Unstable Expression and Thermal Instability of a Species-Specific Cell Surface Epitope Associated with a 66-Kilodalton Antigen Recognized by Monoclonal Antibody EM-7G1 within Serotypes of Listeria monocytogenes Grown in Nonselective and Selective Broths

1998 ◽  
Vol 64 (8) ◽  
pp. 3070-3074 ◽  
Author(s):  
Ramakrishna Nannapaneni ◽  
Robert Story ◽  
Arun K. Bhunia ◽  
Michael G. Johnson
Keyword(s):  
Monoclonal Antibody ◽  
Listeria Monocytogenes ◽  
Cell Surface ◽  
Brain Heart Infusion ◽  
Live Cells ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Cell ◽  
Enrichment Broth ◽  
Species Specific ◽  
Heat Instability

ABSTRACT Conditions that resulted in unstable expression and heat instability of a cell surface epitope associated with a 66-kDa antigen in Listeria monocytogenes serotypes were identified with the probe monoclonal antibody (MAb) EM-7G1 in an enzyme-linked immunosorbent assay. This epitope appeared to be absent in three serotypes (serotypes 3b, 4a, and 4c), which did not react with MAb EM-7G1 irrespective of the enrichment broth tested. The remaining 10 serotypes were detected by MAb EM-7G1 only when cells were grown in nonselective brain heart infusion broth (BHI) or selectiveListeria enrichment broth (LEB). When cells were grown inListeria repair broth (LRB), only 6 of the 13 serotypes were detected by MAb EM-7G1, and recognition of serogroup 4 was completely lost. None of the 13 serotypes was detected by MAb EM-7G1 when cells were grown in two other commonly usedListeria-selective media, UVM1 broth and Fraser broth (FRB), indicating that possible loss of epitope expression occurred under these conditions. MAb EM-7G1 maintained species specificity without cross-reacting with live or heat-killed cells of six otherListeria spp. (Listeria ivanovii,Listeria innocua, Listeria seeligeri,Listeria welshimeri, Listeria grayi, andListeria murrayi) irrespective of the enrichment conditions tested. Due to heat instability of the cell surface epitope when it was exposed to 80 or 100°C for 20 min, MAb EM-7G1 is suitable for detection of live cells of L. monocytogenes in BHI or LEB but not in LRB, UVM1, or FRB enrichment medium.

Reactivities of Genus-Specific Monoclonal Antibody EM-6E11 against Listeria Species and Serotypes of Listeria monocytogenes Grown in Nonselective and Selective Enrichment Broth Media‡

1998 ◽  
Vol 61 (9) ◽  
pp. 1195-1198 ◽  
Author(s):  
RAMAKRISHNA NANNAPANENI ◽  
ROBERT STORY ◽  
ARUN K. BHUNIA ◽  
MICHAEL G. JOHNSON
Keyword(s):  
Monoclonal Antibody ◽  
Cell Surface ◽  
Brain Heart Infusion ◽  
Live Cells ◽  
Specific Cell ◽  
Bacterial Cells ◽  
Cell Surface Antigens ◽  
Enrichment Broth ◽  
Selective Enrichment ◽  
Listeria Species

Depending on the growth medium used for enrichment of bacterial cells prior to assay, the monoclonal antibody (MAb) EM-6E11 recognizing Listeria genus-specific epitope on 43 and 94 to 97 kDa cell-surface antigens (A. K. Bhunia and M. G. Johnson, Appl. Environ. Microbiol. 58:1924–1929–1992) exhibited extensive variability in the detection of Listeria species. MAb EM-6E11 strongly detected live cells of all Listeria species and all serotypes of L. monocytogenes by ELISA when cells were grown in nonselective brain heart infusion (BHI) broth, in selective Listeria enrichment broth (LEB), or in Listeria repair broth (LRB). In contrast, EM-6E11 detected only four of the thirteen serotypes of L. monocytogenes (serotypes ½c, 3b, 4ab, and 7) when cells were grown in the UVM1 formulation of Listeria enrichment broth (UVM1) or Fraser broth (FRB). This MAb failed to react with live cells of four other Listeria species, including L. ivanovii, L. welshimeri, L. grayi, and L. murrayi cells grown in UVM1 or FRB. Heating of Listeria cells at 100°C for 20 min, irrespective of the enrichment media used, led to large losses of MAb EM-6E11 reactivity in ELISA, suggesting that the specific cell-surface epitopes involved may not be heat stable. Our results confirm that MAb EM-6E11 is suitable for detection of live cells but not heat-killed cells of Listeria spp. and can be used in conjunction with an enrichment step in BHI, LEB, or LRB but not in UVM1 or FRB.

scholarly journals Expression of a developmentally regulated antigen on the surface of skeletal and cardiac muscle cells.

