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Sample records for distinct antigenic determinants

  1. Antigenic determinants and functional domains in core antigen and e antigen from hepatitis B virus

    International Nuclear Information System (INIS)

    Salfeld, J.; Pfaff, E.; Noah, M.; Schaller, H.

    1989-01-01

    The precore/core gene of hepatitis B virus directs the synthesis of two polypeptides, the 21-kilodalton subunit (p21c) forming the viral nucleocapsid (serologically defined as core antigen [HBcAg]) and a secreted processed protein (p17e, serologically defined as HBe antigen [HBeAg]). Although most of their primary amino acid sequences are identical, HBcAg and HBeAg display different antigenic properties that are widely used in hepatitis B virus diagnosis. To locate and to characterize the corresponding determinants, segments of the core gene were expressed in Escherichia coli and probed with a panel of polyclonal or monoclonal antibodies in radioimmunoassays or enzyme-linked immunosorbent assays, Western blots, and competition assays. Three distinct major determinants were characterized. It is postulated that HBcAg and HBeAg share common basic three-dimensional structure exposing the common linear determinant HBe1 but that they differ in the presentation of two conformational determinants that are either introduced (HBc) or masked (HBe2) in the assembled core. The simultaneous presentation of HBe1 and HBc, two distinctly different antigenic determinants with overlapping amino acid sequences, is interpreted to indicate the presence of slightly differently folded, stable conformational states of p21c in the hepatitis virus nucleocapsid

  2. Demonstration of two distinct antigenic determinants on hepatitis B e antigen by monoclonal antibodies

    International Nuclear Information System (INIS)

    Imai, M.; Nomura, M.; Gotanda, T.; Sano, T.; Tachibana, K.; Miyamoto, H.; Takahashi, K.; Toyama, S.; Miyakawa, Y.; Mayumi, M.

    1982-01-01

    Mice were immunized against hepatitis B e antigen (HBeAg) isolated from sera of asymptomatic carriers of hepatitis B virus. Their spleen cells were fused with mouse myeloma (NS-1) cells, and 5 clones of hybridoma cells secreting antibody against HBeAg (anti-HBe) were isolated. For the production of anti-HBe in large scale, cells were cultivated both in vitro and in the peritoneal cavity of ascitic mice. Although monoclonal antibodies produced by these clones showed a strong reactivity of anti-HBe in hemagglutination tests, individual monoclonal anti-HBe did not reveal any precipitin line in immunodiffusion. When 2 of the 5 monoclonal antibodies were mixed together, however, some combinations showed a precipitin line against HBeAg, whereas others did not. Utilizing solid-phase radioimmunoassay involving a number of combinations of monoclonal antibodies used for solid-phase and radiolabeling, the 5 antibodies were classified into 2 groups. Three of the anti-HBe antibodies were found to be directed to 1 determinant of HBeAg (determinant a); the remaining 2 to the other determinant (determinant b). Determinants a and b were detected on HBeAg in the serum, as well as on the polypeptide of 19,000 daltons (P19) derived from the nucleocapsid of hepatitis B virus. Monoclonal anti-HBe antibodies with different specificities may provide useful tools in delineating the antigenic structure of HBeAg and also in evaluating immune responses of the host directed to its subdeterminants

  3. Antigen presentation by resting B cells. Radiosensitivity of the antigen-presentation function and two distinct pathways of T cell activation

    International Nuclear Information System (INIS)

    Ashwell, J.D.; DeFranco, A.L.; Paul, W.E.; Schwartz, R.H.

    1984-01-01

    In this report we have examined the ability of small resting B cells to act as antigen-presenting cells (APC) to antigen-specific MHC-restricted T cells as assessed by either T cell proliferation or T cell-dependent B cell stimulation. We found that 10 of 14 in vitro antigen-specific MHC-restricted T cell clones and lines and three of four T cell hybridomas could be induced to either proliferate or secrete IL-2 in the presence of lightly irradiated (1,000 rads) purified B cells and the appropriate foreign antigen. All T cell lines and hybridomas were stimulated to proliferate or make IL-2 by macrophage- and dendritic cell-enriched populations and all T cells tested except one hybridoma caused B cell activation when stimulated with B cells as APC. Furthermore, lightly irradiated, highly purified syngeneic B cells were as potent a source of APC for inducing B cell activation as were low density dendritic and macrophage-enriched cells. Lymph node T cells freshly taken from antigen-primed animals were also found to proliferate when cultured with purified B cells and the appropriate antigen. This APC function was easily measured when the cells were irradiated with 1,000 rads, but was greatly diminished or absent when they were irradiated with 3,300 rads. In addition, this radiosensitivity allowed us to easily distinguish B cell antigen presentation from presentation by the dendritic cell and macrophage, as the latter was resistant to 3,300 rads. Finally, one T cell clone that failed to proliferate when B cells were used as APC was able to recruit allogeneic B cells to proliferate in the presence of syngeneic B cells and the appropriate antigen. This result suggests that there are at least two distinct pathways of activation in T cells, one that leads to T cell proliferation and one that leads to the secretion of B cell recruitment factor(s)

  4. [Radiocompetitive method of H antigen determination].

    Science.gov (United States)

    Semenova, G B; Sokolov, Ia A; Liashenko, V A

    1978-06-01

    The authors describe a radiocompetitive method of H-d-monomere determination with the sensitivity of 2 ng/ml in double antibodies modification; this method was used for comparing the immunological affinity of the affiliated H-antigens. A difference between the immunological affinity to the antibodies in a monomere, polymere and the flagellum was shown.

  5. The neural cell adhesion molecule L1 is distinct from the N-CAM related group of surface antigens BSP-2 and D2

    DEFF Research Database (Denmark)

    Faissner, A; Kruse, J; Goridis, C

    1984-01-01

    The neural cell adhesion molecule L1 and the group of N-CAM related molecules, BSP-2 and D2 antigen, are immunochemically distinct molecular species. The two groups of surface molecules are also functionally distinct entities, since inhibition of Ca2+-independent adhesion among early post-natal m...

  6. Mosaic VSGs and the scale of Trypanosoma brucei antigenic variation.

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    James P J Hall

    Full Text Available A main determinant of prolonged Trypanosoma brucei infection and transmission and success of the parasite is the interplay between host acquired immunity and antigenic variation of the parasite variant surface glycoprotein (VSG coat. About 0.1% of trypanosome divisions produce a switch to a different VSG through differential expression of an archive of hundreds of silent VSG genes and pseudogenes, but the patterns and extent of the trypanosome diversity phenotype, particularly in chronic infection, are unclear. We applied longitudinal VSG cDNA sequencing to estimate variant richness and test whether pseudogenes contribute to antigenic variation. We show that individual growth peaks can contain at least 15 distinct variants, are estimated computationally to comprise many more, and that antigenically distinct 'mosaic' VSGs arise from segmental gene conversion between donor VSG genes or pseudogenes. The potential for trypanosome antigenic variation is probably much greater than VSG archive size; mosaic VSGs are core to antigenic variation and chronic infection.

  7. Determination of Diagnostic Antigens in Cattle Amphistomiasis Using Western Blotting

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    A Halajian

    2009-05-01

    Full Text Available "nBackground: Mixed infection with amphistomes seems common in native cattle of Iran. The aim of this study was to determine diagnostic antigens in cattle mixed amphistomiasis."nMethods: Specific antigens of Cotylophoron cotylophorum, Gastrothylax crumenifer and Paramphisto­mum cervi (mixed infection, the most common species, were collected from cattle was deter­mined. Adult trematodes were collected from the rumen of naturally infected cattle at meat inspec­tion. After their homogenization and centrifugation, somatic antigens were prepared and ana­lyzed by SDS-PAGE. Specific antigens were determinated by western blot with homologous and heterolo­gous sera. SDS-PAGE of whole worms extract was performed at different concentrations and subse­quent gels staining. Immunoblotting analysis using sera from cattle naturally infected with am­phistomes, Dicrocoelium dendriticum, Fasciola spp. and hydatid cyst was performed."nResults: Electrophorese analysis of somatic antigens revealed the presence of 10 and 21 protein bands at 4 µgr/ml and 8 µgr/ml with molecular weights ranging from 25-120 and 25-150 kDa, respectively. The best result was taken at 8 mg/ml concentration. Although western blot of these proteins demon­strate 5 major antigenic polypeptides ranging from 50 to 100 kDa which were recognized by serum of cat­tle naturally infected with mixed amphistomes.

  8. Antigenic relatedness of primate procollagens as determined by a competitive radioimmunoassay

    International Nuclear Information System (INIS)

    Taubman, M.B.; Goldberg, B.

    1978-01-01

    A radioimmunoassay specific for the nonhelical carboxy terminal portion of human type I procollagen was used to study the antigenic relatedness of primate procollagens. The assay identified reactive antigen in primate sera and in the media of primate fibroblast cultures. The displacement curves generated in the assay indicated that human and ape type I procollagens have antigenically identical carboxy terminal determinants which are partially cross-reactive with those from Old and New World monkeys. (author)

  9. Use of immunoradiometric analysis to determine Rickettsia antigens in cell cultures and chick embryos

    Energy Technology Data Exchange (ETDEWEB)

    Prozorovskiy, S.V.; Alekseeva, N.V.; Knyazeva, E.N.; Ignatovich, V.F.; Barkhatova, O.T.

    1984-02-01

    A modification of an immunoradiometric analysis to determine Rickettsia antigens in various biological substrates was studied, using rickettsious diagnostricums, egg and cell cultures of Rickettsia. The method was highly sensitive for the determination of minimal quantities of antigens in these substrates. The method appears to be promising for studies related to the detection of microorganisms and their antigens. 5 references.

  10. Critical study and applications of the radioimmunological determination of carcinoembryonic antigen

    International Nuclear Information System (INIS)

    Troupel, Solange.

    1974-01-01

    This paper outlines our research on the development of a radioimmunological method to determine the carcinoembryonic antigen of the digestive system (ACE). The carcinoembryonic antigen is defined and situated in the framework of antigens associated with human tumours. The general principles of the radioimmunological determination are then reviewed. A detailed technical study is devoted to each of the elements involved in the reaction and to the working conditions of each method tried. A labelling procedure and a radioactive protein separation method have been worked out, guaranteeing a high specific radioactivity consistent with a good immunoreactivity. The period of effectiveness of this protein has also been determined, taking account of its deiodination. The antiserum is a very important factor in the sensitivity of the measurement. A ewe antiserum of good antibody content and volume yield was chosen, its disadvantage being the length of the determination imposed by the 48 hour preincubation time. Ammonium sulphate precipitation and double antibody techniques were used for the labelled antigen-antibody separation. In seric solution the ammonium sulphate precipitation carries down non-specifically, in the standards, a large amount of labelled antigen. This disadvantage has been offset by a method of calculation which shows the actual contribution of the labelled complex. The double antibody technique requires a special adjustment to balance quantity of second antiserum and precipitation time. The system sometimes needs an addition of serum from the animal donor of the first antibody in order to obtain an adequate separation. Where techniques are concerned, although the macro-method is suitable for determinations on perchloric extract and is still in common use we prefer to use the one described here under the name of micro-method. Finally the results obtained in experimental and clinical applications are presented [fr

  11. Longitudinal multiparameter single-cell analysis of macaques immunized with pneumococcal protein-conjugated or unconjugated polysaccharide vaccines reveals distinct antigen specific memory B cell repertoires.

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    Bin Jia

    Full Text Available The efficacy of protein-conjugated pneumococcal polysaccharide vaccines has been well characterized for children. The level of protection conferred by unconjugated polysaccharide vaccines remains less clear, particularly for elderly individuals who have had prior antigenic experience through immunization with unconjugated polysaccharide vaccines or natural exposure to Streptococcus pneumoniae.We compared the magnitude, diversity and genetic biases of antigen-specific memory B cells in two groups of adult cynomolgus macaques that were immunized with a 7-valent conjugated vaccine and boosted after five years with either a 13-valent pneumococcal polysaccharide conjugate vaccine (13vPnC or a 23-valent unconjugated pneumococcal polysaccharide vaccine (23vPS using microengraving (a single-cell analysis method and single-cell RT-PCR.Seven days after boosting, the mean frequency of antigen-specific memory B cells was significantly increased in macaques vaccinated with 13vPnC compared to those receiving 23vPS. The 13vPnC-vaccinated macaques also exhibited a more even distribution of antibody specificities to four polysaccharides in the vaccine (PS4, 6B, 14, 23F that were examined. However, single-cell analysis of the antibody variable region sequences from antigen-specific B cells elicited by unconjugated and conjugated vaccines indicated that both the germline gene segments forming the heavy chains and the average lengths of the Complementary Determining Region 3 (CDR3 were similar.Our results confirm that distinctive differences can manifest between antigen-specific memory B cell repertoires in nonhuman primates immunized with conjugated and unconjugated pneumococcal polysaccharide vaccines. The study also supports the notion that the conjugated vaccines have a favorable profile in terms of both the frequency and breadth of the anamnestic response among antigen-specific memory B cells.

