Tumor antigens: Biological activity of components of soluble antigens of MTV-induced mammary tumors

Summary Antigens (AMMT) of MTV-induced mammary tumors of BALB/cfC3H CRGL mice were solubilized by treatment of homogenates of the tumor with 1 M perchloric acid. The soluble antigens exhibited biological activity by their ability to induce DNA synthesis in spleen cells of mice bearing syngeneic transplants of the tumor. AMMT-induced DNA synthesis, however, was abrogated by serum from tumor-bearing mice. AMMT neutralized complement-dependent rabbit anti-ML(r) antibody activity and intracytoplasmic fixed cell immunofluorescent activity of the same antibodies. One component of AMMT has an electrophoretic mobility pattern identical to that of ML(r) antigen. Thus, AMMT may share antigenicity with MTV-specific antigens.     

The use of Fab-masking antigens to enhance the activity of immobilized antibodies

This study demonstrates that masking the Feb regions of a monoclonal antibody (Mab) with synthetic antigens prior to covalent immobilization efficiency. Water-soluble adducts of poly(2-methyloxazoline) polymers and a syntheticpeptide epitope for the Mab were constructed. These synthetic antigens are referred to as Fab-masking antigents (FMAs). The antibody used in this study is a Ca(2+)-dependent murine monoclonal lgG directed against the plasma protein, human protein C (hPC). The FMAs were pre-equilibrated with Mab in the presence of calcium prior to immobilization and were then removed by EDTA, which destabilized the FMA-Mab complexes. The antigen binding efficiency and accessibility of the Fab domain of the immobilized antibody was significantly increased for Mab immobilized in the presence of FMA relative to those Mab immobilized without FMA. The increase in binding efficiency was most pronounced for the largest FMA employed. No appreciable differences were detected in the avidity of hPC-Mab complexes formed by immunosorbents produced by either masked or unmaked antibody. These results provide evidence that orientgation may play an important role in the binding activity of immobilized antibodies.     

A comparison of bovine lymphocyte antigens

In Canberra, 31 antigens have been described on the surface of bovine lymphocytes. Seven antigens are subgroups of other antigens. Eleven antigens are similar to the eleven antigens which have been described in Melbourne. Fourteen antigens are similar to twelve international-workshop antigens and two European-workshop antigens.     

Immunochemical identification of multiple cell surface antigens appearing during specific stages of mouse spermatogenesis

Cell surface antigens that appear in a defined temporal sequence during mouse spermatogenesis were previously detected serologically, but not identified biochemically, with four heterologous antibodies prepared against purified populations of pachytene spermatocytes (AP), round spermatids (ARS), vas deferens spermatozoa (AVDS), and mixed seminiferous cells (ASC) [Millette and Bellvé, J Cell Biol 74:86–97, 1977]. These antigens have now been identified immunochemically on nitrocellulose blots from SDS polyacrylamide gels. Three antisera (AP, ARS, and ASC) recognize a similar subset of determinants on one-dimensional immunoblots of germ cells and plasma membranes prepared from a mixed population of late spermatogenic cells. Comparisons of minor bands to reveal differences among these antisera. AVDS exhibits the least complex binding pattern. The results indicate that at least ten surface constituents appear during the pachytene stage of meiosis, coincident with a period of maximal RNA and protein synthesis [Monesi, Exp Cell Res 39:197–224, 1965]. Furthermore, two-dimensional immunoblot comparisons of plasma membranes isolated from pachytene spermatocytes and round spermatids reveal differences between surface determinants detectable at these two spermatogenic stages. For example, ASC recognizes two newly described proteins that are restricted to pachytene spermatocytes (? M_r 57,000, pI 6.45) and to round spermatids (? M_r 39,500, pI 4.85), respectively.     