1985 ◽  
Vol 100 (6) ◽  
pp. 1977-1987 ◽  
Author(s):  
S J Kaufman ◽  
R F Foster ◽  
K R Haye ◽  
L E Faiman
Keyword(s):  
Cell Surface ◽  
Cardiac Muscle ◽  
Surface Antigen ◽  
Cardiac Cells ◽  
Cell Surface Antigen ◽  
Myoblast Differentiation ◽  
Specific Cell ◽  
Developmentally Regulated ◽  
Cardiac Muscle Cells ◽  
Species Specific

H36 is a species-specific, cell-surface antigen on differentiating newborn rat skeletal myoblasts and myogenic lines. This membrane antigen has been defined by a monoclonal antibody raised by the fusion of SP 2/0-Ag14 myeloma cells with spleen cells from mice immunized with myotubes derived from the myogenic E63 line. H36 antigen, isolated by immunoaffinity chromatography, is comprised of two polypeptides with apparent molecular weights of 98,000 and 117,000. Fluorescence photometry and radioimmunoassays have been used to follow quantitative and topographic changes in the H36 determinant during myogenesis. H36 is present at a basal level on replicating myoblasts; it increases on prefusion myoblasts and persists on myotubes. At or near the time of prefusion, it becomes concentrated between adjacent aligned myoblasts and localized on membrane "blebs". H36 is present on both skeletal and cardiac cells but absent from a variety of cells that include fibroblasts, neuronal cells, and smooth muscle. There are approximately 4 x 10(5) determinants per myoblast, and the Ka of the antibody is 3.8 x 10(8) liters/mol. The distributions of H36 on the top and attached surfaces of myoblasts and myotubes are distinct, which suggests localized specialization of these surfaces. H36 is an integral membrane component and upon cross-linking, it associates with the detergent-insoluble cytoskeletal framework. Inhibition of myogenesis by 5-bromodeoxyuridine or by calcium deprivation prevents the developmentally associated changes in the expression of H36. H36 is also absent or markedly reduced on the fu- and Ama102 developmentally defective mutant myoblast lines. We conclude that H36 is a muscle-specific, developmentally regulated cell-surface antigen that may have a role in myoblast differentiation and that can be used to determine the embryonic lineages of skeletal and cardiac muscle.

scholarly journals Vitronectin Concentrates Proteolytic Activity on the Cell Surface and Extracellular Matrix by Trapping Soluble Urokinase Receptor-Urokinase Complexes

Blood ◽  
1998 ◽  
Vol 91 (7) ◽  
pp. 2305-2312 ◽  
Author(s):  
Triantafyllos Chavakis ◽  
Sandip M. Kanse ◽  
Barbara Yutzy ◽  
H. Roger Lijnen ◽  
Klaus T. Preissner
Keyword(s):  
Monoclonal Antibody ◽  
Extracellular Matrix ◽  
Cell Surface ◽  
Plasminogen Activator ◽  
Umbilical Vein ◽  
Urokinase Receptor ◽  
Enzyme Linked Immunosorbent Assay ◽  
Tissue Remodelling ◽  
Amino Terminal ◽  
Plasminogen Activator Activity

Abstract Urokinase-type-plasminogen activator (uPA) and its receptor are localized in the vessel wall where they are involved in cellular activation and remodelling processes. Besides the cell surface glycolipid (GPI)-anchored urokinase receptor (uPAR), which binds uPA with high affinity, recent evidence points to the existence of soluble uPAR (suPAR), as well. In the present study, the origin, binding mechanism, and cellular effects of suPAR were examined. Under basal conditions human vascular smooth muscle cells (HVSMC), human umbilical vein endothelial cells (HUVEC), and monocytic cells released 0.1 to 2 ng/mL suPAR, which was increased twofold to fivefold after phorbol ester (PMA) stimulation, as measured by a function-dependent enzyme-linked immunosorbent assay (ELISA). suPAR alone did not bind to HVSMC or HUVEC, but reduced cellular uPA binding by 50% to 70%. However, after removal of GPI-uPAR with phosphatidylinositol-specific phospholipase C, suPAR dose-dependently increased uPA binding by fourfold to fivefold. This increase in binding was completely inhibited by vitronectin (VN) and by a monoclonal antibody against VN, but not by other matrix proteins or antibodies. Thus, VN-mediated uPA binding to cells was regulated by the ratio of soluble to surface-associated uPAR. In a uPAR-deficient cell line (LM-TK−), suPAR increased uPA binding up to 10-fold, whereas the truncated receptor lacking the amino-terminal uPA-binding domain was ineffective. The formation of a ternary uPA/suPAR/VN-complex on the cell surface and the free extracellular matrix could be inhibited by a monoclonal antibody against VN, as well as by plasminogen activator inhibitor-1 (PAI-1). Moreover, VN-mediated binding of the uPA/suPAR-complex led to a fivefold increase in plasminogen activator activity. Through this novel pathway, VN concentrates the uPA/suPAR-complex to cell surfaces and extracellular matrix sites, leading to the accumulation of plasminogen activator activity required for cell migration and tissue remodelling processes.

scholarly journals Anti-Mac-1 selectively inhibits the mouse and human type three complement receptor.