  12. Determinants of wheat antigen and fungal alpha-amylase exposure in bakeries.

    Science.gov (United States)

    Burstyn, I; Teschke, K; Bartlett, K; Kennedy, S M

    1998-05-01

    The study's objectives were to measure flour antigen exposure in bakeries and define the determinants of exposure. Ninety-six bakery workers, employed in seven different bakeries, participated in the study. Two side-by-side full-shift inhalable dust samples were obtained from each study participant on a single occasion. The flour antigen exposure was measured as wheat antigen and fungal alpha-amylase content of the water-soluble fraction of inhalable dust, assayed via enzyme-linked immunosorbent assays. During the entire sampling period bakers were observed and information on 14 different tasks was recorded at 15-minute intervals. Other production characteristics were also recorded for each sampling day and used in statistical modeling to identify significant predictors of exposure. The mean alpha-amylase antigen exposure was 22.0 ng/m3 (ranging from below the limit of detection of 0.1 ng/m3 to 307.1 ng/m3) and the mean wheat antigen exposure was 109 micrograms/m3 (ranging from below the limit of detection of 1 microgram/m3 to 1018 micrograms/m3). Regression models that explained 74% of variability in wheat antigen and alpha-amylase antigen exposures were constructed. The models indicated that tasks such as weighing, pouring, and operating dough-brakers increased flour antigen exposure, while packing and decorating resulted in lower exposures. Croissant, puff-pastry, and bread/bun production lines were associated with increased exposure, while cake production and substitution of dusting with the use of divider oil were associated with decreased exposure. Exposure levels can be reduced by the automation of forming tasks, alteration of tasks requiring pouring of flour, and changes to the types of products manufactured.

  13. Determination of carcinoembryonic antigen: experiences with a new radioimmunoassay

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    Lamerz, R [Univ., Munich; Ruider, H

    1976-04-01

    The determination of the carcinoembryonic antigen (CEA) as RIA-test was examined and tested. Labelling was carried out with /sup 125/I according to the chloramin T-method, as RIA in the form of a competitive double antibody examination. The method was tested on patients with colonic and pancreatic carcinomas.

  14. Nephritogenic antigen determinants in epidermal and renal basement membranes of kindreds with Alport-type familial nephritis.

    Science.gov (United States)

    Kashtan, C; Fish, A J; Kleppel, M; Yoshioka, K; Michael, A F

    1986-10-01

    We probed epidermal basement membranes (EBM) of acid-urea denatured skin from members of kindreds with Alport-type familial nephritis (FN) for the presence of antigens reactive with Goodpasture sera (GPS) and serum (FNS) from an Alport patient who developed anti-glomerular basement membrane (GBM) nephritis in a renal allograft. By immunoblotting, GPS reacted primarily with the 28,000 molecular weight (mol wt) monomer but also the 24,000 mol wt and 26,000 mol wt monomers of the noncollagenous globular domain (NC1) of type IV collagen from normal human GBM, while FNS identified only the 26,000-mol wt monomer. FNS reacted with EBM of 12 controls and nine unaffected male kindred members but not EBM of eight affected males. Five affected females exhibited interrupted reactivity of FNS with EBM. GPS showed variable reactivity with EBM and was not discriminating with respect to Alport-type FN. FNS did not stain renal basement members of five affected males. However, the EBM, tubular basement membrane, and Bowman's capsules of affected males contained antigens reactive with GPS. These immunochemical studies suggest that the FNS antigen is distinct from Goodpasture antigen(s). The expression of FNS antigen located on the NC1 domain of type IV collagen is altered in basement membranes of patients with Alport-type FN, and the distribution of this antigenic anomaly within kindreds suggests X-linked dominant transmission of a defective gene.

  15. Characterisation of monoclonal antibodies for human luteinising hormone, and mapping of antigenic determinants on the hormone

    International Nuclear Information System (INIS)

    Soos, M.; Siddle, K.

    1983-01-01

    Twelve mouse monoclonal antibodies for human luteinising hormone were produced. The affinities varied from 4 X 10 7 to 1 X 10 10 l/mol. The specificity of each antibody was assessed by determining the relative reactivities with luteinising hormone, thyroid stimulating hormone, follicle stimulating hormone and chorionic gonadotrophin. Six antibodies bound to the α-subunit as shown by similar reactivity with all hormones, and the remainder to the β-subunit as shown by specificity for luteinising hormone. This latter group of antibodies cross-reacted only weakly with thyroid stimulating hormone (approximately 10%) and follicle stimulating hormone (approximately 3%). Three of these antibodies also showed low reactivity towards chorionic gonadotrophin (<10%), though the others did not (80-300%). The ability of different antibodies to bind simultaneously to luteinising hormone was examined and it was shown that several distinct antigenic determinants existed on both subunits. The characterisation of monoclonal binding sites is discussed in relation to the use of antibodies in two-site immunoradiometric assays. (Auth.)

  16. A procedure for the radioimmunochemical determination of antigens with different structures

    International Nuclear Information System (INIS)

    Galoci, J.; Machan, V.

    1990-01-01

    To a solution of a specific antibody, its antigen labelled with iodine 125, and of the unknown sample is added animal (e.g., goat) antibody against rabbit immunoglobulin immobilized on modified microcrystalline cellulose; always the same quantities are applied always for the same time. After incubation for 1 hr at 20 degC, the immunochemical complex formed is centrifuged and its radioactivity is measured, and the amount of the antigen in sample is determined by comparison with standards. The procedure is applicable to antigens with any structure. Assets of the separating system include a very good stability in solution, a low value of nonspecific bonding and a low sedimentation velocity of the particles used. The procedure is well suited to the quantitation of antigens present in very low concentrations in blood serum, urine, milk, and extracts from tissues or animal feed. In a modification of the procedure, the animal antibody against rabbit immunoglobulin is immobilized on the surface of modified cellulose particles 5 to 15 μm in size exhibiting a very low sedimentation velocity, activated with 1,1-carbonyldiimidazole. (M.D.)

  17. Studies on antigenic cross-reactivity of Trichuris ovis with host mucosal antigens in goat

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    Gautam Patra

    2015-12-01

    Full Text Available Objective: To ascertain whether immunodominant antigens of Trichuris ovis might share and cross react with host molecule. Methods: Two crude protein preparations from anterior and posterior parts of Trichuris ovis were characterized along with host mucosal antigen by double immunodiffusion, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting technique. Conventional scanning electron microscopy was performed as per standard procedure. Results: Sharp and distinct bands of three antigens have been found in double immunodiffusion using hyperimmune serum raised in rabbit indicating the presence of specific antibody against each antigen. All three antigens have shown major and minor bands with molecular weight ranging from 15 to 110 kDa during sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Conclusions: The antigenic cross-reactivity was thought to result from shared antigens. The existence of paracloacal papillae found in the anterior part of the male was not a unique feature for species differentiation.

  18. Genetic Determinism and the Innate-Acquired Distinction in Medicine

    Science.gov (United States)

    2009-01-01

    This article illustrates in which sense genetic determinism is still part of the contemporary interactionist consensus in medicine. Three dimensions of this consensus are discussed: kinds of causes, a continuum of traits ranging from monogenetic diseases to car accidents, and different kinds of determination due to different norms of reaction. On this basis, this article explicates in which sense the interactionist consensus presupposes the innate–acquired distinction. After a descriptive Part 1, Part 2 reviews why the innate–acquired distinction is under attack in contemporary philosophy of biology. Three arguments are then presented to provide a limited and pragmatic defense of the distinction: an epistemic, a conceptual, and a historical argument. If interpreted in a certain manner, and if the pragmatic goals of prevention and treatment (ideally specifying what medicine and health care is all about) are taken into account, then the innate–acquired distinction can be a useful epistemic tool. It can help, first, to understand that genetic determination does not mean fatalism, and, second, to maintain a system of checks and balances in the continuing nature–nurture debates. PMID:20234831

  19. Impaired Cytokine Responses to Epstein-Barr Virus Antigens in Systemic Lupus Erythematosus Patients

    DEFF Research Database (Denmark)

    Draborg, Anette Holck; Sandhu, Noreen; Larsen, Nanna

    2016-01-01

    We analyzed cytokine responses against latent and lytic Epstein-Barr virus (EBV) antigens in systemic lupus erythematosus (SLE) patients and healthy controls (HCs) to obtain an overview of the distinctive immune regulatory response in SLE patients and to expand the previously determined impaired...

  20. Distinction of Fly Artifacts from Human Blood using Immunodetection.

    Science.gov (United States)

    Rivers, David B; Acca, Gillian; Fink, Marc; Brogan, Rebecca; Chen, Dorothy; Schoeffield, Andrew

    2018-02-21

    Insect stains produced by necrophagous flies are indistinguishable morphologically from human bloodstains. At present, no diagnostic tests exist to overcome this deficiency. As the first step toward developing a chemical test to recognize fly artifacts, polyclonal antisera were generated in rats against three distinct antigenic sequences of fly cathepsin D-like proteinase, an enzyme that is structurally distinct in cyclorrhaphous Diptera from other animals. The resulting rat antisera bound to artifacts produced by Protophormia terraenovae and synthetic peptides used to generate the polyclonal antisera, but not with any type of mammalian blood tested in immunoassays. Among the three antisera, anti-md3 serum displayed the highest reactivity for fly stains, demonstrated cross-reactivity for all synthetic peptides representing antigenic sequences of the mature fly enzyme, and bound artifacts originating from the fly digestive tract. Further work is needed to determine whether the antisera are suitable for non-laboratory conditions. © 2018 American Academy of Forensic Sciences.

  1. Mycobacterium leprae antigens involved in human immune responses. I. Identification of four antigens by monoclonal antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Britton, W.J.; Hellqvist, L.; Basten, A.; Raison, R.L.

    1985-12-01

    Four distinct antigens were identified in soluble sonicates of Mycobacterium leprae by using a panel of 11 monoclonal antibodies. Cross-reactivity studies with other mycobacterial species were conducted by using ELISA and immunoblot assays, and demonstrated that determinants on two of the antigens were present in many mycobacteria, whereas the other two were limited in distribution. Competitive inhibition experiments with radiolabeled monoclonal antibodies showed cross-inhibition between antibodies identifying two of the four antigenicbands. These two bands, of M/sub tau/ 4.5 to 6 KD and 30 to 40 KD, were resistant to protease treatment after immunoblotting. In contrast the two other bands of 16 and 70 KD were protease-sensitive. Although all four bands reacted with some human lepromatous leprosy sera in immunoblots, the 4.5 to 6 KD and 30 to 40 KD bands were most prominent. Lepromatous leprosy sera also inhibited the binding of radiolabeled monoclonal antibodies to each of the four antigens, with the mean titer causing 50% inhibition being higher for antibodies reacting with the 4.5 to 6 KD and 30 to 40 KD bands. These findings indicated that all four antigens were involved in the human B cell response to M. leprae.

  2. Mycobacterium leprae antigens involved in human immune responses. I. Identification of four antigens by monoclonal antibodies

    International Nuclear Information System (INIS)

    Britton, W.J.; Hellqvist, L.; Basten, A.; Raison, R.L.

    1985-01-01

    Four distinct antigens were identified in soluble sonicates of Mycobacterium leprae by using a panel of 11 monoclonal antibodies. Cross-reactivity studies with other mycobacterial species were conducted by using ELISA and immunoblot assays, and demonstrated that determinants on two of the antigens were present in many mycobacteria, whereas the other two were limited in distribution. Competitive inhibition experiments with radiolabeled monoclonal antibodies showed cross-inhibition between antibodies identifying two of the four antigenicbands. These two bands, of M/sub tau/ 4.5 to 6 KD and 30 to 40 KD, were resistant to protease treatment after immunoblotting. In contrast the two other bands of 16 and 70 KD were protease-sensitive. Although all four bands reacted with some human lepromatous leprosy sera in immunoblots, the 4.5 to 6 KD and 30 to 40 KD bands were most prominent. Lepromatous leprosy sera also inhibited the binding of radiolabeled monoclonal antibodies to each of the four antigens, with the mean titer causing 50% inhibition being higher for antibodies reacting with the 4.5 to 6 KD and 30 to 40 KD bands. These findings indicated that all four antigens were involved in the human B cell response to M. leprae

  3. Geographical and temporal conservation of antibody recognition of Plasmodium falciparum variant surface antigens

    DEFF Research Database (Denmark)

    Nielsen, Morten A; Vestergaard, Lasse S; Lusingu, John

    2004-01-01

    The slow acquisition of protection against Plasmodium falciparum malaria probably reflects the extensive diversity of important antigens. The variant surface antigens (VSA) that mediate parasite adhesion to a range of host molecules are regarded as important targets of acquired protective immunity......, but their diversity makes them questionable vaccine candidates. We determined levels of VSA-specific immunoglobulin G (IgG) in human plasma collected at four geographically distant and epidemiologically distinct localities with specificity for VSA expressed by P. falciparum isolates from three African countries...

  4. Topographic antigenic determinants recognized by monoclonal antibodies on human choriogonadotropin beta-subunit

    International Nuclear Information System (INIS)

    Bidart, J.M.; Troalen, F.; Salesse, R.; Bousfield, G.R.; Bohuon, C.J.; Bellet, D.H.

