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C Wagener

Publications and source records attributed to C Wagener.

At least 19 recordsLinked to original sources
PubMed1992

A monoclonal anti-idiotypic antibody bearing the image of an epitope specific to the human carcinoembryonic antigen.

One concept for immune therapy of patients bearing carcinomas involves monoclonal anti-idiotypic antibodies (Malds) to trigger the immune system of the host into a response against the tumor cells. Current theory states that so-called internal image Malds bearing epitopes specific to a given tumor-associated antigen would be most suited for that purpose. We report here the generation of syngeneic Malds generated against the murine monoclonal immunoglobulin T84.66 (Ab1), which defines a single epitope on the protein moiety of the carcinoembryonic antigen (CEA). This antigenic determinant is unique to CEA, as it is absent in other members of the CEA gene family that are expressed in a variety of normal human tissues, including granulocytes. The Mald 6G6.C4 (Ab2) exhibits the immunochemical features of an internal image antibody mimicking the epitope recognized by the idiotype T84.66. In enzyme immunoassays the binding of Ab1 to Ab2 is completely inhibited by CEA. In addition, Mald 6G6.C4 only binds to native but not to the denatured or reduced idiotype. Immunization of rabbits with F(ab')2-fragments of 6G6.C4 results in antisera (Ab3) that show specificity to CEA in both binding and inhibition enzyme immunoassays as well as in Western blots. Finally, Ab3 did not detect NCA, a major CEA-related glycoprotein in Western blots, either in a purified form or in a crude tumor extract, indicating a high specificity of the anti-anti-idiotypic response. In summary, these immunochemical data show that the monoclonal anti-idiotype 6G6.C4 can functionally mimic a CEA-specific epitope in rabbits and may do so in humans. Therefore, this antibody may have a clinical potential as a network antigen for active immune therapy in patients suffering from CEA-positive carcinomas.

Animals↗
PubMed1992

Specificity and affinity of monoclonal antibodies against carcinoembryonic antigen.

The binding specificities of 52 well-characterized monoclonal antibodies (Mabs) against carcinoembryonic antigen (CEA) from 12 different research groups were studied by immunohistochemistry and immuno flow cytometry. In addition, the binding constant for the interaction between Mab and CEA was determined by a solution-phase assay. Cryostat sections of colon carcinoma and normal colon, stomach, liver, pancreas, and spleen were studied by immunohistochemistry. Peripheral blood granulocytes, monocytes, and lymphocytes were assayed by immuno flow cytometry. The Mabs used here have previously been classified into five essentially nonoverlapping epitope groups (GOLD 1-5) (Cancer Res., 49: 4852-4858, 1989). Most Mabs cross-reacted with different normal tissues, ranging from highly cross-reactive Mabs (positive reaction with 8 of 9 discriminating tissues) to relatively specific Mabs (positive reaction with 1 of 9 discriminating tissues). Five Mabs (10%) were specific, reacting only with colon carcinoma, normal colon mucosa, and normal gastric foveola. There was a correlation between epitope group and binding specificity. Mabs with a high degree of CEA specificity almost exclusively belonged to epitope groups 1, 2, and 3, while highly cross-reactive Mabs belonged to epitope groups 4 and 5. There was no correlation between antibody specificity and affinity for CEA. Specific Mabs with high as well as low affinity were found.

Adult↗
PubMed1992

Guided tissue regeneration in interproximal defects in the monkey.

The potential for guided tissue regeneration was evaluated in one-walled interproximal sites in Macaca fascicularis. Histologic differences were evaluated at 1 and 3 months. Within the experimental (barrier) group, 100% of the root surfaces with potential for regeneration were covered with new cementum, whereas the control specimens had 20% or less new cementum. The amount of regeneration was determined by the position of the barrier membrane; the more coronal the barrier, the greater the regeneration. Observations indicated that the optimal time for barrier removal is between 1 and 3 months.

Animals↗
PubMed1991

Purification and N-terminal amino acid sequence of a chondroitin sulphate/dermatan sulphate proteoglycan isolated from intima/media preparations of human aorta.