1982 ◽  
Vol 156 (4) ◽  
pp. 1000-1009 ◽  
Author(s):  
D I Beller ◽  
T A Springer ◽  
R D Schreiber
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Cell Surface ◽  
Polymorphonuclear Leukocytes ◽  
Specific Cell ◽  
Complement Receptor ◽  
Murine Macrophage ◽  
Differentiation Antigen ◽  
Human Macrophages ◽  
Specific Cell Surface

Anti-Mac-1 (M1/70), a rat monoclonal antibody that reacts with mouse and human macrophages, polymorphonuclear leukocytes (PMNL), and natural killer cells, selectively inhibited complement receptor-mediated rosetting by murine macrophages and human PMNL. Preincubation of macrophages with anti-Mac-1 inhibited formation of rosettes with sheep erythrocytes bearing IgM antibody and murine C3 fragments. No inhibition was observed when other monoclonal antibodies that react with macrophages (such as anti-Ly5, anti-H-2, or anti-pan-leukocyte) were tested at 10-fold higher concentrations. Anti-Mac-1 did not affect macrophage Fc receptor-mediated rosetting. Erythrocytes bearing homogeneous human C3 fragments C3b (EC3b) or C3bi (EC3bi) were used to test the specificity of the murine macrophage and human PMNL complement receptor inhibited by anti-Mac-1. In both cases, anti-Mac-1 inhibited CR3-mediated rosetting of EC3bi but not CR1-dependent rosetting of EC3b. The results show that Mac-1 is either identical to CR3 or closely associated with CR3 function. This is one of the first cases in which a monoclonal antibody-defined differentiation antigen has been associated with a specific cell surface function.

scholarly journals Comparative study of adenoviruses with monoclonal antibodies

1992 ◽  
Vol 34 (1) ◽  
pp. 19-26 ◽  
Author(s):  
Terezinha Maria de Paiva ◽  
Sueko Takimoto ◽  
María Akíko Ishida ◽  
María Candida Oliveira de Souza ◽  
Tuneo Ishimaru ◽  
...  
Keyword(s):  
Cell Culture ◽  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Comparative Study ◽  
Neutralization Test ◽  
Human Adenovirus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Viral Neutralization ◽  
Species Specific ◽  
Viral Neutralization Test

The obtainment of monoclonal antibodies for adenovirus species 4(Ad4) is described.The specificities of selected monoclonal antibodies were determined by means of viral neutralization test in cell culture, immunofluorescence and Enzyme-Linked Immunosorbent Assay (ELISA), in the presence of the following species of human adenovirus: 1, 2, 5 (subgenus C), 4 (subgenus E), 7 and 16 (subgenus B) and 9 (subgenus D). Two monoclonal antibodies species specific to adenovirus 4 (1CIII and 3DIII) and one monoclonal antibody that cross reacted with adenovirus species 4 and 7 (2HIII) were obtained.

Corneal epithelial-specific cell surface antigen recognized by a monoclonal antibody

Development ◽  
1986 ◽  
Vol 94 (1) ◽  
pp. 163-172
Author(s):  
Marian G. Langer ◽  
C. V. Sundarraj ◽  
Nirmala Sundarraj
Keyword(s):  
Epithelial Cells ◽  
Cell Surface ◽  
Primary Cultures ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Cell ◽  
Immunoperoxidase Technique ◽  
Igg Antibody ◽  
Corneal Epithelial Cells ◽  
Corneal Epithelial ◽  
Human Corneal Epithelial Cells

Monoclonal antibodies, specific against cell surface differentiation antigens of human corneal epithelial cells, were developed using epithelial cells resected from human corneas as the immunogens. One of these antibodies reacted specifically with corneal epithelial cells and not with epithelial cells of other tissues when tested by an indirect immunoperoxidase technique. Nonidet P-40 extracts of different subcellular fractions of human corneal epithelial cells were tested for their reactivity against this antibody using an enzyme-linked immunosorbent assay. The results indicated that the antigen recognized by this antibody is associated with the plasma membrane. This was further verified by immuno-electron-microscopic analysis using ferritin-conjugated anti-mouse IgG antibody. This antigen was not detectable in the corneal epithelial cells in primary cultures nor in the epithelial cells from early stages of developing cornea (12 to 18 weeks in utero) but was present in the epithelial cells in the corneas of an 8-month-old infant. Therefore, this surface-associated antigen identified in the present study is a developmentally regulated marker of human corneal epithelium.