    1987-01-01

    We describe a first attempt to study the antibody-combining sites recognized by monoclonal antibodies raised against the beta-subunit of human choriogonadotropin (hCG). Two groups of antibodies were first defined by their ability to recognize only the free beta-subunit or the free and combined subunit. Antibodies FBT-11 and FBT-11-L bind only to hCG beta-subunit but not to hCG, whereas antibodies FBT-10 and D1E8 bind to both the beta-subunit and the hormone. In both cases, the antigenic determinants were localized to the core of the protein (residues 1-112), indicating the weak immunogenicity of the specific carboxyl-terminal extension of hCG-beta. Nine synthetic peptides spanning different regions of hCG-beta and lutropin-beta were assessed for their capacity to inhibit antibody binding. A synthetic peptide inclusive of the NH2-terminal region (residues 1-7) of the hCG beta-subunit was found to inhibit binding to the radiolabeled subunit of a monoclonal antibody specific for free hCG-beta (FBT-11). Further delineation of the antigenic site recognized by this antibody provided evidence for the involvement of fragment 82-92. Moreover, monoclonal antibody FBT-11 inhibited the recombination of hCG-beta to hCG-alpha, indicating that its antigenic determinant might be located nearby or in the hCG-beta portion interacting with the alpha-subunit. Binding of monoclonal antibody FBT-10, corresponding to the second antigenic determinant, was weakly inhibited by fragment 82-105 and did not impair the recombination of the hCG beta-subunit to the hCG alpha-subunit. Its combining site appeared to be located in a region of the intact native choriogonadotropin present at the surface of the hormone-receptor complex

  5. Distinction of the memory B cell response to cognate antigen versus bystander inflammatory signals.

    Science.gov (United States)

    Benson, Micah J; Elgueta, Raul; Schpero, William; Molloy, Michael; Zhang, Weijun; Usherwood, Edward; Noelle, Randolph J

    2009-08-31

    The hypothesis that bystander inflammatory signals promote memory B cell (B(MEM)) self-renewal and differentiation in an antigen-independent manner is critically evaluated herein. To comprehensively address this hypothesis, a detailed analysis is presented examining the response profiles of B-2 lineage B220(+)IgG(+) B(MEM) toward cognate protein antigen in comparison to bystander inflammatory signals. After in vivo antigen encounter, quiescent B(MEM) clonally expand. Surprisingly, proliferating B(MEM) do not acquire germinal center (GC) B cell markers before generating daughter B(MEM) and differentiating into plasma cells or form structurally identifiable GCs. In striking contrast to cognate antigen, inflammatory stimuli, including Toll-like receptor agonists or bystander T cell activation, fail to induce even low levels of B(MEM) proliferation or differentiation in vivo. Under the extreme conditions of adjuvanted protein vaccination or acute viral infection, no detectable bystander proliferation or differentiation of B(MEM) occurred. The absence of a B(MEM) response to nonspecific inflammatory signals clearly shows that B(MEM) proliferation and differentiation is a process tightly controlled by the availability of cognate antigen.

  6. On the localisation of antigenic determinants in a Bence Jones protein

    NARCIS (Netherlands)

    Eyk, H.G. van; Myszkowska, K.

    1967-01-01

    1. 1. The presence of a low molecular weight protein (1.2 S), having antigenic determinants in common with the homologous Bence Jones protein (3.4 S), has been observed in the urine of a patient with multiple myeloma. Its amino acid composition and α-NH2-terminal amino acid residue make it likely

  7. Evidence Suggesting That Francisella tularensis O-Antigen Capsule Contains a Lipid A-Like Molecule That Is Structurally Distinct from the More Abundant Free Lipid A.

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    Jason H Barker

    Full Text Available Francisella tularensis, the Gram-negative bacterium that causes tularemia, produces a high molecular weight capsule that is immunologically distinct from Francisella lipopolysaccharide but contains the same O-antigen tetrasaccharide. To pursue the possibility that the capsule of Francisella live vaccine strain (LVS has a structurally unique lipid anchor, we have metabolically labeled Francisella with [14C]acetate to facilitate highly sensitive compositional analysis of capsule-associated lipids. Capsule was purified by two independent methods and yielded similar results. Autoradiographic and immunologic analysis confirmed that this purified material was largely devoid of low molecular weight LPS and of the copious amounts of free lipid A that the Francisellae accumulate. Chemical hydrolysis yielded [14C]-labeled free fatty acids characteristic of Francisella lipid A but with a different molar ratio of 3-OH C18:0 to 3-OH C16:0 and different composition of non-hydroxylated fatty acids (mainly C14:0 rather than C16:0 than that of free Francisella lipid A. Mild acid hydrolysis to induce selective cleavage of KDO-lipid A linkage yielded a [14C]-labeled product that partitioned during Bligh/Dyer extraction and migrated during thin-layer chromatography like lipid A. These findings suggest that the O-antigen capsule of Francisella contains a covalently linked and structurally distinct lipid A species. The presence of a discrete lipid A-like molecule associated with capsule raises the possibility that Francisella selectively exploits lipid A structural heterogeneity to regulate synthesis, transport, and stable bacterial surface association of the O-antigen capsular layer.

  8. Regulation of T cell response to leishmania antigens by determinants of histocompatibility leukocyte class I and II molecules

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    Bacellar O.

    1998-01-01

    Full Text Available It has been shown that HLA class I molecules play a significant role in the regulation of the proliferation of T cells activated by mitogens and antigens. We evaluated the ability of mAb to a framework determinant of HLA class I molecules to regulate T cell proliferation and interferon gamma (IFN-g production against leishmania, PPD, C. albicans and tetanus toxoid antigens in patients with tegumentary leishmaniasis and healthy subjects. The anti-major histocompatibility complex (MHC mAb (W6/32 suppressed lymphocyte proliferation by 90% in cultures stimulated with aCD3, but the suppression was variable in cultures stimulated with leishmania antigen. This suppression ranged from 30-67% and was observed only in 5 of 11 patients. IFN-g production against leishmania antigen was also suppressed by anti-HLA class I mAb. In 3 patients IFN-g levels were suppressed by more than 60%, while in the other 2 cultures IFN-g levels were 36 and 10% lower than controls. The suppression by HLA class I mAb to the proliferative response in leishmaniasis patients and in healthy controls varied with the antigens and the patients or donors tested. To determine whether the suppression is directed at antigen presenting cells (APCs or at the responding T cells, experiments with antigen-primed non-adherent cells, separately incubated with W6/32, were performed. Suppression of proliferation was only observed when the W6/32 mAb was added in the presence of T cells. These data provide evidence that a mAb directed at HLA class I framework determinants can suppress proliferation and cytokine secretion in response to several antigens.

  9. Structure-guided evolution of antigenically distinct adeno-associated virus variants for immune evasion.

    Science.gov (United States)

    Tse, Longping Victor; Klinc, Kelli A; Madigan, Victoria J; Castellanos Rivera, Ruth M; Wells, Lindsey F; Havlik, L Patrick; Smith, J Kennon; Agbandje-McKenna, Mavis; Asokan, Aravind

    2017-06-13

    Preexisting neutralizing antibodies (NAbs) against adeno-associated viruses (AAVs) pose a major, unresolved challenge that restricts patient enrollment in gene therapy clinical trials using recombinant AAV vectors. Structural studies suggest that despite a high degree of sequence variability, antibody recognition sites or antigenic hotspots on AAVs and other related parvoviruses might be evolutionarily conserved. To test this hypothesis, we developed a structure-guided evolution approach that does not require selective pressure exerted by NAbs. This strategy yielded highly divergent antigenic footprints that do not exist in natural AAV isolates. Specifically, synthetic variants obtained by evolving murine antigenic epitopes on an AAV serotype 1 capsid template can evade NAbs without compromising titer, transduction efficiency, or tissue tropism. One lead AAV variant generated by combining multiple evolved antigenic sites effectively evades polyclonal anti-AAV1 neutralizing sera from immunized mice and rhesus macaques. Furthermore, this variant displays robust immune evasion in nonhuman primate and human serum samples at dilution factors as high as 1:5, currently mandated by several clinical trials. Our results provide evidence that antibody recognition of AAV capsids is conserved across species. This approach can be applied to any AAV strain to evade NAbs in prospective patients for human gene therapy.

  10. Carcinoembryonic antigen (CEA)

    International Nuclear Information System (INIS)

    Ephraim, K.H.; Cox, P.H.; Hamer, C.J.A. v.d.; Berends, W.; Delhez, H.

    1977-01-01

    The carcinoembryonic antigen (CEA) is a complex of antigen determinants and also the carrier of these determinants. Chemically it is a glycoprotein. Its occurrence in blood serum or urine is correlated with malignant disease. Several radioimmunoassays (RIA) have been developed, one by Hoffmann-Laroche and one by the Rotterdam Radiotherapeutic Institute. Both methods and the Hoffmann assay kit are tested. Specifications are given for isolation of the antigen, preparation of the antiserum, and the execution of the RIA. Biochemical and clinical aspects are discussed

  11. Detection of Burkholderia pseudomallei O-antigen serotypes in near-neighbor species

    Directory of Open Access Journals (Sweden)

    Stone Joshua K

    2012-11-01

    Full Text Available Abstract Background Burkholderia pseudomallei is the etiological agent of melioidosis and a CDC category B select agent with no available effective vaccine. Previous immunizations in mice have utilized the lipopolysaccharide (LPS as a potential vaccine target because it is known as one of the most important antigenic epitopes in B. pseudomallei. Complicating this strategy are the four different B. pseudomallei LPS O-antigen types: A, B, B2, and rough. Sero-crossreactivity is common among O-antigens of Burkholderia species. Here, we identified the presence of multiple B. pseudomallei O-antigen types and sero-crossreactivity in its near-neighbor species. Results PCR screening of O-antigen biosynthesis genes, phenotypic characterization using SDS-PAGE, and immunoblot analysis showed that majority of B. mallei and B. thailandensis strains contained the typical O-antigen type A. In contrast, most of B. ubonensis and B. thailandensis-like strains expressed the atypical O-antigen types B and B2, respectively. Most B. oklahomensis strains expressed a distinct and non-seroreactive O-antigen type, except strain E0147 which expressed O-antigen type A. O-antigen type B2 was also detected in B. thailandensis 82172, B. ubonensis MSMB108, and Burkholderia sp. MSMB175. Interestingly, B. thailandensis-like MSMB43 contained a novel serotype B positive O-antigen. Conclusions This study expands the number of species which express B. pseudomallei O-antigen types. Further work is required to elucidate the full structures and how closely these are to the B. pseudomallei O-antigens, which will ultimately determine the efficacy of the near-neighbor B serotypes for vaccine development.

  12. Genetic and antigenic analysis of the G attachment protein of bovine respiratory syncytial virus strains

    DEFF Research Database (Denmark)

    Elvander, M.; Vilcek, S.; Baule, C.

    1998-01-01

    Antigenic and genetic studies of bovine respiratory syncytial virus (BRSV) were made on isolates obtained from three continents over 27 years. Antigenic variation between eight isolates was initially determined using protein G-specific monoclonal antibodies. Four distinct reaction patterns were...... of a 731 nucleotide fragment in the G protein gene. Nine of the BRSV strains were analysed by direct sequencing of RT-PCR amplicons whereas sequences of 18 BRSV and three human respiratory syncytial virus (HRSV) strains were obtained from GenBank. The analysis revealed similarities of 88-100% among BRSV...

  13. Limited antigenic variation in the Trypanosoma cruzi candidate vaccine antigen TSA-1.

    Science.gov (United States)

    Knight, J M; Zingales, B; Bottazzi, M E; Hotez, P; Zhan, B

    2014-12-01

    Chagas disease (American trypanosomiasis caused by Trypanosoma cruzi) is one of the most important neglected tropical diseases in the Western Hemisphere. The toxicities and limited efficacies of current antitrypanosomal drugs have prompted a search for alternative technologies such as a therapeutic vaccine comprised of T. cruzi antigens, including a recombinant antigen encoding the N-terminal 65 kDa portion of Trypomastigote surface antigen-1 (TSA-1). With at least six known genetically distinct T. cruzi lineages, variability between the different lineages poses a unique challenge for the development of broadly effective therapeutic vaccine. The variability across the major lineages in the current vaccine candidate antigen TSA-1 has not previously been addressed. To assess the variation in TSA-1, we cloned and sequenced TSA-1 from several different T. cruzi strains representing three of the most clinically relevant lineages. Analysis of the different alleles showed limited variation in TSA-1 across the different strains and fit with the current theory for the evolution of the different lineages. Additionally, minimal variation in known antigenic epitopes for the HLA-A 02 allele suggests that interlineage variation in TSA-1 would not impair the range and efficacy of a vaccine containing TSA-1. © 2014 John Wiley & Sons Ltd.

  14. Determining the binding affinity of therapeutic monoclonal antibodies towards their native unpurified antigens in human serum.

    Directory of Open Access Journals (Sweden)

    Christine Bee

    Full Text Available Monoclonal antibodies (mAbs are a growing segment of therapeutics, yet their in vitro characterization remains challenging. While it is essential that a therapeutic mAb recognizes the native, physiologically occurring epitope, the generation and selection of mAbs often rely on the use of purified recombinant versions of the antigen that may display non-native epitopes. Here, we present a method to measure both, the binding affinity of a therapeutic mAb towards its native unpurified antigen in human serum, and the antigen's endogenous concentration, by combining the kinetic exclusion assay and Biacore's calibration free concentration analysis. To illustrate the broad utility of our method, we studied a panel of mAbs raised against three disparate soluble antigens that are abundant in the serum of healthy donors: proprotein convertase subtilisin/kexin type 9 (PCSK9, progranulin (PGRN, and fatty acid binding protein (FABP4. We also determined the affinity of each mAb towards its purified recombinant antigen and assessed whether the interactions were pH-dependent. Of the six mAbs studied, three did not appear to discriminate between the serum and recombinant forms of the antigen; one mAb bound serum antigen with a higher affinity than recombinant antigen; and two mAbs displayed a different affinity for serum antigen that could be explained by a pH-dependent interaction. Our results highlight the importance of taking pH into account when measuring the affinities of mAbs towards their serum antigens, since the pH of serum samples becomes increasingly alkaline upon aerobic handling.