A proteoglycan (PG) was purified to homogeneity from intima/media preparations of human aorta specimens by the following chromatographic steps: Sepharose Q anion exchange, Sepharose CL-4B size exclusion, hydroxyapatite, MonoQ anion exchange and TSK G 4000 SW size exclusion. The purity of the preparation was established by SDS/PAGE using direct staining by silver or Dimethylmethylene Blue, as well as by Western blots of biotin-labelled samples. The electrophoretic mobility of the native PG was less than that of a 200,000-Mr standard protein. After treatment with chondroitin sulphate lyase ABC, a core protein of Mr 15,000 was revealed. The Mr of the glycosaminoglycan (GAG) peptides was less than 24,000, by comparison with a keratan sulphate peptide. The composition of the GAG chains was determined by differential digestion of the PG by chondroitin sulphate lyases AC/ABC or chondroitin sulphate lyase AC alone followed by anion-exchange chromatography of the resulting disaccharides. The GAG chains are composed of approximately one-third of dermatan sulphate and two-thirds chondroitin sulphate disaccharide units. The sequence of the 20 N-terminal amino acids is identical with the sequence previously reported for PG I isolated from human developing bone [Fisher, Termine & Young (1989) J. Biol. Chem. 264, 4571-4576]. The assignment of glycosylation sites to the serine residues in positions 5 and 10 was confirmed. The findings indicate that the chondroitin sulphate/dermatan sulphate PG is a major PG in intima/media preparations of human aorta and represents a biglycan-type PG.

Amino Acid Sequence↗
PubMed1991

Identification of membrane antigens in granulocytes and colonic carcinoma cells by a monoclonal antibody specific for biliary glycoprotein, a member of the carcinoembryonic antigen family.

The gene coding for 'biliary glycoprotein (BGP)' is a member of the carcinoembryonic antigen (CEA) gene family. A monoclonal antibody (MAb) was induced against a BGP-preparation isolated from human bile. The antibody did not crossreact with the carcinoembryonic antigen (CEA) and different non-specific crossreacting antigens. The anti-BGP MAb was used to identify BGP-related antigens in membrane extracts from granulocytes and the colonic carcinoma cell line HT-29. In granulocyte membranes, a single antigen of Mr 160,000 was bound. In membranes from HT-29 cells, a main antigen of Mr 85,000 was present. At high antigen concentration, an additional antigen of Mr 115,000 was identified. Since several transcripts of the BGP gene have been identified, the different BGP related antigens are probably products of alternatively spliced mRNAs.

Antibodies, Monoclonal↗
PubMed1991

The putative role of members of the CEA-gene family (CEA, NCA an BGP) as ligands for the bacterial colonization of different human epithelial tissues.

Immobilized purified CEA (carcinoembryonic antigen), NCA (non-specific crossreacting antigen) and BGP I (biliary glycoprotein I) bind strains of E. coli (including EPEC) and some Salmonella species (including S. typhi, S. paratyphi A + B and S. java) while Shigella-, Yersinia- and Bacteroides- strains showed no adhesion. The binding was of high avidity, heat sensitive, dose dependent, saturable and nearly completely abolished in the presence of 10 mM alpha-methylmannoside. From inhibition studies with aromatic mannose compounds, it was suggested that in contrast to Salmonella strains E. coli strains exhibit a higher hydrophobicity in the binding region adjacent to the CEA-, NCA- and BGP-binding site. By further inhibition experiments it could be demonstrated that E. coli and Salmonella strains bind to high-mannose type oligosaccharides of these molecules via lectins on bacterial type I fimbriae. We conclude that the expression of products of this gene family on different human epithelial cells (colon-, bile canaliculi, uroepithel etc.) may function as ligands for bacterial colonization of epithelial tissues.

Antigens, CD↗
PubMed1991

Binding of Escherichia coli and Salmonella strains to members of the carcinoembryonic antigen family: differential binding inhibition by aromatic alpha-glycosides of mannose.

Various Escherichia coli and Salmonella strains bound to glycoproteins of the family of carcinoembryonic antigens (CEA). As judged from plateau regions of the binding curves, CEA, nonspecific cross-reacting antigen of Mr 55,000 (NCA-55), and biliary glycoprotein of Mr 85,000 (BGP-85) showed similar binding activities. The binding to ovalbumin was significantly lower and the binding to fetuin was insignificant under identical experimental conditions. The binding of E. coli and S. typhi to the different glycoproteins was similar as judged from the binding curves. In comparison with alpha-methyl-D-mannopyranoside, aromatic alpha-glycosides of mannose were more potent binding inhibitors of E. coli but not of salmonellae to CEA and NCA-55. These results are similar to those previously obtained with intestinal epithelial cells and yeast cells (N. Firon, S. Ashkenazi, D. Mirelman, I. Ofek, and N. Sharon, Infect. Immun. 55:472-476, 1987). The binding of E. coli to CEA was inhibited by purified type 1 fimbriae. On the basis of the distribution of CEA-like glycoproteins in tissues and body fluids, the results indicate that glycoproteins of the CEA family may be involved in the recognition of bacteria and the regulation of bacterial colonization.

Animals↗
PubMed1991

Binding of nonspecific cross-reacting antigen, a granulocyte membrane glycoprotein, to Escherichia coli expressing type 1 fimbriae.