Detection and Isolation of Yersinia pestis Without Fraction 1 Antigen by Monoclonal Antibody in Foods and Water

2012 ◽  
Vol 75 (9) ◽  
pp. 1555-1561 ◽  
Author(s):  
TONG ZHAO ◽  
PING ZHAO ◽  
MICHAEL P. DOYLE
Keyword(s):  
Monoclonal Antibody ◽  
Yersinia Pestis ◽  
Magnetic Beads ◽  
Yersinia Pseudotuberculosis ◽  
Brain Heart Infusion ◽  
Enteric Bacteria ◽  
Immunoblot Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Whole Cells ◽  
A Minor

Most available immunoassays for Yersinia pestis are based on the detection of fraction 1 antigen (F1) when yersiniae are grown at 37°C. A monoclonal antibody (MAb) was developed based on the detection of surface antigens that are not F1. F1-deficient Y. pestis cells were induced and used to immunize BALB/c mice from which MAb (immunoglobulin G1), which specifically recognizes Y. pestis, with or without F1, was obtained. This MAb (6B5) did not cross-react with enteric bacteria, including Yersinia enterocolitica. Enzyme-linked immunosorbent assay results revealed that MAb 6B5 is specific for Y. pestis, with the exception of a minor cross-reaction with Yersinia pseudotuberculosis. Western immunoblot analysis revealed that MAb 6B5 recognizes a Y. pestis outer membrane protein of ca. 30 kDa. Magnetic beads that were coated with MAb 6B5 were compared with beads coated with polyclonal antibody (PAb; rabbit) against Y. pestis for the isolation of Y. pestis in food and water samples by using a PATHATRIX cell concentration apparatus. Enrichment cultures of Y. pestis in different foods by using two different times (6 and 24 h) in brain heart infusion broth at 37°C were evaluated. Results revealed MAb 6B5–coated magnetic beads were equivalent to magnetic beads coated with PAb against Y. pestis A1122 whole cells in concentrating Y. pestis for isolation, especially when samples were enriched for 6 h. However, the selectivity for Y. pestis of the magnetic beads coated with MAb 6B5 was greater than that coated with PAb.

scholarly journals Protection against Candidiasis by an Immunoglobulin G3 (IgG3) Monoclonal Antibody Specific for the Same Mannotriose as an IgM Protective Antibody

2000 ◽  
Vol 68 (3) ◽  
pp. 1649-1654 ◽  
Author(s):  
Yongmoon Han ◽  
Marcia H. Riesselman ◽  
Jim E. Cutler
Keyword(s):  
Monoclonal Antibody ◽  
Cell Surface ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Survival Times ◽  
Vaginal Infection ◽  
Igg Antibody ◽  
Disseminated Candidiasis

ABSTRACT We previously reported that a liposome-mannan vaccine (L-mann) ofCandida albicans induces production of mouse antibodies that protect against disseminated candidiasis and vaginal infection. Immunoglobulin M (IgM) monoclonal antibody (MAb) B6.1, specific for aC. albicans cell surface β-1,2-mannotriose, protects mice against both infections. Another IgM MAb, termed B6, which is specific for a different cell surface mannan epitope, does not protect against disseminated candidiasis. The B6.1 epitope is displayed homogeneously over the entire cell surface, compared to a patchy distribution of the B6 epitope. To determine if protection is restricted to an IgM class of antibody, we tested an IgG antibody. MAb C3.1 was obtained from L-mann-immunized mice. By results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, capture enzyme-linked immunosorbent assay, and immunodiffusion tests, MAb C3.1 is an IgG3 isotype. By epitope inhibition assays, we determined that MAb C3.1 is specific for same mannotriose as MAb B6.1. As expected by the results of the inhibition assays, immunofluorescence microscopy showed that the C3.1 epitope is distributed on the yeast cell surface in a pattern identical to that of the B6.1 epitope. Kidney CFU and mean survival times of infected mice pretreated with MAb C3.1 indicated that the antibody enhanced resistance of mice against disseminated candidiasis. Mice in pseudoestrus that were given MAb C3.1 prior to vaginal infection developed fewer vaginal Candida CFU than control animals that received buffered saline instead of the antibody. The finding that an IgG3 antibody is protective is consistent with our hypothesis that epitope specificity and complement activation are related to the ability of an antibody to protect against candidiasis.

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