  15. Use of radio-immuno-inhibition assay for the study of the y, d and w determinants of hepatitis B surface antigen

    Energy Technology Data Exchange (ETDEWEB)

    Donea-Debroise, B; Brocteur, J; Andre, A; Remacle, M B [Liege Univ. (Belgium)

    1979-01-01

    A radioimmunoassay determination of the HBs antigen subtypes is discussed, this simple but effective technique was used in association with the use of the Austria II kit (Abbott Laboratories). This method consists of an inhibition reaction of the Austria II test, by previous incubation of the antigen to be subtyped with a monospecific antibody. With this method we were able to distinguish the y and the d antigens as well as the w1, w3, w4 determinants of hepatitis B surface antigen. We have included a frequency table of the various HBs subtypes found among donor and patient populations in Liege.

  16. Mature IgM-expressing plasma cells sense antigen and develop competence for cytokine production upon antigenic challenge

    Science.gov (United States)

    Blanc, Pascal; Moro-Sibilot, Ludovic; Barthly, Lucas; Jagot, Ferdinand; This, Sébastien; de Bernard, Simon; Buffat, Laurent; Dussurgey, Sébastien; Colisson, Renaud; Hobeika, Elias; Fest, Thierry; Taillardet, Morgan; Thaunat, Olivier; Sicard, Antoine; Mondière, Paul; Genestier, Laurent; Nutt, Stephen L.; Defrance, Thierry

    2016-01-01

    Dogma holds that plasma cells, as opposed to B cells, cannot bind antigen because they have switched from expression of membrane-bound immunoglobulins (Ig) that constitute the B-cell receptor (BCR) to production of the secreted form of immunoglobulins. Here we compare the phenotypical and functional attributes of plasma cells generated by the T-cell-dependent and T-cell-independent forms of the hapten NP. We show that the nature of the secreted Ig isotype, rather than the chemical structure of the immunizing antigen, defines two functionally distinct populations of plasma cells. Fully mature IgM-expressing plasma cells resident in the bone marrow retain expression of a functional BCR, whereas their IgG+ counterparts do not. Antigen boost modifies the gene expression profile of IgM+ plasma cells and initiates a cytokine production program, characterized by upregulation of CCL5 and IL-10. Our results demonstrate that IgM-expressing plasma cells can sense antigen and acquire competence for cytokine production upon antigenic challenge. PMID:27924814

  17. Antigenic determinants of prostate-specific antigen (PSA) and development of assays specific for different forms of PSA.

    OpenAIRE

    Nilsson, O.; Peter, A.; Andersson, I.; Nilsson, K.; Grundstr?m, B.; Karlsson, B.

    1997-01-01

    Monoclonal antibodies were raised against prostate-specific antigen (PSA) by immunization with purified free PSA, i.e. not in complex with any protease inhibitor (F-PSA) and PSA in complex with alpha1-anti-chymotrypsin (PSA-ACT). Epitope mapping of PSA using the established monoclonal antibody revealed a complex pattern of independent and partly overlapping antigenic domains in the PSA molecule. Four independent antigenic domains and at least three partly overlapping domains were exposed both...

  18. Prognostic value of determination of carcinoembryonic antigen and α-fetoprotein level in blood plasma in patients with cancer stomach

    International Nuclear Information System (INIS)

    Smyslova, V.N.; Vygonnyj, I.I.

    1986-01-01

    60 donors and 129 patients with cancer stomach were examined. Tumor antigens were determined in blood plasma by the method of radioimmunoassay. The upper boundary of the norm of alpha-fetoprotein (AFP) and carcino-embryonic antigen (CEA) is 12 ng/ml. Increased concentration of antigens studied is detected in most patients. It is established that the level of antigens increases depending on generalization of the process, cancer stage, tumor propagation in the stomach wall, patient's age. High volumes of AFP and CEA after operation give evidence about non-radicality of operation and bad prognosis

  19. Tumor Associated Antigenic Peptides in Prostate Cancer

    National Research Council Canada - National Science Library

    Tiwari, Raj

    1999-01-01

    .... We proposed to identify these novel antigens in an experimental rat model using purified preparations of the heat shock protein gp96 and a library of synthetic distinct antibodies that were available...

  20. Novel 6xHis tagged foot-and-mouth disease virus vaccine bound to nanolipoprotein adjuvant via metal ions provides antigenic distinction and effective protective immunity

    Energy Technology Data Exchange (ETDEWEB)

    Rai, Devendra K.; Segundo, Fayna Diaz-San [Foreign Animal Disease Research Unit, United States Department of Agriculture, Agricultural Research Service, Plum Island Animal Disease Center, Greenport, NY 11944 (United States); Department of Pathobiology and Veterinary Science, CANR, University of Connecticut, Storrs, CT 06269 (United States); Schafer, Elizabeth [Foreign Animal Disease Research Unit, United States Department of Agriculture, Agricultural Research Service, Plum Island Animal Disease Center, Greenport, NY 11944 (United States); Burrage, Thomas G. [Department of Homeland Security, S & T, Targeted Advance Development, Virus, Cellular and Molecular Imaging Agriculture, Agricultural Research Service, Plum Island Animal Disease Center, Greenport, NY 11944 (United States); Rodriguez, Luis L.; Santos, Teresa de los [Foreign Animal Disease Research Unit, United States Department of Agriculture, Agricultural Research Service, Plum Island Animal Disease Center, Greenport, NY 11944 (United States); Hoeprich, Paul D. [Physical and Life Sciences Directorate, Lawrence Livermore National Laboratory, Livermore, CA (United States); Rieder, Elizabeth, E-mail: Elizabeth.Rieder@ars.usda.gov [Foreign Animal Disease Research Unit, United States Department of Agriculture, Agricultural Research Service, Plum Island Animal Disease Center, Greenport, NY 11944 (United States)

    2016-08-15

    Here, we engineered two FMD viruses with histidine residues inserted into or fused to the FMDV capsid. Both 6xHis viruses exhibited growth kinetics, plaque morphologies and antigenic characteristics similar to wild-type virus. The 6xHis tag allowed one-step purification of the mutant virions by Co{sup 2+} affinity columns. Electron microscopy and biochemical assays showed that the 6xHis FMDVs readily assembled into antigen: adjuvant complexes in solution, by conjugating with Ni{sup 2+}-chelated nanolipoprotein and monophosphoryl lipid A adjuvant (MPLA:NiNLP). Animals Immunized with the inactivated 6xHis-FMDV:MPLA:NiNLP vaccine acquired enhanced protective immunity against FMDV challenge compared to virions alone. Induction of anti-6xHis and anti-FMDV neutralizing antibodies in the immunized animals could be exploited in the differentiation of vaccinated from infected animals needed for the improvement of FMD control measures. The novel marker vaccine/nanolipid technology described here has broad applications for the development of distinctive and effective immune responses to other pathogens of importance. - Highlights: • 6xHis-tags in A{sub 24} FMDV enable purification and biding to adjuvants via metal ions. • 6xHis A{sub 24} FMDV:MPLA:NiNLP vaccine enhanced protective immunity against FMDV. • Surface exposed capsid tags allow distinction of infected from vaccinated animals.

  1. Novel 6xHis tagged foot-and-mouth disease virus vaccine bound to nanolipoprotein adjuvant via metal ions provides antigenic distinction and effective protective immunity

    International Nuclear Information System (INIS)

    Rai, Devendra K.; Segundo, Fayna Diaz-San; Schafer, Elizabeth; Burrage, Thomas G.; Rodriguez, Luis L.; Santos, Teresa de los; Hoeprich, Paul D.; Rieder, Elizabeth

    2016-01-01

    Here, we engineered two FMD viruses with histidine residues inserted into or fused to the FMDV capsid. Both 6xHis viruses exhibited growth kinetics, plaque morphologies and antigenic characteristics similar to wild-type virus. The 6xHis tag allowed one-step purification of the mutant virions by Co"2"+ affinity columns. Electron microscopy and biochemical assays showed that the 6xHis FMDVs readily assembled into antigen: adjuvant complexes in solution, by conjugating with Ni"2"+-chelated nanolipoprotein and monophosphoryl lipid A adjuvant (MPLA:NiNLP). Animals Immunized with the inactivated 6xHis-FMDV:MPLA:NiNLP vaccine acquired enhanced protective immunity against FMDV challenge compared to virions alone. Induction of anti-6xHis and anti-FMDV neutralizing antibodies in the immunized animals could be exploited in the differentiation of vaccinated from infected animals needed for the improvement of FMD control measures. The novel marker vaccine/nanolipid technology described here has broad applications for the development of distinctive and effective immune responses to other pathogens of importance. - Highlights: • 6xHis-tags in A_2_4 FMDV enable purification and biding to adjuvants via metal ions. • 6xHis A_2_4 FMDV:MPLA:NiNLP vaccine enhanced protective immunity against FMDV. • Surface exposed capsid tags allow distinction of infected from vaccinated animals.

  2. The use of radio-immuno-inhibition assay for the study of the y, d and w determinants of hepatitis B surface antigen

    International Nuclear Information System (INIS)

    Donea-Debroise, B.; Brocteur, J.; Andre, A.; Remacle, M.B.

    1979-01-01

    A radioimmunoassay determination of the HBs antigen subtypes is discussed, this simple but effective technique was used in association with the use of the Austria II kit (Abbott Laboratories). This method consists of an inhibition reaction of the Austria II test, by previous incubation of the antigen to be subtyped with a monospecific antibody. With this method we were able to distinguish the y and the d antigens as well as the w1, w3, w4 determinants of hepatitis B surface antigen. We have included a frequency table of the various HBs subtypes found among donor and patient populations in Liege

  3. Characterization of antigen association with accessory cells: specific removal of processed antigens from the cell surface by phospholipases

    International Nuclear Information System (INIS)

    Falo, L.D. Jr.; Haber, S.I.; Herrmann, S.; Benacerraf, B.; Rock, K.L.

    1987-01-01

    To characterize the basis for the cell surface association of processed antigen with the antigen-presenting cell (APC) the authors analyzed its sensitivity to enzymatic digestion. Antigen-exposed APC that are treated with phospholipase and then immediately fixed lose their ability to stimulate antigen-plus-Ia-specific T-T hybridomas. This effect is seen with highly purified phospholipase A 2 and phospholipase C. In addition it is observed with three distinct antigens - ovalbumin, bovine insulin, and poly(LGlu 56 LLys 35 LPhe 9 )[(GluLysPhe)/sub n/]. The effect of phospholipases is highly specific. Identically treated APC are equivalent to control in their ability to stimulate alloreactive hybridomas specific for precisely the same Ia molecule that is corecognized by antigen-plus-Ia-specific hybrids. Furthermore, the antigen-presenting function of enzyme-treated, fixed APC can be reconstituted by the addition of exogenous in vitro processed or processing independent antigens. In parallel studies 125 I-labeled avidin was shown to specifically bind to APC that were previously exposed and allowed to process biotin-insulin. Biotin-insulin-exposed APC that are pretreated with phospholipase bind significantly less 125 I-labeled avidin than do untreated, exposed APC. Identical enzyme treatment does not reduce the binding of avidin to a biotinylated antibody already bound to class II major histocompatibility complex molecules of APC. These studies demonstrate that phospholipase effectively removes processed cell surface antigen

  4. Three-site sandwich radioimmunoassay with monoclonal antibodies for a sensitive determination of human alpha-fetoprotein

    International Nuclear Information System (INIS)

    Nomura, M.; Imai, M.; Takahashi, K.; Kumakura, T.; Tachibana, K.; Aoyagi, S.; Usuda, S.; Nakamura, T.; Miyakawa, Y.; Mayumi, M.

    1983-01-01

    Utilizing monoclonal antibodies against human alpha-fetoprotein, 3 distinct antigenic determinants were identified. These antigenic determinants, provisionally designated a, b and c, were arranged in such a manner that the binding of one determinant with the corresponding antibody did not inhibit, or only barely inhibited the binding of antibodies directed to the other 2 determinants. Monoclonal antibodies with 3 different specificities were, therefore, applied to develop a sandwich-type solid-phase radioimmunoassay of the antigen in which wells were coated with anti-a, and radiolabeled anti-b together with radiolabeled anti-c was employed to detect the bound antigen. The 3-site sandwich radioimmunoassay involving 3 different determinants gave a higher sensitivity than 2-site assays in which only anti-b or anti-c was employed as a radiolabeled reagent, because the radioactivity of the 2 labeled antibodies was added on the antigen bound to immobilized anti-a. (Auth.)