Nonspecific cross-reacting antigen (NCA) is a well-characterized membrane glycoprotein on granulocytes, macrophages, and lung epithelium. Structural studies at the protein and genomic levels have revealed that NCA is a member of the immunoglobulin supergene family, and hybridization studies showed that the transcript level of NCA is induced by treatment with gamma interferon. These studies, as well as the expression of NCA on granulocytes, suggest a role for NCA in immune response. For a first step in studying this possible role, we have examined the binding of two glycoforms of NCA designated NCA-50 (Mr, 50,000) and TEX-75 (Mr, 75,000). Here we report the results from binding assays which demonstrate carbohydrate-mediated binding of Escherichia coli expressing type 1 fimbriae and of isolated type 1 fimbriae to NCA-50. TEX-75 did not bind to the purified fimbriae but bound slightly to the bacterial strain. Inhibition studies showed that the binding to NCA-50 involved interaction of mannose moieties on NCA-50 and lectins on the fimbriae. The binding of NCA-50 to bacterial fimbriae was confirmed by electron microscopy studies, using immunolabeling techniques. In addition, we show that the surface expression of NCA-50 (and presumably of other NCA species) on isolated polymorphonuclear leukocytes is increased following activation with the bacterial peptide formylmethionyl-leucyl-phenylalanine, consistent with a role for NCA in immune response.

Antigens, Neoplasm↗
PubMed1990

Escherichia coli of human origin binds to carcinoembryonic antigen (CEA) and non-specific crossreacting antigen (NCA).

Immobilized carcinoembryonic antigen (CEA) and non-specific crossreacting antigen (NCA) bound 3 strains of E. coli of human origin. The binding was dose dependent, saturable, and of high avidity. Binding of the bacteria to CEA and NCA was completely abolished in the presence of 10 mM alpha-methyl D-mannopyranoside. Bacteria did not bind to concanavalin A. In addition, binding to deglycosylated CEA was either absent or significantly reduced. These findings indicate that the E. coli strains bind to D-mannosyl residues in CEA and NCA. Considering the tissue distribution of CEA (brush border of colonic epithelium) and NCA (granulocytes), these glycoproteins may be involved in the recognition of bacteria.

Antigens, Neoplasm↗
PubMed1989

Determination of epitope specificities and affinities of monoclonal antibodies in solution phase using biotin-labeled carcinoembryonic antigen and avidin as precipitating agent.

A generally applicable method for the selective precipitation of antigens from solution phase is presented. The method is based on the rapid and quantitative binding of avidin to biotin-labeled antigen in the presence of biotinylated carrier protein. Using carcinoembryonic antigen (CEA) as a model antigen biotin-labeled CEA was precipitated to greater than 97% at amounts of up to 5000 ng/tube. The method was applied to the determination of epitope specificities and affinities of monoclonal anti-CEA antibodies in solution phase. Non-specific precipitation did not exceed 2% for radiolabeled Fab fragments and 6% for radiolabeled IgG. Because antigen-antibody binding takes place in solution, artifacts introduced by the immobilization of reactants to solid supports are omitted.

Animals↗
PubMed1989

Characterization of biotin-labeled proteoglycans by electrophoretic separation on minigels and blotting onto nylon membranes prior and after enzymatic digestion.

Biotinylated proteoglycans were separated by sodium dodecyl sulfate electrophoresis prior and after enzymatic digestion by glycan-specific enzymes using polyacrylamide minigels. The biotin-labeled compounds were blotted onto nylon membranes either by electrophoresis or by diffusion and detected by avidin-enzyme conjugates. The method allows the nonisotopic detection of native proteoglycans and core proteins. Proteoglycans can be visualized at protein amounts as low as 0.7 ng per lane. In comparison with sensitive protein stains, compounds of enzyme preparations do not interfere with bands corresponding to core proteins. Electrophoresis, blotting, and staining of up to 12 samples per gel are accomplished in less than 3 h.

Amino Acids↗
PubMed1989

Activation of the phosphatidylinositol cycle in spreading cells.

Metabolites of the phosphatidylinositol cycle were analyzed in BHK-21 (C13) cells spreading on fibronectin-coated culture plates in comparison with attached nonspreading cells 45 min after plating. Among the water-soluble metabolites (glycerophosphoinositol, inositol, inositol monophosphate, inositol bisphosphate, inositol trisphosphate, and inositol tetrakisphosphate), significant elevations were found for inositol monophosphate, inositol bisphosphate, and inositol tetrakisphosphate. In the lipid fraction, phosphatidylinositol 4-monophosphate and phosphatidylinositol 4,5-bisphosphate were significantly elevated. The activation of the phosphatidylinositol cycle in spreading versus nonspreading attached BHK-21 (C13) cells may be involved in the permissive effect of the extracellular matrix on cell proliferation.

Animals↗