Detection of cross reactive antigens between Pestalotiopsis theae and tea leaves and their cellular location

Among the 12 varieties of tea tested against three isolates of Pestalotiopsis theae, causal agent of grey blight disease, Teen Ali-17/1/54 and TV-23 were found to be highly susceptible while CP-1 and TV-26 were resistant under identical conditions. Leaf antigens were prepared from all the tea varieties, three isolates of P. theae and a non-pathogen of tea (Bipolaris tetramera). Polyclonal antisera were raised against mycelial suspensions of P. theae (isolate Pt-2) and leaf antigens of Teen Ali-17/1/54 and CP-1. These were compared an immunodiffusion test and enzyme-linked immunosorbent assay to detect cross reactive antigens (CRA) shared the host and the parasite. CRA were found among the susceptible varieties and isolates of P. theae (Pt-1, 2 and 3). Such antigens were not detected between isolates of P. theae and resistant varieties, B. tetramera and tea varieties or isolates of P. theae. Indirect staining of antibodies using fluorescein isothiocyanate (FITC) indicated that in cross sections of tea leaves, the CRA was concentrated in the epidermal cells and mesophyll tissues. CRA was present in the young hyphal tips of the mycelia and on the setulae and appendages of the conidia of P. theae.     

Surface antigens of Biomphalaria glabrata (Gastropoda) hemocytes: evidence for linkage-independence of some hemolymph-like surface antigens and Con A receptor-bearing macromolecules

The structural relationship between hemolymph-like surface antigens and concanavalin A (Con A)-reactive macromolecules on circulating hemocytes of Biomphalaria glabrata was assessed using a double-ligand labeling method. It was determined that Con A-induced clearance of its own receptor complexes resulted in a significant reduction, but of complete elimination, of hemolymph-like antigens, i.e., antigens cross-reactive with an anti-B. glabrata hemolymph antiserum, suggesting the presence of at least two separate antigenic populations with anti-hemolymph reactivity; one group structurally linked and another group structurally independent of Con A-binding membrane components. The latter group of surface antigens appears to be (1) chemically related to the higher-molecular-weight hemolymph components, most probably hemoglobin, (2) Pronase resistant, and (3) partially composed of a subpopulation of cryptic (hidden) cross-reacting antigens uncovered at the cell surface as a consequence of Con A-receptor clearance. Results of this study demonstrate not only that individual hemocytes of B. glabrata may possess different populations of hemolymph-like antigens, but also that the interaction between some membrane components and appropriate ligands (e.g., carbohydrate-binding lectins) could result in a modulation in expression of other groups of surface antigens.     

Species-specific and cross-reactive antigens of Entamoeba

Specific and cross-reactive antigens were defined in four species of Entamoeba: invadens, moshkovskii, Laredo and histolytica strains HM1, HM3, HM38 and HK9. Among these species extensive common reactivities were observed by immunoblot. Eight E. histolytica antigenic markers were revealed after blocking common specificities with antigens of other Entamoeba species. A monoclonal antibody (mAb) defined two protein markers of E. histolytica, M, 29 and 25 kDa. The four strains of E. histolytica, which varied in virulence as determined by the development of liver abscesses in hamsters, showed the same antigenic patterns with the mAb and with polyclonal antibodies.     

Quantitation of human mammary epithelial antigens in cells cultured from normal and cancerous breast tissues

Summary A sensitive radioimmunoassay technique was developed to quantitatite the level of human breast celltype specific antigens on cells from normal breast and from various established cell lines of breast and nonbreast origins. Polyacrylamide gel electrophoresis revealed four major proteinaceous components (150,000; 75,000; 60,000; and 48,000) in human milk fat globule membranes that were used to immunize rabbits in order to elicit antimammary epithelial cell antibody. Antisera obtained were rendered specific by abosorptions and were able to recognize three specific mammary epithelial components of the breast epithelial cell. Human mammary epithelial (HME) antigen expression was highest (1290 ng/10⁶ cells) in normal breast epithelial cells from primary cultures of normal breasts. Lower levels (range: 955 to 330 ng/10⁶ cells) were found in breast epithelial cells from cell lines established from cancerous breast tissue. Cells of nonbreast origins as well as fibroblasts from breast gave much lower values (less than 30 ng/10⁶ cells). On treatment, with trypsin, of two breast epithelial cell lines (MDA-MB-157 and MCF-7) 80 to 85% of their HME antigen expression was lost, suggesting that a majority of these breast antigens reside on the cell surface. This work was Supported by Grant PTD-99 from the American Cancer Society, Grant CA19455 and CA20286 from the National Cancer Institute, and Biomedical Research Support Grant RR05467 from the National Institutes of Health. Most cells used in the present study were produced with support from National Cancer Institute Contract Y01-CP8-0500, Biological Carcinogenesis Branch, Division of Cancer Cause and Prevention, under the auspices of the Office of Naval Research and the Regents of the University of California.     