  5. FIBER OPTIC IMMUNOSENSORS BASED ON ENHANCED CHEMILUMINESCENCE AND THEIR APPLICATION TO DETERMINE DIFFERENT ANTIGENS

    NARCIS (Netherlands)

    STARODUB, NF; ARENKOV, PJ; STARODUB, AN; BEREZIN, VA

    The main scheme of the cell for measuring as well as the sensor device itself, the analysis conditions and the results of quantitative determination of several antigens such as estradiol-17, alpha-2-interferon, chorionic honadotropine, antibodies to the influenza virus, total IgG and cells of

  6. In vivo xenogeneic scaffold fate is determined by residual antigenicity and extracellular matrix preservation.

    Science.gov (United States)

    Wong, Maelene L; Wong, Janelle L; Vapniarsky, Natalia; Griffiths, Leigh G

    2016-06-01

    The immunological potential of animal-derived tissues and organs is the critical hurdle to increasing their clinical implementation. Glutaraldehyde-fixation cross-links proteins in xenogeneic tissues (e.g., bovine pericardium) to delay immune rejection, but also compromises the regenerative potential of the resultant biomaterial. Unfixed xenogeneic biomaterials in which xenoantigenicity has been ameliorated and native extracellular matrix (ECM) architecture has been maintained have the potential to overcome limitations of current clinically utilized glutaraldehyde-fixed biomaterials. The objective of this work was to determine how residual antigenicity and ECM architecture preservation modulate recipient immune and regenerative responses towards unfixed bovine pericardium (BP) ECM scaffolds. Disruption of ECM architecture during scaffold generation, with either SDS-decellularization or glutaraldehyde-fixation, stimulated recipient foreign body response and resultant fibrotic encapsulation following leporine subpannicular implantation. Conversely, BP scaffolds subjected to stepwise removal of hydrophilic and lipophilic antigens using amidosulfobetaine-14 (ASB-14) maintained native ECM architecture and thereby avoided fibrotic encapsulation. Removal of hydrophilic and lipophilic antigens significantly decreased local and systemic graft-specific, adaptive immune responses and subsequent calcification of BP scaffolds compared to scaffolds undergoing hydrophile removal only. Critically, removal of antigenic components and preservation of ECM architecture with ASB-14 promoted full-thickness recipient non-immune cellular repopulation of the BP scaffold. Further, unlike clinically utilized fixed BP, ASB-14-treated scaffolds fostered rapid intimal and medial vessel wall regeneration in a porcine carotid patch angioplasty model. This work highlights the importance of residual antigenicity and ECM architecture preservation in modulating recipient immune and regenerative

  7. Distinct galactofuranose antigens in the cell wall and culture supernatants as a means to differentiate Fusarium from AspergillusspeciesAnnegret

    DEFF Research Database (Denmark)

    Wiedemann, Annegret; Kakoschke, Tamara Katharina; Speth, Cornelia

    2016-01-01

    tDetection of carbohydrate antigens is an important means for diagnosis of invasive fungal infections. Fordiagnosis of systemic Aspergillus infections, galactomannan is commonly used, the core antigenic struc-ture of which consists of chains of several galactofuranose moieties. In this study, we ....... fumigatus and Fusar-ium hyphae in immunohistology. Moreover, since Fusarium releases the AB135-8 antigen, it appears tobe a promising target antigen for a serological detection of Fusarium infections....

  8. Co-stimulatory signaling determines tumor antigen sensitivity and persistence of CAR T cells targeting PSCA+ metastatic prostate cancer.

    Science.gov (United States)

    Priceman, Saul J; Gerdts, Ethan A; Tilakawardane, Dileshni; Kennewick, Kelly T; Murad, John P; Park, Anthony K; Jeang, Brook; Yamaguchi, Yukiko; Yang, Xin; Urak, Ryan; Weng, Lihong; Chang, Wen-Chung; Wright, Sarah; Pal, Sumanta; Reiter, Robert E; Wu, Anna M; Brown, Christine E; Forman, Stephen J

    2018-01-01

    Advancing chimeric antigen receptor (CAR)-engineered adoptive T cells for the treatment of solid cancers is a major focus in the field of immunotherapy, given impressive recent clinical responses in hematological malignancies. Prostate cancer may be amenable to T cell-based immunotherapy since several tumor antigens, including prostate stem-cell antigen (PSCA), are widely over-expressed in metastatic disease. While antigen selectivity of CARs for solid cancers is crucial, it is problematic due to the absence of truly restricted tumor antigen expression and potential safety concerns with "on-target off-tumor" activity. Here, we show that the intracellular co-stimulatory signaling domain can determine a CAR's sensitivity for tumor antigen expression. A 4-1BB intracellular co-stimulatory signaling domain in PSCA-CARs confers improved selectivity for higher tumor antigen density, reduced T cell exhaustion phenotype, and equivalent tumor killing ability compared to PSCA-CARs containing the CD28 co-stimulatory signaling domain. PSCA-CARs exhibit robust in vivo anti-tumor activity in patient-derived bone-metastatic prostate cancer xenograft models, and 4-1BB-containing CARs show superior T cell persistence and control of disease compared with CD28-containing CARs. Our study demonstrates the importance of co-stimulation in defining an optimal CAR T cell, and also highlights the significance of clinically relevant models in developing solid cancer CAR T cell therapies.

  9. Co-stimulatory signaling determines tumor antigen sensitivity and persistence of CAR T cells targeting PSCA+ metastatic prostate cancer

    Science.gov (United States)

    Priceman, Saul J.; Gerdts, Ethan A.; Tilakawardane, Dileshni; Kennewick, Kelly T.; Murad, John P.; Park, Anthony K.; Jeang, Brook; Yamaguchi, Yukiko; Urak, Ryan; Weng, Lihong; Chang, Wen-Chung; Wright, Sarah; Pal, Sumanta; Reiter, Robert E.; Brown, Christine E.; Forman, Stephen J.

    2018-01-01

    ABSTRACT Advancing chimeric antigen receptor (CAR)-engineered adoptive T cells for the treatment of solid cancers is a major focus in the field of immunotherapy, given impressive recent clinical responses in hematological malignancies. Prostate cancer may be amenable to T cell-based immunotherapy since several tumor antigens, including prostate stem-cell antigen (PSCA), are widely over-expressed in metastatic disease. While antigen selectivity of CARs for solid cancers is crucial, it is problematic due to the absence of truly restricted tumor antigen expression and potential safety concerns with “on-target off-tumor” activity. Here, we show that the intracellular co-stimulatory signaling domain can determine a CAR's sensitivity for tumor antigen expression. A 4-1BB intracellular co-stimulatory signaling domain in PSCA-CARs confers improved selectivity for higher tumor antigen density, reduced T cell exhaustion phenotype, and equivalent tumor killing ability compared to PSCA-CARs containing the CD28 co-stimulatory signaling domain. PSCA-CARs exhibit robust in vivo anti-tumor activity in patient-derived bone-metastatic prostate cancer xenograft models, and 4-1BB-containing CARs show superior T cell persistence and control of disease compared with CD28-containing CARs. Our study demonstrates the importance of co-stimulation in defining an optimal CAR T cell, and also highlights the significance of clinically relevant models in developing solid cancer CAR T cell therapies. PMID:29308300

  10. Radioimmunochemical characterization of hemoglobins Lepore and Kenya: unique antigenic determinants located on hybrid hemoglobins

    International Nuclear Information System (INIS)

    Garver, F.A.; Altay, G.; Baker, M.M.; Gravely, M.; Huisman, T.H.J.

    1978-01-01

    Antisera were produced in rabbits to the three known types of Lepore hemoglobins, which contain hybrid delta-β non-α-chains, and to hemoglobin Kenya, which has a hybrid γ-β non-α-chain. By using a sensitive radioimmunoassay technique, the absorbed antisera were shown to contain an antibody population that was specific for the hybrid hemoglobin and did not cross-react with normal hemoglobins. However, with the absorbed Lepore-specific antisera, the three known types of Lepore hemoglobins were antigenically indistinguishable from each other, suggesting that antibodies are not produced to the primary structural differences which define the three non-α-chains of the Lepore hemoglobins. These studies demonstrate that the non-α-subunits of hemoglobins Lepore and Kenya possess unique antigenic determinant sites, evidently resulting from an altered polypeptide conformation

  11. Chlorphenesin: an antigen-associated immunosuppressant.

    Science.gov (United States)

    Whang, H Y; Neter, E

    1970-07-01

    Chlorphenesin (3-p-chlorophenoxy-1,2-propanediol), when injected intravenously together with either of two common bacterial antigens, inhibits the antibody response of the rabbit. The antigens studied are those common to Enterobacteriaceae and to gram-positive bacteria. The immunosuppression is contingent upon incubation of chlorphenesin and antigen in vitro prior to administration, since separate injection of antigen and inhibitor or of mixtures without prior incubation yields undiminished antibody response. Chlorphenesin, as shown by hemagglutination-inhibition tests, does not alter the antigenic determinants, because antibody neutralization occurs in the presence or absence of the drug. The immunosuppressive effect is reversible, since precipitation of chlorphenesin at 4 C substantially restores immunogenicity. Animals immunized with antigen-drug mixtures, which fail to respond with significant antibody production, nonetheless are immunologically primed. It is concluded that chlorphenesin represents another example of antigen-associated immunosuppressants.

  12. Expression and Antigenic Evaluation of VacA Antigenic Fragment of Helicobacter Pylori

    Science.gov (United States)

    Hasanzadeh, Leila; Ghaznavi-Rad, Ehsanollah; Soufian, Safieh; Farjadi, Vahideh; Abtahi, Hamid

    2013-01-01

    Objective(s) : Helicobacter pylori, a human specific gastric pathogen is a causative agent of chronic active gastritis. The vacuolating cytotoxin (VacA) is an effective virulence factor involved in gastric injury. The aim of this study was to construct a recombinant protein containing antigenic region of VacA gene and determine its antigenicity. Materials and Methods: The antigenic region of VacA gene was detected by bioinformatics methods. The polymerase chain reaction method was used to amplify a highly antigenic region of VacA gene from chromosomal DNA of H. pylori. The eluted product was cloned into the prokaryotic expression vector pET32a. The target protein was expressed in the Escherichia coli BL21 (DE3) pLysS. The bacteria including pET32a-VacA plasmids were induced by IPTG. The antigenicity was finally studied by western blotting using sera of 15 H. pylori infected patients after purification. Results: Enzyme digestion analysis, PCR and DNA sequencing results showed that the target gene was inserted correctly into the recombinant vector. The expressed protein was purified successfully via affinity chromatography. Data indicated that antigenic region of VacA protein from Helicobacter pylori was recognized by all 15 patient’s sera. Conclusion : Our data showed that antigenic region of VacA protein can be expressed by in E. co.li. This protein was recognized by sera patients suffering from H. pylori infection. the recombinant protein has similar epitopes and close antigenic properties to the natural form of this antigen. Recombinant antigenic region of VacA protein also seems to be a promising antigen for protective and serologic diagnosis . PMID:23997913

  13. Expression and Antigenic Evaluation of VacA Antigenic Fragment of Helicobacter Pylori

    Directory of Open Access Journals (Sweden)

    Leila Hasanzadeh

    2013-07-01

    Full Text Available Objective(s: Helicobacter pylori, a human specific gastric pathogen is a causative agent of chronic active gastritis. The vacuolating cytotoxin (VacA is an effective virulence factor involved in gastric injury. The aim of this study was to construct a recombinant protein containing antigenic region of VacA gene and determine its antigenicity.   Materials and Methods: The antigenic region of VacA gene was detected by bioinformatics methods. The polymerase chain reaction method was used to amplify a highly antigenic region of VacA gene from chromosomal DNA of H. pylori. The eluted product was cloned into the prokaryotic expression vector pET32a. The target protein was expressed in the Escherichia coli BL21 (DE3 pLysS. The bacteria including pET32a-VacA plasmids were induced by IPTG. The antigenicity was finally studied by western blotting using sera of 15 H. pylori infected patients after purification. Results: Enzyme digestion analysis, PCR and DNA sequencing results showed that the target gene was inserted correctly into the recombinant vector. The expressed protein was purified successfully via affinity chromatography. Data indicated that antigenic region of VacA protein from Helicobacter pylori was recognized by all 15 patient’s sera. Conclusion : Our data showed that antigenic region of VacA protein can be expressed by in E. co.li. This protein was recognized by sera patients suffering from H. pylori infection. the recombinant protein has similar epitopes and close antigenic properties to the natural form of this antigen. Recombinant antigenic region of VacA protein also seems to be a promising antigen for protective and serologic diagnosis .

  14. Abnormal expression of blood group-related antigens in uterine endometrial cancers.

    Science.gov (United States)

    Tsukazaki, K; Sakayori, M; Arai, H; Yamaoka, K; Kurihara, S; Nozawa, S

    1991-08-01

    The expression of A, B, and H group antigens, Lewis group antigens (Lewis(a), Lewis(b), Lewis(x), and Lewis(y)), and Lc4 and nLc4 antigens, the precursor antigens of both groups, was examined immunohistochemically with monoclonal antibodies in 9 normal endometria, 6 endometrial hyperplasias, and 31 endometrial cancers. 1) A, B and/or H antigens were detected in endometrial cancers at an incidence of 51.6%, while no distinct localization of these antigens was observed in normal endometria. H antigen, the precursor of A and B antigens, was particularly frequently detected in endometrial cancers. 2) An increased rate of expression of Lewis group antigens, particularly Lewis(b) antigen, was observed in endometrial cancers compared with its expression in normal endometria. 3) Lc4 and nLc4 antigens were detected in endometrial cancers at rates of 41.9% and 38.7%, respectively, these expressions being increased compared with those in normal endometria. 4) These results suggest that a highly abnormal expression of blood group-related antigens in endometrial cancers occurs not only at the level of A, B, and H antigens and Lewis group antigens, but also at the level of their precursor Lc4 and nLc4 antigens. 5) Lewis(a), Lewis(b), and Lc4 antigens, built on the type-1 chain, are more specific to endometrial cancers than their respective positional isomers, Lewis(x), Lewis(y), and nLc4 antigens, built on the type-2 chain.

