have used monoclonal antibodies raised against isolated native calf thymus centrosomes to probe the structure and composition of the pericentriolar material. To distinguish prospective antibodies as specific to conserved elements of this material, we screened clones by their identification of microtubule organizing centers (MTOCs) in different animal and plant cells. Among the clonal antibodies that reacted with MTOCs in both plant and mammalian cells, we describe one (mAb 6C6) that was found to immunostain centrosomes in a variety of bovine and human cells. In cycling cells this signal persisted through the entire cell cycle. Microscopy showed that the mAb 6C6 antigen was a component of the pericentriolar material and this was confirmed by biochemical analysis of centrosomes. Using immunoblot analysis of protein fractions derived from purified components of centrosomes, we have characterized the mAb 6C6 antigen as a 180 kDa polypeptide. We conclude that we have identified a protein component permanently associated with the pericentriolar material. Surprisingly, monoclonal antibody 6C6 also stained other mitotic organelles in mammalian cells, in a cell-cycle-dependent manner. During prometaphase and metaphase the antibody stained both centrosomes and kinetochores. At the onset of anaphase the kinetochore-specific staining dissociated from chromosomes and was subsequently redistributed onto a newly characterized organelle, the telophase disc while the centrosomal stain remained intact. It is not known if the 180 kDa centrosomal protein itself redistributes during mitosis, or if the pattern observed represents other antigens with shared epitopes. The pericentriolar material is thought to be composed of conserved elements, which appeared very early during the evolution of eukaryotes. Our results strongly suggest that mAb 6C6 identifies one of these elements.

scholarly journals The centrin-based cytoskeleton of Chlamydomonas reinhardtii: distribution in interphase and mitotic cells.

1988 ◽  
Vol 107 (2) ◽  
pp. 635-641 ◽  
Author(s):  
J L Salisbury ◽  
A T Baron ◽  
M A Sanders
Keyword(s):  
Nuclear Envelope ◽  
Polyclonal Antibodies ◽  
Flagellar Apparatus ◽  
Immunogold Labeling ◽  
Basal Bodies ◽  
Flagellar Roots ◽  
Cell Biol ◽  
Immunofluorescence Studies ◽  
Descending Fibers

Monoclonal and polyclonal antibodies raised against algal centrin, a protein of algal striated flagellar roots, were used to characterize the occurrence and distribution of this protein in interphase and mitotic Chlamydomonas cells. Chlamydomonas centrin, as identified by Western immunoblot procedures, is a low molecular (20,000-Mr) acidic protein. Immunofluorescence and immunogold labeling demonstrates that centrin is a component of the distal fiber. In addition, centrin-based flagellar roots link the flagellar apparatus to the nucleus. Two major descending fibers extend from the basal bodies toward the nucleus; each descending fiber branches several times giving rise to 8-16 fimbria which surround and embrace the nucleus. Immunogold labeling indicates that these fimbria are juxtaposed to the outer nuclear envelope. Earlier studies have demonstrated that the centrin-based linkage between the flagellar apparatus and the nucleus is contractile, both in vitro and in living Chlamydomonas cells (Wright, R. L., J. Salisbury, and J. Jarvik. 1985. J. Cell Biol. 101:1903-1912; Salisbury, J. L., M. A. Sanders, and L. Harpst. 1987. J. Cell Biol. 105:1799-1805). Immunofluorescence studies show dramatic changes in distribution of the centrin-based system during mitosis that include a transient contraction at preprophase; division, separation, and re-extension during prophase; and a second transient contraction at the metaphase/anaphase boundary. These observations suggest a fundamental role for centrin in motile events during mitosis.

Monoclonal antibody recognizing gp80, a membrane glycoprotein implicated in intercellular adhesion of Dictyostelium discoideum

1983 ◽  
Vol 3 (5) ◽  
pp. 863-870
Author(s):  
B A Murray ◽  
H L Niman ◽  
W F Loomis
Keyword(s):  
Monoclonal Antibody ◽  
Dictyostelium Discoideum ◽  
Posttranslational Modification ◽  
Rabbit Antiserum ◽  
Intercellular Adhesion ◽  
Blocking Effect ◽  
Antigenic Determinants ◽  
Membrane Glycoprotein ◽  
Developmentally Regulated ◽  
Contact Sites

WE have raised a monoclonal antibody, designated E28D8, which reacts with an 80,000-dalton membrane glycoprotein (gp80) of Dictyostelium discoideum. gp80 has been implicated in the formation of the EDTA-resistant adhesions ("contact sites A") which appear during development. The monoclonal antibody reacted with other developmentally regulated proteins of D. discoideum, confirming previous results indicating the presence of common antigenic determinants recognized by polyclonal rabbit antibodies directed to gp80. Periodate sensitivity of the determinants suggests that carbohydrate may be necessary for reactivity. Thus, the determinant recognized by E28D8 may result from a posttranslational modification common to a number of proteins. Some of the proteins that carry the determinant were preferentially localized to posterior cells in slugs. Monoclonal antibody E28D8 did not inhibit contact-sites-A-mediated intercellular adhesion. However, gp80 affinity purified on immobilized monoclonal antibody was able to neutralize the adhesion-blocking effect of rabbit antiserum to gp80. Although gp80 itself may not be essential for cell-cell adhesion, it appears to carry the determinants associated with adhesion.

Monoclonal antibody analysis of serotype-restricted and unrestricted bluetongue viral antigenic determinants

Virology ◽  
1983 ◽  
Vol 124 (2) ◽  
pp. 286-299 ◽  
Author(s):  
Judith A. Appleton ◽  
Geoffrey J. Letchworth
Keyword(s):  
Monoclonal Antibody ◽  
Antigenic Determinants

Differential cytolocalization of prosomes in axolotl during oogenesis and meiotic maturation

1988 ◽  
Vol 90 (4) ◽  
pp. 543-553 ◽  
Author(s):  
J. Gautier ◽  
J.K. Pal ◽  
M.F. Grossi de Sa ◽  
J.C. Beetschen ◽  
K. Scherrer
Keyword(s):  
Mammalian Cells ◽  
De Novo ◽  
Germinal Vesicle ◽  
Immunoblot Analysis ◽  
Ambystoma Mexicanum ◽  
Meiotic Maturation ◽  
Crescent Formation ◽  
Oocyte Growth ◽  
Oocyte Cytoplasm

The prosomes, a novel type of small RNA-protein complex previously characterized in avian and mammalian cells, were isolated from axolotl (Ambystoma mexicanum) oocytes and identified by sedimentation analysis and protein composition. The prosomal nature of these particles was further ascertained by immunoblot analysis with anti-duck prosome monoclonal antibodies. By in vitro [35S]methionine labelling, de novo synthesis of prosomal proteins could be detected neither during oogenesis nor meiotic maturation. The results obtained by both indirect immunofluorescence and immunoblot analyses demonstrated a dramatic change in the localization of prosomal antigens during oocyte development. They were initially detected in the oocyte cytoplasm, during oocyte growth. At the end of vitellogenesis (stages V-VI), they entered the nucleus (germinal vesicle) and were accumulated there to the highest concentration. During oocyte maturation, after nuclear envelope breakdown, prosomal antigens were found to be localized again in the cytoplasm, until fertilization. No specific localization of prosomal antigens in mature oocytes, unfertilized and fertilized eggs was observed within the oocyte cytoplasm in relation to the cytoplasmic rearrangements leading to grey crescent formation.

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