Immunoregulatory activity of the lungs in relation to corpuscular and soluble antigens

The immunoregulatory activity of the lungs in normal Wistar rats has been evaluated by difference between primary and secondary immune response to the same dose of the antigen introduced into the respiratory tract or intravenously. As shown in this investigation, intratracheal immunization with corpuscular antigens is accompanied by faintly pronounced antibody formation and a high degree of delayed hypersensitivity, while the introduction of soluble antigens into the respiratory tract leads to the active production of antibodies. The immunoregulatory activity of the lungs is T-dependent. The preliminary introduction of corpuscular or soluble antigen into the respiratory tract is accompanied by the formation of the local mechanism in the lungs for suppressing immune response to the subsequent intratracheal immunization.     

Synthesis and release of proteins homologous to excretory-secretory antigens during embryogenesis ofSetaria digitata

Localization of antigens homologous to excretory-secretory proteins in developing embryos ofSetaria digitata has been carried out by indirect fluorescent antibody test, [¹⁴C] labelling studies and Western blotting. Indirect fluorescent antibody test showed binding of excretory-secretory antibodies at perivitelline space. The fluorescent antibody binding was almost absent at small morulae stage and increasing in intensity in the successive developmental stages with maximum at coiled microfilaria stage. Hatched microfilaria did not show the presence of antigens by immunofluorescence. Immuno-complex of excretory-secretory antiserum against “amniotic fluid” collected from developing embryos ofSetaria digitata labelled with [¹⁴C] amino acids showed highest radioactivity at coiled and tadpole stages and differed significantly from small morulae, big morulae and hatched microfilaria. Immunoblot analysis of amniotic fluid showed two proteins, 16.5 and 11 kDa, to be highly antigenic. The antigenic protein (11 kDa) content as seen by immuno blotting increased during embryogenesis and decreased at the stage of hatching.     

Surface antigens of Biomphalaria glabrata (Gastropoda) hemocytes: occurrence of membrane-associated hemolymph-like factors antigenically related to snail hemoglobin

A polyvalent antiserum (anti-H_PR) generated in rabbits to cell-free hemolymph from a PR albino (M-line) stock of snail, Biomphalaria glabrata, was employed as a membrane probe to determine if antigens related to snail hemolymph were associated with the surface membranes of phosphate-buffered saline (PBS) washed hemocytes from a schistosome-susceptible (PR albino) and refractory (10-R2) stock of B. glabrata. Immunofluorescent and immunoelectron microscopical analyses revealed a strong cross-reactivity between anti-H_PR antibodies and hemocytes from both PR albino and 10-R2 snails indicating the presence of surface-associated hemolymph or hemolymph-like antigens. Hemoglobin isolated from PR albino B. glabrata hemolymph competitively inhibited the binding of anti-H_PR to hemocytes suggesting that cross-reactive membrane components were, at least in part, antigenically related to snail hemoglobin. Antigens reactive with antihemolymph antibodies also were resistant to protease treatment. No antigenic differences between PR albino and 10-R2 snail hemocytes could be detected due to the heterospecific nature of the probe antiserum, however, it is believed that the major cross-reactive membrane components, e.g., hemoglobin-like determinants, are shared in common by hemocytes of both snail stocks.     

Specifically induced suppression of T cell and NK-like cytolytic activity by ingested soluble antigens

The effect of feeding xenoserum (xs) on cytolytic cell activity induced by parenteral injection was examined in C3H/N mice. Spleen cells were cultured with xs and then assayed for cytolytic activity against a panel of 51Cr-labeled YAC-1, AKR-A, or P815 target cells. Prior feeding resulted in significant suppression of responses stimulated by injection and culture. The induction of these responses was antigen specific for xs whereas the effector stage represented polyclonal activation of cytolytic cells. Some effector cells were lysed by either anti-Lyt 2 or anti-NK- 1.2 and complement and some were blocked by anti-Lyt 2 or anti-T200 in the cytotoxicity assay. Thus, both cytolytic T and NK-like cells were suppressed by antigen feeding. Activity of TH cell-derived factors which enhance cytolytic activity ("promoter" factor, interferon, and interleukin 2) also was diminished in culture supernatants of cells from mice fed soluble antigens. The conclusion that polyclonal cytolytic responses induced by soluble antigen can be regulated by prior enteric stimulation is made.     