  15. Allosensibilisation to erythrocyte antigens (literature review

    Directory of Open Access Journals (Sweden)

    N. V. Mineeva

    2015-01-01

    Full Text Available In this article literature review of the causes of allosensibilisation to erythrocyte antigens are presented. It is shown that the ability to produce antierythrocyte antibodies is affected by many factors, principal of whom it is difficult to identify. For the allosensibilisation development requires genetically determined differences in erythrocyte antigens phenotypes of donor and recipient, mother and fetus, which can lead to immune response and antibodies production. The biochemical nature of erythrocyte antigens, antigen dose (the amount of transfused doses, the number of antigens determinants on donor and fetus erythrocytes, the number of pregnancies are important. Individual patient characteristics: age, gender, diseases, the use of immunosuppressive therapy and the presence of inflammatory processes, are also relevant. Note that antibody to one erythrocyte antigens have clinical value, and to the other – have no. The actual data about frequency of clinically significant antibodies contribute to the development of post-transfusion hemolytic complications prophylaxis as well as the improvement of laboratory diagnosis of hemolytic disease of the newborn in the presence of maternal antierythrocyte antibodies.

  16. Designed ankyrin repeat proteins: a new approach to mimic complex antigens for diagnostic purposes?

    Directory of Open Access Journals (Sweden)

    Stefanie Hausammann

    Full Text Available Inhibitory antibodies directed against coagulation factor VIII (FVIII can be found in patients with acquired and congenital hemophilia A. Such FVIII-inhibiting antibodies are routinely detected by the functional Bethesda Assay. However, this assay has a low sensitivity and shows a high inter-laboratory variability. Another method to detect antibodies recognizing FVIII is ELISA, but this test does not allow the distinction between inhibitory and non-inhibitory antibodies. Therefore, we aimed at replacing the intricate antigen FVIII by Designed Ankyrin Repeat Proteins (DARPins mimicking the epitopes of FVIII inhibitors. As a model we used the well-described inhibitory human monoclonal anti-FVIII antibody, Bo2C11, for the selection on DARPin libraries. Two DARPins were selected binding to the antigen-binding site of Bo2C11, which mimic thus a functional epitope on FVIII. These DARPins inhibited the binding of the antibody to its antigen and restored FVIII activity as determined in the Bethesda assay. Furthermore, the specific DARPins were able to recognize the target antibody in human plasma and could therefore be used to test for the presence of Bo2C11-like antibodies in a large set of hemophilia A patients. These data suggest, that our approach might be used to isolate epitopes from different sets of anti-FVIII antibodies in order to develop an ELISA-based screening assay allowing the distinction of inhibitory and non-inhibitory anti-FVIII antibodies according to their antibody signatures.

  17. Radioimmunoassay for hepatitis B core antigen

    International Nuclear Information System (INIS)

    Sagnelli, E.; Pereira, C.; Triolo, G.; Vernace, S.; Paronetto, F.

    1982-01-01

    Serum hepatitis B core antigen (HBcAg) is an important marker of hepatitis B virus replication. We describe an easy, sensitive radioimmunoassay for determination of HBcAg in detergent-treated serum pellets containing Dane particles. Components of a commercial kit for anticore determination are used, and HBcAG is measured by competitive inhibition of binding of 125 I-labeled antibodies to HBcAg with HBcAg-coated beads. We assayed for HBcAG in the sera of 49 patients with hepatitis B surface antigen (HBsAg)-positive chronic hepatitis, 50 patients with HBsAg-negative chronic hepatitis, and 30 healthy volunteers. HBcAg was detected in 41% of patients with HBsAg-positive chronic hepatitis but not in patients with HBsAg-negative chronic hepatitis. Hepatitis Be antigen (an antigen closely associated with the core of Dane particles) determined in the same sera by radioimmunoassay, was not detected in 50% of HBcAg-positive sera

  18. Bone marrow-derived thymic antigen-presenting cells determine self-recognition of Ia-restricted T lymphocytes

    International Nuclear Information System (INIS)

    Longo, D.L.; Kruisbeek, A.M.; Davis, M.L.; Matis, L.A.

    1985-01-01

    The authors previously have demonstrated that in radiation-induced bone marrow chimeras, T-cell self-Ia restriction specificity appeared to correlate with the phenotype of the bone marrow-derived antigen-presenting (or dendritic) cell in the thymus during T-cell development. However, these correlations were necessarily indirect because of the difficulty in assaying thymic function directly by adult thymus transplant, which has in the past been uniformly unsuccessful. They now report success in obtaining functional T cells from nude mice grafted with adult thymuses reduced in size by treatment of the thymus donor with anti-thymocyte globulin and cortisone. When (B10 Scn X B10.D2)F1 nude mice (I-Ab,d) are given parental B10.D2 (I-Ad) thymus grafts subcutaneously, their T cells are restricted to antigen recognition in association with I-Ad gene products but not I-Ab gene products. Furthermore, thymuses from (B10 X B10.D2)F1 (I-Ab,d)----B10 (I-Ab) chimeras transplanted 6 months or longer after radiation (a time at which antigen-presenting cell function is of donor bone marrow phenotype) into (B10 X B10.D2)F1 nude mice generate T cells restricted to antigen recognition in association with both I-Ad and I-Ab gene products. Thymuses from totally allogeneic bone marrow chimeras appear to generate T cells of bone marrow donor and thymic host restriction specificity. Thus, when thymus donors are radiation-induced bone marrow chimeras, the T-cell I-region restriction of the nude mice recipients is determined at least in part by the phenotype of the bone marrow-derived thymic antigen presenting cells or dendritic cells in the chimeric thymus

  19. Properties of the simian virus 40 (SV40) large T antigens encoded by SV40 mutants with deletions in gene A.

    Science.gov (United States)

    Cole, C N; Tornow, J; Clark, R; Tjian, R

    1986-01-01

    The biochemical properties of the large T antigens encoded by simian virus 40 (SV40) mutants with deletions at DdeI sites in the SV40 A gene were determined. Mutant large T antigens containing only the first 138 to 140 amino acids were unable to bind to the SV40 origin of DNA replication as were large T antigens containing at their COOH termini 96 or 97 amino acids encoded by the long open reading frame located between 0.22 and 0.165 map units (m.u.). All other mutant large T antigens were able to bind to the SV40 origin of replication. Mutants with in-phase deletions at 0.288 and 0.243 m.u. lacked ATPase activity, but ATPase activity was normal in mutants lacking origin-binding activity. The 627-amino acid large T antigen encoded by dlA2465, with a deletion at 0.219 m.u., was the smallest large T antigen displaying ATPase activity. Mutant large T antigens with the alternate 96- or 97-amino acid COOH terminus also lacked ATPase activity. All mutant large T antigens were found in the nuclei of infected cells; a small amount of large T with the alternate COOH terminus was also located in the cytoplasm. Mutant dlA2465 belonged to the same class of mutants as dlA2459. It was unable to form plaques on CV-1p cells at 37 or 32 degrees C but could form plaques on BSC-1 monolayers at 37 degrees C but not at 32 degrees C. It was positive for viral DNA replication and showed intracistronic complementation with any group A mutant whose large T antigen contained a normal carboxyl terminus. These findings and those of others suggest that both DNA binding and ATPase activity are required for the viral DNA replication function of large T antigen, that these two activities must be located on the same T antigen monomer, and that these two activities are performed by distinct domains of the polypeptide. These domains are distinct and separable from the domain affected by the mutation of dlA2465 and indicate that SV40 large T antigen is made up of at least three separate functional

  20. Differential recognition and hydrolysis of host carbohydrate antigens by Streptococcus pneumoniae family 98 glycoside hydrolases.

    Science.gov (United States)

    Higgins, Melanie A; Whitworth, Garrett E; El Warry, Nahida; Randriantsoa, Mialy; Samain, Eric; Burke, Robert D; Vocadlo, David J; Boraston, Alisdair B

    2009-09-18

    The presence of a fucose utilization operon in the Streptococcus pneumoniae genome and its established importance in virulence indicates a reliance of this bacterium on the harvesting of host fucose-containing glycans. The identities of these glycans, however, and how they are harvested is presently unknown. The biochemical and high resolution x-ray crystallographic analysis of two family 98 glycoside hydrolases (GH98s) from distinctive forms of the fucose utilization operon that originate from different S. pneumoniae strains reveal that one enzyme, the predominant type among pneumococcal isolates, has a unique endo-beta-galactosidase activity on the LewisY antigen. Altered active site topography in the other species of GH98 enzyme tune its endo-beta-galactosidase activity to the blood group A and B antigens. Despite their different specificities, these enzymes, and by extension all family 98 glycoside hydrolases, use an inverting catalytic mechanism. Many bacterial and viral pathogens exploit host carbohydrate antigens for adherence as a precursor to colonization or infection. However, this is the first evidence of bacterial endoglycosidase enzymes that are known to play a role in virulence and are specific for distinct host carbohydrate antigens. The strain-specific distribution of two distinct types of GH98 enzymes further suggests that S. pneumoniae strains may specialize to exploit host-specific antigens that vary from host to host, a factor that may feature in whether a strain is capable of colonizing a host or establishing an invasive infection.

  1. Radioimmunoassay and some properties of human antibodies to hepatitis B core antigen

    Energy Technology Data Exchange (ETDEWEB)

    Neurath, A R; Szmuness, W; Stevens, C E; Strick, N; Harley, E J [New York Blood Center, N.Y. (USA)

    1978-03-01

    A solid-phase radioimmunoassay for antibodies to hepatitis B core antigen (anti-HBsub(c)) is described. Polystyrene beads coated with anti-HBsub(c), hepatitis B core antigen prepared from pooled sera of humans infected with hepatitis B virus (HBV) and /sup 125/I-labelled anti-HBsub(c) were used for the test. Distinct patterns of development and changes of anti-HBsub(c) and their immunological properties are all related to variations of other markers specific for HBV infections. Knowledge concerning the detailed features of the immune response to hepatitis B core antigen may provide deeper insight into the pathogenesis of HBV infections.

  2. Antigen-Specific Th17 Cells Are Primed by Distinct and Complementary Dendritic Cell Subsets in Oropharyngeal Candidiasis.

    Directory of Open Access Journals (Sweden)

    Kerstin Trautwein-Weidner

    2015-10-01

    Full Text Available Candida spp. can cause severe and chronic mucocutaneous and systemic infections in immunocompromised individuals. Protection from mucocutaneous candidiasis depends on T helper cells, in particular those secreting IL-17. The events regulating T cell activation and differentiation toward effector fates in response to fungal invasion in different tissues are poorly understood. Here we generated a Candida-specific TCR transgenic mouse reactive to a novel endogenous antigen that is conserved in multiple distant species of Candida, including the clinically highly relevant C. albicans and C. glabrata. Using TCR transgenic T cells in combination with an experimental model of oropharyngeal candidiasis (OPC we investigated antigen presentation and Th17 priming by different subsets of dendritic cells (DCs present in the infected oral mucosa. Candida-derived endogenous antigen accesses the draining lymph nodes and is directly presented by migratory DCs. Tissue-resident Flt3L-dependent DCs and CCR2-dependent monocyte-derived DCs collaborate in antigen presentation and T cell priming during OPC. In contrast, Langerhans cells, which are also present in the oral mucosa and have been shown to prime Th17 cells in the skin, are not required for induction of the Candida-specific T cell response upon oral challenge. This highlights the functional compartmentalization of specific DC subsets in different tissues. These data provide important new insights to our understanding of tissue-specific antifungal immunity.

  3. Antigen Availability Shapes T Cell Differentiation and Function during Tuberculosis.

    Science.gov (United States)

    Moguche, Albanus O; Musvosvi, Munyaradzi; Penn-Nicholson, Adam; Plumlee, Courtney R; Mearns, Helen; Geldenhuys, Hennie; Smit, Erica; Abrahams, Deborah; Rozot, Virginie; Dintwe, One; Hoff, Søren T; Kromann, Ingrid; Ruhwald, Morten; Bang, Peter; Larson, Ryan P; Shafiani, Shahin; Ma, Shuyi; Sherman, David R; Sette, Alessandro; Lindestam Arlehamn, Cecilia S; McKinney, Denise M; Maecker, Holden; Hanekom, Willem A; Hatherill, Mark; Andersen, Peter; Scriba, Thomas J; Urdahl, Kevin B

    2017-06-14

    CD4 T cells are critical for protective immunity against Mycobacterium tuberculosis (Mtb), the cause of tuberculosis (TB). Yet to date, TB vaccine candidates that boost antigen-specific CD4 T cells have conferred little or no protection. Here we examined CD4 T cell responses to two leading TB vaccine antigens, ESAT-6 and Ag85B, in Mtb-infected mice and in vaccinated humans with and without underlying Mtb infection. In both species, Mtb infection drove ESAT-6-specific T cells to be more differentiated than Ag85B-specific T cells. The ability of each T cell population to control Mtb in the lungs of mice was restricted for opposite reasons: Ag85B-specific T cells were limited by reduced antigen expression during persistent infection, whereas ESAT-6-specific T cells became functionally exhausted due to chronic antigenic stimulation. Our findings suggest that different vaccination strategies will be required to optimize protection mediated by T cells recognizing antigens expressed at distinct stages of Mtb infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Merkel cell polyomavirus in Merkel cell carcinogenesis: small T antigen-mediates c-Jun phosphorylation.

    Science.gov (United States)

    Wu, Julie H; Simonette, Rebecca A; Nguyen, Harrison P; Rady, Peter L; Tyring, Stephen K

    2016-06-01

    Merkel cell carcinoma (MCC) is a highly aggressive neuroendocrine skin cancer associated with the Merkel cell polyomavirus (MCPyV). The MCPyV genome, which is clonally integrated in the majority of MCCs, encodes the regulatory small T (sT) antigen. Previously, reports have established MCPyV sT antigen as a potent oncogene capable of inducing cell transformation. In the current study, we demonstrate a distinct role for c-Jun hyperactivation in MCPyV sT antigen pathogenesis. As MCPyV sT antigen's association with aggressive cancer growth has been previously established, this finding may represent a potential therapeutic target for the treatment of MCCs.

  5. On Determinatives and the Category-Function Distinction: A Reply to Brett Reynolds

    Science.gov (United States)

    Lenchuk, Iryna; Ahmed, Amer

    2014-01-01

    This article examines the arguments made in the article "Determiners, Feline Marsupials, and the Category-Function Distinction: A Critique of ELT Grammars" by Brett Reynolds recently published in the "TESL Canada Journal" (2013). In our response, we demonstrate that the author's arguments are problematic on both…

  6. Rationalization and Design of the Complementarity Determining Region Sequences in an Antibody-Antigen Recognition Interface

    Science.gov (United States)

    Chen, Ing-Chien; Lee, Yu-Ching; Chen, Jun-Bo; Tsai, Keng-Chang; Chen, Ching-Tai; Chang, Jeng-Yih; Yang, Ei-Wen; Hsu, Po-Chiang; Jian, Jhih-Wei; Hsu, Hung-Ju; Chang, Hung-Ju; Hsu, Wen-Lian; Huang, Kai-Fa; Ma, Alex Che; Yang, An-Suei

    2012-01-01

    Protein-protein interactions are critical determinants in biological systems. Engineered proteins binding to specific areas on protein surfaces could lead to therapeutics or diagnostics for treating diseases in humans. But designing epitope-specific protein-protein interactions with computational atomistic interaction free energy remains a difficult challenge. Here we show that, with the antibody-VEGF (vascular endothelial growth factor) interaction as a model system, the experimentally observed amino acid preferences in the antibody-antigen interface can be rationalized with 3-dimensional distributions of interacting atoms derived from the database of protein structures. Machine learning models established on the rationalization can be generalized to design amino acid preferences in antibody-antigen interfaces, for which the experimental validations are tractable with current high throughput synthetic antibody display technologies. Leave-one-out cross validation on the benchmark system yielded the accuracy, precision, recall (sensitivity) and specificity of the overall binary predictions to be 0.69, 0.45, 0.63, and 0.71 respectively, and the overall Matthews correlation coefficient of the 20 amino acid types in the 24 interface CDR positions was 0.312. The structure-based computational antibody design methodology was further tested with other antibodies binding to VEGF. The results indicate that the methodology could provide alternatives to the current antibody technologies based on animal immune systems in engineering therapeutic and diagnostic antibodies against predetermined antigen epitopes. PMID:22457753

  7. Presentation of lipid antigens to T cells.

    Science.gov (United States)

    Mori, Lucia; De Libero, Gennaro

    2008-04-15

    T cells specific for lipid antigens participate in regulation of the immune response during infections, tumor immunosurveillance, allergy and autoimmune diseases. T cells recognize lipid antigens as complexes formed with CD1 antigen-presenting molecules, thus resembling recognition of MHC-peptide complexes. The biophysical properties of lipids impose unique mechanisms for their delivery, internalization into antigen-presenting cells, membrane trafficking, processing, and loading of CD1 molecules. Each of these steps is controlled at molecular and celular levels and determines lipid immunogenicity. Lipid antigens may derive from microbes and from the cellular metabolism, thus allowing the immune system to survey a large repertoire of immunogenic molecules. Recognition of lipid antigens facilitates the detection of infectious agents and the initiation of responses involved in immunoregulation and autoimmunity. This review focuses on the presentation mechanisms and specific recognition of self and bacterial lipid antigens and discusses the important open issues.

  8. Antigenicity and diagnostic potential of vaccine candidates in human Chagas disease.

    Directory of Open Access Journals (Sweden)

    Shivali Gupta

    Full Text Available Chagas disease, caused by Trypanosoma cruzi, is endemic in Latin America and an emerging infectious disease in the US and Europe. We have shown TcG1, TcG2, and TcG4 antigens elicit protective immunity to T. cruzi in mice and dogs. Herein, we investigated antigenicity of the recombinant proteins in humans to determine their potential utility for the development of next generation diagnostics for screening of T. cruzi infection and Chagas disease.Sera samples from inhabitants of the endemic areas of Argentina-Bolivia and Mexico-Guatemala were analyzed in 1(st-phase for anti-T. cruzi antibody response by traditional serology tests; and in 2(nd-phase for antibody response to the recombinant antigens (individually or mixed by an ELISA. We noted similar antibody response to candidate antigens in sera samples from inhabitants of Argentina and Mexico (n=175. The IgG antibodies to TcG1, TcG2, and TcG4 (individually and TcG(mix were present in 62-71%, 65-78% and 72-82%, and 89-93% of the subjects, respectively, identified to be seropositive by traditional serology. Recombinant TcG1- (93.6%, TcG2- (96%, TcG4- (94.6% and TcG(mix- (98% based ELISA exhibited significantly higher specificity compared to that noted for T. cruzi trypomastigote-based ELISA (77.8% in diagnosing T. cruzi-infection and avoiding cross-reactivity to Leishmania spp. No significant correlation was noted in the sera levels of antibody response and clinical severity of Chagas disease in seropositive subjects.Three candidate antigens were recognized by antibody response in chagasic patients from two distinct study sites and expressed in diverse strains of the circulating parasites. A multiplex ELISA detecting antibody response to three antigens was highly sensitive and specific in diagnosing T. cruzi infection in humans, suggesting that a diagnostic kit based on TcG1, TcG2 and TcG4 recombinant proteins will be useful in diverse situations.

  9. The value of serum Hepatitis B surface antigen quantification in ...

    African Journals Online (AJOL)

    The value of serum Hepatitis B surface antigen quantification in determining viralactivity in chronic Hepatitis B virus infection. ... ofCHB andalso higher in hepatitis e antigen positive patients compared to hepatitis e antigen negative patients.

  10. Analysis of antigen-specific B-cell memory directly ex vivo.

    Science.gov (United States)

    McHeyzer-Williams, Louise J; McHeyzer-Williams, Michael G

    2004-01-01

    Helper T-cell-regulated B-cell memory develops in response to initial antigen priming as a cellular product of the germinal center (GC) reaction. On antigen recall, memory response precursors expand rapidly with exaggerated differentiation into plasma cells to produce the high-titer, high-affinity antibody(Ab) that typifies the memory B-cell response in vivo. We have devised a high-resolution flow cytometric strategy to quantify the emergence and maintenance of antigen-specific memory B cells directly ex vivo. Extended cell surface phenotype establishes a level of cellular diversity not previously appreciated for the memory B-cell compartment. Using an "exclusion transfer" strategy, we ascertain the capacity of two distinct memory B-cell populations to transfer antigen-specific memory into naive adoptive hosts. Finally, we sequence expressed messenger ribonucleic acid (mRNA) from single cells within the population to estimate the level of somatic hypermutation as the best molecular indicator of B-cell memory. In this chapter, we describe the methods used in each of these four sections that serve to provide high-resolution quantification of antigen-specific B-cell memory responses directly ex vivo.

  11. Tissue polypeptide-specific antigen (TPS) determinations before and during intermittent maximal androgen blockade in patients with metastatic prostatic carcinoma

    NARCIS (Netherlands)

    Kil, P. J. M.; Goldschmidt, H. M. J.; Wieggers, B. J. A.; Kariakine, O. B.; Studer, U. E.; Whelan, P.; Hetherington, J.; de Reijke, Th M.; Hoekstra, J. W.; Collette, L.

    2003-01-01

    To evaluate the prognostic significance of serially measured tissue polypeptide-specific antigen (TPS) levels in patients with metastatic prostatic carcinoma treated with intermittent maximal androgen blockade (MAB). To determine its value with respect to predicting response to treatment and time to

  12. COLONOSCOPY AND CARCINOEMBRYONIC ANTIGEN VARIATIONS

    Directory of Open Access Journals (Sweden)

    Rita G SOUSA

    2014-03-01

    Full Text Available Context Colonoscopy is essential for synchronous and metachronous cancer detection. Carcinoembryonic antigen is a colorectal cancer tumor marker, important as a follow-up tool in patients with previous colorectal cancer. False-positive carcinoembryonic antigen elevation results in multiples exams and in patient anxiety. In literature, there is reference to transient carcinoembryonic antigen increase with colonoscopy. Objective To evaluate the influence of bowel preparation and colonoscopy in carcinoembryonic antigen blood levels. Methods We prospectively studied subjects that underwent routine colonoscopy in our institution. Blood samples were collected (1 before bowel cleaning, (2 before colonoscopy and (3 immediately after colonoscopy. Blood carcinoembryonic antigen levels were determined by “Sandwich” immunoassay. The statistical methods used were the paired t-test and ANOVA. Results Thirty-seven patients (22M/15F were included; age range 28-84 (mean 56 years. Mean carcinoembryonic antigen values were 1.9, 2 and 1.8 for (1, (2 and (3, respectively. An increase in value (2 compared with (1 was observed in 20/37 patients (P = 0.018, mainly in younger patients and in patients requiring more endoluminal interventions. In 29/37 patients, the CEA value decreased from (2 to (3 (P = 1.3x10-7. Conclusions A trend for carcinoembryonic antigen increase after bowel cleaning was observed, especially in younger patients and in patients with more endoluminal interventions, but without clinical meaning.

  13. Genetic distribution of noncapsular meningococcal group B vaccine antigens in Neisseria lactamica.

    Science.gov (United States)

    Lucidarme, Jay; Gilchrist, Stefanie; Newbold, Lynne S; Gray, Stephen J; Kaczmarski, Edward B; Richardson, Lynne; Bennett, Julia S; Maiden, Martin C J; Findlow, Jamie; Borrow, Ray

    2013-09-01

    The poor immunogenicity of the meningococcal serogroup B (MenB) capsule has led to the development of vaccines targeting subcapsular antigens, in particular the immunodominant and diverse outer membrane porin, PorA. These vaccines are largely strain specific; however, they offer limited protection against the diverse MenB-associated diseases observed in many industrialized nations. To broaden the scope of its protection, the multicomponent vaccine (4CMenB) incorporates a PorA-containing outer membrane vesicle (OMV) alongside relatively conserved recombinant protein components, including factor H-binding protein (fHbp), Neisseria adhesin A (NadA), and neisserial heparin-binding antigen (NHBA). The expression of PorA is unique to meningococci (Neisseria meningitidis); however, many subcapsular antigens are shared with nonpathogenic members of the genus Neisseria that also inhabit the nasopharynx. These organisms may elicit cross-protective immunity against meningococci and/or occupy a niche that might otherwise accommodate pathogens. The potential for 4CMenB responses to impact such species (and vice versa) was investigated by determining the genetic distribution of the primary 4CMenB antigens among diverse members of the common childhood commensal, Neisseria lactamica. All the isolates possessed nhba but were devoid of fhbp and nadA. The nhba alleles were mainly distinct from but closely related to those observed among a representative panel of invasive MenB isolates from the same broad geographic region. We made similar findings for the immunogenic typing antigen, FetA, which constitutes a major part of the 4CMenB OMV. Thus, 4CMenB vaccine responses may impact or be impacted by nasopharyngeal carriage of commensal neisseriae. This highlights an area for further research and surveillance should the vaccine be routinely implemented.

  14. Antigen storage compartments in mature dendritic cells facilitate prolonged cytotoxic T lymphocyte cross-priming capacity.

    Science.gov (United States)

    van Montfoort, Nadine; Camps, Marcel G; Khan, Selina; Filippov, Dmitri V; Weterings, Jimmy J; Griffith, Janice M; Geuze, Hans J; van Hall, Thorbald; Verbeek, J Sjef; Melief, Cornelis J; Ossendorp, Ferry

    2009-04-21

    Dendritic cells (DCs) are crucial for priming of naive CD8(+) T lymphocytes to exogenous antigens, so-called "cross-priming." We report that exogenous protein antigen can be conserved for several days in mature DCs, coinciding with strong cytotoxic T lymphocyte cross-priming potency in vivo. After MHC class I peptide elution, protein antigen-derived peptide presentation is efficiently restored, indicating the presence of an intracellular antigen depot. We characterized this depot as a lysosome-like organelle, distinct from MHC class II compartments and recently described early endosomal compartments that allow acute antigen presentation in MHC class I. The storage compartments we report here facilitate continuous supply of MHC class I ligands. This mechanism ensures sustained cross-presentation by DCs, despite the short-lived expression of MHC class I-peptide complexes at the cell surface.

  15. Immunophenotyping of Waldenstroms macroglobulinemia cell lines reveals distinct patterns of surface antigen expression: potential biological and therapeutic implications.