The stage-specific expression of TEC-1, -2, -3, and -4 antigens on bovine preimplantation embryos

The preimplantation developmental period is associated with constant changes within the embryo, and some of these changes are apparent on the embryo cell surface. For example, during transition from maternal to embryonic genome control and the compaction and differentiation of embryonic cells, the cell surface undergoes morphologic alterations that reflect changes in gene control. In order to gain insight into the events occurring during embryonic development and cellular differentiation, monoclonal antibodies specific for cell surface antigens (TEC antigens) of embryonic cells have been generated previously and shown to recognise either the carbohydrate moiety of embryoglycan or a developmentally regulated protein epitope. The TEC antigens have been identified on mouse preimplantation embryos, and their expression is specific to particular developmental stages. To determine whether these antigens are conserved in higher mammals, we examined the expression of four TEC antigens (TEC-1 to TEC-4) on in vitro–derived bovine and murine embryos during the preimplantation stage of development. It was found that bovine oocytes and embryos derived from in vitro maturation (IVM) and in vitro fertilisation (IVF) showed stage-specific expression of each of the TEC antigens investigated, with the pattern of expression overlapping but not identical to that seen in the mouse. Immunoprecipitation together with Western blot analysis showed that the TEC monoclonal antibodies recognised a single glycoprotein band with an apparent molecular weight of 70 kDa. Confocal microscopy of immunofluorescence staining of the bovine cells showed this protein to be located on the cell surface. The apparent negative expression of these TEC antigens by immunohistochemistry and immunoprecipitation at particular stages of development appears to be due to the epitopes being inaccessible to the TEC antibodies, since Western blotting revealed the TEC antigens to be present at all stages of development examined. Antibodies identifying stage-specific antigens will provide useful markers to characterise early embryonic cells, monitor normal embryonic development in vitro, and identify cell surface structures having a function in cell-cell interactions during embryogenesis and differentiation. Mol. Reprod. Dev. 49:19–28, 1998. © 1998 Wiley-Liss, Inc.     

BoLA antigens are associated with increased frequency of persistent lymphocytosis in bovine leukaemia virus infected cattle and with increased incidence of antibodies to bovine leukaemia virus

The association between bovine major histocompatibility system (BoLA) type and persistent lymphocytosis in cattle with antibodies to bovine leukaemia virus was examined by comparing antigen frequencies in cattle with persistent lymphocytosis to controls matched for age, sex, breed and presence of antibodies to BLV. The cattle came from nine dairy herds in south-east Queensland, Australia; six herds were Australian Illawarra Shorthorn (AIS), two herds were Jersey and one herd was Friesian. Antigen W6 and Eu28R were more common in cattle with persistent lymphocytosis than in controls. Antigen W8 was less common in AIS cattle with persistent lymphocytosis. A study of 24 offspring from one sire, heterozygous for W10 and Eu28R, showed that offspring inheriting Eu28R from the sire were significantly more likely to have antibodies to BLV than offspring inheriting the opposing W10 haplotype.     

Isolation and functional characterization of Plasmodium falciparum merozoite antigens

Merozoite surface proteins are thought to play an important role during the invasion of red blood cells by merozoites. In this article the strategies for the chromatographic isolation and for the functional and molecular characterisation of isolated antigens from freshly harvested Plasmodium falciparum merozoites from cultures are described.     

Immunolocalization of theca antigens on Alexandrium catenella and their use to isolate cells from fixed phytoplankton samples

Murine monoclonal antibodies were generated and selected for their ability to specifically recognize theca antigens of Alexandrium catenella (Whedon et Kafoid) Balech cells. The specificity of the monoclonal antibodies for theca antigens was shown by indirect immunofluorescence and by confocal microscopic analysis. Using these antibodies we demonstrate, for the first time, the presence of different theca antigens on the cell surface. The fluorescent signal analysis suggests that these antigens differ in their distribution and quantities in the theca.Also, using the antibodies we developed a rapid method to isolate A. catenella cells from a lugol-fixed phytoplanktonic sample. The method uses a mixture of different monoclonal antibodies to bind the cells, which then are pulled off from the sample by means of a second anti-mouse antibody coupled to 0.8 μm magnetic beads.     

Surface antigens on cercariae,schistosomula and adult worms of Schistosoma mansoni

Shah J. and Ramasamy R. 1982. Surface antigens on cercariae, schistosomula and adult worms of Schistosoma mansoni. International Journal for Parasitology12: 451–461. The surface protein antigens of Schistosoma mansoni were radiolabelled by lactoperoxidase catalysed I¹²⁵-iodination and analysed by immune-precipitation and polyacrylamide gel electrophoresis. The results showed that regularly labelled surface antigens of mol. wts >150,000, 78,000, 45,000 and 22,000 were present on adult worms. Common surface antigens were observed on the cercariae, schistosomula and adult worms. It is suggested that surface antigens released from living adult worms can sensitise a host to react against the invading schistosomula of a secondary infection. However, the failure to vaccinate mice using material containing adult worm surface antigens suggests that the induction of protective immunity is a complex phenomenon.     

Relationships of Phenylobacterium immobile and purple nonsulfur bacteria on the basis of surface antigens

Abstract Indirect immunofluorescence tests with antisera against whole cells of Phenylobacterium immobile strains revealed a serological relationship to Pseudomonas vesicularis, Aquaspirillum itersonii and Rhodospirillum rubrum , three members of the purple nonsulfur bacteria (group I) and also to Gluconobacter oxydans and Azotobacter vinelandii . Antisera against whole cells of Gluconobacter oxydans and Pseudomonas vesicularis reacted positively with the Phenylobacterium immobile strains, tested. Furthermore, a serological relationship of Gluconobacter oxydans to Acetobacter aceti , and of Pseudomonas vesicularis to Pseudomonas diminuta and Aquaspirillum itersonii could be demonstrated.     

Identification of capillaries in sections from skeletal muscle by use of lectins and monoclonal antibodies reacting with histo-blood group ABH antigens

This study was performed to evaluate the application of different lectins and monoclonal antibodies against ABH antigens to detect and characterize carbohydrate structures in capillaries of skeletal muscle from humans and laboratory animals. Blood group specific lectins (Griffonia simplicifolia, Griffonia simplicifolia isolectin B4,Lotus tetragonlobus, Ulex europaeus, andDolichos biflorus) and monoclonal antibodies reacting with histo-blood group carbohydrate antigens belonging to type 1 (Le^a) and type 2 (H, A and Le^y) chains were used as histological markers for capillaries in sections from skeletal muscle. The material consisted of 20 human masseter muscle biopsies from individuals with known blood types: (eight blood group O, nine blood group A, two blood group B, and one blood group AB) and masseter muscles specimens from different laboratory animals (mouse, rat, rabbit, cat, dog, pig, cow, and macaca monkey). Unfixed sections and an avidin alkaline phosphatase method were used to visualize the specific reaction.Ulex lectin stained capillaries in all human biopsies either strongly or moderately. Strong muscle capillary reaction was observed in biopsies from O, B and AB individuals while capillaries from A individuals were only moderately stained.Griffonia simplicifolia marked capillaries in A, B, and AB individuals andGriffonia simplicifolia isolectin B4 stained capillaries in muscle biopsies from B and AB donors.Dolichos biflorus was a weak marker of muscle capillaries from A individuals. Only capillaries from O individuals were stained with the antibody against H type 2. Capillary reaction was not observed with the other antibodies used.Girffonia simplicifolia was an excellent marker for capillaries in mouse muscle whileGriffonia simplicifolia isolectin B4 is recommended for rat muscles. Periodic acid treatment and subsequentLotus tetragonolobus staining is suitable to visualize capillaries in mouse, rat and pig muscle. Using a sensitive histochemical technique for staining with lectins and monoclonal antibodies reacting with blood group related antigens the microvascular density in human skeletal muscle may be estimated. Further, the carbohydrate compounds in the muscle capillaries reflect the individual blood type. A selection of lectins is suitable for demonstration of capillaries in animal skeletal muscle.