    Directory of Open Access Journals (Sweden)

    Aneel Paulus

    Full Text Available Waldenströms macroglobulinemia (WM is a subtype of Non-Hodgkin's lymphoma in which the tumor cell population is markedly heterogeneous, consisting of immunoglobulin-M secreting B-lymphocytes, plasmacytoid lymphocytes and plasma cells. Due to rarity of disease and scarcity of reliable preclinical models, many facets of WM molecular and phenotypic architecture remain incompletely understood. Currently, there are 3 human WM cell lines that are routinely used in experimental studies, namely, BCWM.1, MWCL-1 and RPCI-WM1. During establishment of RPCI-WM1, we observed loss of the CD19 and CD20 antigens, which are typically present on WM cells. Intrigued by this observation and in an effort to better define the immunophenotypic makeup of this cell line, we conducted a more comprehensive analysis for the presence or absence of other cell surface antigens that are present on the RPCI-WM1 model, as well as those on the two other WM cell lines, BCWM.1 and MWCL-1. We examined expression of 65 extracellular and 4 intracellular antigens, comprising B-cell, plasma cell, T-cell, NK-cell, myeloid and hematopoietic stem cell surface markers by flow cytometry analysis. RPCI-WM1 cells demonstrated decreased expression of CD19, CD20, and CD23 with enhanced expression of CD28, CD38 and CD184, antigens that were differentially expressed on BCWM.1 and MWCL-1 cells. Due to increased expression of CD184/CXCR4 and CD38, RPCI-WM1 represents a valuable model in which to study the effects anti-CXCR4 or anti-CD38 targeted therapies that are actively being developed for treatment of hematologic cancers. Overall, differences in surface antigen expression across the 3 cell lines may reflect the tumor clone population predominant in the index patients, from whom the cell lines were developed. Our analysis defines the utility of the most commonly employed WM cell lines as based on their immunophenotype profiles, highlighting unique differences that can be further studied for

  16. Myeloid antigens in childhood lymphoblastic leukemia:clinical data point to regulation of CD66c distinct from other myeloid antigens

    Directory of Open Access Journals (Sweden)

    Madzo Jozef

    2005-04-01

    Full Text Available Abstract Background Aberrant expression of myeloid antigens (MyAgs on acute lymphoblastic leukemia (ALL cells is a well-documented phenomenon, although its regulating mechanisms are unclear. MyAgs in ALL are interpreted e.g. as hallmarks of early differentiation stage and/or lineage indecisiveness. Granulocytic marker CD66c – Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6 is aberrantly expressed on ALL with strong correlation to genotype (negative in TEL/AML1 and MLL/AF4, positive in BCR/ABL and hyperdiploid cases. Methods In a cohort of 365 consecutively diagnosed Czech B-precursor ALL patients, we analyze distribution of MyAg+ cases and mutual relationship among CD13, CD15, CD33, CD65 and CD66c. The most frequent MyAg (CD66c is studied further regarding its stability from diagnosis to relapse, prognostic significance and regulation of surface expression. For the latter, flow cytometry, Western blot and quantitative RT-PCR on sorted cells is used. Results We show CD66c is expressed in 43% patients, which is more frequent than other MyAgs studied. In addition, CD66c expression negatively correlates with CD13 (p Conclusion In contrast to general notion we show that different MyAgs in lymphoblastic leukemia represent different biological circumstances. We chose the most frequent and tightly genotype-associated MyAg CD66c to show its stabile expression in patients from diagnosis to relapse, which differs from what is known on the other MyAgs. Surface expression of CD66c is regulated at the gene transcription level, in contrast to previous reports.

  17. Identification of a novel splice variant form of the influenza A virus M2 ion channel with an antigenically distinct ectodomain.

    Directory of Open Access Journals (Sweden)

    Helen M Wise

    Full Text Available Segment 7 of influenza A virus produces up to four mRNAs. Unspliced transcripts encode M1, spliced mRNA2 encodes the M2 ion channel, while protein products from spliced mRNAs 3 and 4 have not previously been identified. The M2 protein plays important roles in virus entry and assembly, and is a target for antiviral drugs and vaccination. Surprisingly, M2 is not essential for virus replication in a laboratory setting, although its loss attenuates the virus. To better understand how IAV might replicate without M2, we studied the reversion mechanism of an M2-null virus. Serial passage of a virus lacking the mRNA2 splice donor site identified a single nucleotide pseudoreverting mutation, which restored growth in cell culture and virulence in mice by upregulating mRNA4 synthesis rather than by reinstating mRNA2 production. We show that mRNA4 encodes a novel M2-related protein (designated M42 with an antigenically distinct ectodomain that can functionally replace M2 despite showing clear differences in intracellular localisation, being largely retained in the Golgi compartment. We also show that the expression of two distinct ion channel proteins is not unique to laboratory-adapted viruses but, most notably, was also a feature of the 1983 North American outbreak of H5N2 highly pathogenic avian influenza virus. In identifying a 14th influenza A polypeptide, our data reinforce the unexpectedly high coding capacity of the viral genome and have implications for virus evolution, as well as for understanding the role of M2 in the virus life cycle.

  18. Mass spectrometric characterization of circulating and functional antigens derived from piperacillin in patients with cystic fibrosis1

    Science.gov (United States)

    Whitaker, Paul; Meng, Xiaoli; Lavergne, Sidonie N.; El-Ghaiesh, Sabah; Monshi, Manal; Earnshaw, Caroline; Peckham, Daniel; Gooi, Jimmy; Conway, Steve; Pirmohamed, Munir; Jenkins, Rosalind E.; Naisbitt, Dean J.; Park, B. Kevin

    2011-01-01

    A mechanistic understanding of the relationship between the chemistry of drug antigen formation and immune function is lacking. Thus, mass spectrometric methods were employed to detect and fully characterize circulating antigens derived from piperacillin in patients undergoing therapy and the nature of the drug derived-epitopes on protein which can function as an antigen to stimulate T-cells. Albumin modification with piperacillin in vitro resulted in the formation of two distinct haptens, one formed directly from piperacillin and a second in which the dioxopiperazine ring had undergone hydrolysis. Modification was time- and concentration-dependent, with selective modification of Lys541 observed at low concentrations, whereas at higher concentrations up to 13/59 lysine residues were modified, four of which (Lys190, 195, 432 and 541) were detected in patients’ plasma. Piperacillin-specific T-lymphocyte responses (proliferation, cytokines and granzyme-B release) were detected ex vivo with cells from hypersensitive patients, and analysis of incubation medium showed that modification of the same lysine residues in albumin occurred in situ. The antigenicity of piperacillin-modified albumin was confirmed by stimulation of T-cells with characterized synthetic conjugates. Analysis of minimally-modified T-cell stimulatory albumin conjugates revealed peptide sequences incorporating Lys190, 432 and 541 as principal functional epitopes for T-cells. This study has characterized the multiple haptenic structures on albumin in patients, and showed that they constitute functional antigenic determinants for T-cells. PMID:21606251

  19. Cationic liposomes promote antigen cross-presentation in dendritic cells by alkalizing the lysosomal pH and limiting the degradation of antigens

    Directory of Open Access Journals (Sweden)

    Gao J

    2017-02-01

    Full Text Available Jie Gao,1–3 Lukasz J Ochyl,1,3 Ellen Yang,4 James J Moon1,3,5 1Department of Pharmaceutical Sciences, University of Michigan, Ann Arbor, MI, USA; 2Department of Pharmaceutical Sciences, School of Pharmacy, Second Military Medical University, Shanghai, People’s Republic of China; 3Biointerfaces Institute, 4Department of Chemistry, 5Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA Abstract: Cationic liposomes (CLs have been widely examined as vaccine delivery nanoparticles since they can form complexes with biomacromolecules, promote delivery of antigens and adjuvant molecules to antigen-presenting cells (APCs, and mediate cellular uptake of vaccine components. CLs are also known to trigger antigen cross-presentation – the process by which APCs internalize extracellular protein antigens, degrade them into minimal CD8+ T-cell epitopes, and present them in the context of major histocompatibility complex-I (MHC-I. However, the precise mechanisms behind CL-mediated induction of cross-presentation and cross-priming of CD8+ T-cells remain to be elucidated. In this study, we have developed two distinct CL systems and examined their impact on the lysosomal pH in dendritic cells (DCs, antigen degradation, and presentation of peptide:MHC-I complexes to antigen-specific CD8+ T-cells. To achieve this, we have used 3β-[N-(N',N'-dimethylaminoethane-carbamoyl] cholesterol (DC-Chol and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP as the prototypical components of CLs with tertiary amine groups and compared the effect of CLs and anionic liposomes on lysosomal pH, antigen degradation, and cross-presentation by DCs. Our results showed that CLs, but not anionic liposomes, elevated the lysosomal pH in DCs and reduced antigen degradation, thereby promoting cross-presentation and cross-priming of CD8+ T-cell responses. These studies shed new light on CL-mediated cross-presentation and suggest that intracellular fate of vaccine

  20. Original antigenic sin responses to influenza viruses.

    Science.gov (United States)

    Kim, Jin Hyang; Skountzou, Ioanna; Compans, Richard; Jacob, Joshy

    2009-09-01

    Most immune responses follow Burnet's rule in that Ag recruits specific lymphocytes from a large repertoire and induces them to proliferate and differentiate into effector cells. However, the phenomenon of "original antigenic sin" stands out as a paradox to Burnet's rule of B cell engagement. Humans, upon infection with a novel influenza strain, produce Abs against older viral strains at the expense of responses to novel, protective antigenic determinants. This exacerbates the severity of the current infection. This blind spot of the immune system and the redirection of responses to the "original Ag" rather than to novel epitopes were described fifty years ago. Recent reports have questioned the existence of this phenomenon. Hence, we revisited this issue to determine the extent to which original antigenic sin is induced by variant influenza viruses. Using two related strains of influenza A virus, we show that original antigenic sin leads to a significant decrease in development of protective immunity and recall responses to the second virus. In addition, we show that sequential infection of mice with two live influenza virus strains leads to almost exclusive Ab responses to the first viral strain, suggesting that original antigenic sin could be a potential strategy by which variant influenza viruses subvert the immune system.

  1. ANTIGENICITY OF COW'S MILK PROTEINS IN TWO ANIMAL MODELS

    OpenAIRE

    T.R. Neyestani; M. Djalali M. I'ezeshki

    2000-01-01

    Antigenicity of proteins found in cow's milk is age dependent. This is primarily due to infants possessing a more permeable intestinal wall than that in adults. Thus infants may acquire cow's milk allergy during their first year of life. While milk antigen specific IgE may cause allergy in susceptible subjects, there is some evidence indicating that milk antigen specific IgG may play some role in chronic disease development. The puropose of this study was to determine the antigenicity of cow'...

  2. Identification of antigenic proteins of setaria cervi by immunoblotting technique

    International Nuclear Information System (INIS)

    Kaushal, N.A.; Kaushal, D.C.; Ghatak, S.

    1987-01-01

    Identification and characterization of antigenic proteins of Setaria cervi (bovine filarial parasite) adults and microfilariae was done by immunoblotting technique using hyperimmune rabbit sera against S. cervi and Brugia malayi. The antigens recognized by these sera were detected by using 125 I protein-A followed by autoradiography. Fifteen different antigens were observed to be common between adult and microfilarial stages of the parasite. Some stage specific antigens were also identified. Many antigens of S. cervi adults and microfilariae were also recognized by rabbit anti-B.malayi serum showing the existence of common antigenic determinants between the bovine and human filarial parasites

  3. Reagents for radioimmunological determination of carcinoembryonic antigen (CEA)

    International Nuclear Information System (INIS)

    Albert, Z.; Balbierz, H.; Breberowicz, J.

    1978-01-01

    The work was undertaken to prepare the reagents for carcinoembryonic antigen (CEA) radioimmunoassay with double antibody method. The CEA standard of high immunoreactivity was prepared and purified. The purified CEA was used for immunozation of goats. The goat anti - CEA sera were received. IgG fraction from normal goat serum was purified and used for the production of horse anti-goat IgG serum which was then used in the radioimmunoassay of CEA. The labelling of CEA with iodine-125 has been carried out be means of the enzymatic method.(Z.R.)

  4. Viral sequestration of antigen subverts cross presentation to CD8(+ T cells.

    Directory of Open Access Journals (Sweden)

    Eric F Tewalt

    2009-05-01

    Full Text Available Virus-specific CD8(+ T cells (T(CD8+ are initially triggered by peptide-MHC Class I complexes on the surface of professional antigen presenting cells (pAPC. Peptide-MHC complexes are produced by two spatially distinct pathways during virus infection. Endogenous antigens synthesized within virus-infected pAPC are presented via the direct-presentation pathway. Many viruses have developed strategies to subvert direct presentation. When direct presentation is blocked, the cross-presentation pathway, in which antigen is transferred from virus-infected cells to uninfected pAPC, is thought to compensate and allow the generation of effector T(CD8+. Direct presentation of vaccinia virus (VACV antigens driven by late promoters does not occur, as an abortive infection of pAPC prevents production of these late antigens. This lack of direct presentation results in a greatly diminished or ablated T(CD8+ response to late antigens. We demonstrate that late poxvirus antigens do not enter the cross-presentation pathway, even when identical antigens driven by early promoters access this pathway efficiently. The mechanism mediating this novel means of viral modulation of antigen presentation involves the sequestration of late antigens within virus factories. Early antigens and cellular antigens are cross-presented from virus-infected cells, as are late antigens that are targeted to compartments outside of the virus factories. This virus-mediated blockade specifically targets the cross-presentation pathway, since late antigen that is not cross-presented efficiently enters the MHC Class II presentation pathway. These data are the first to describe an evasion mechanism employed by pathogens to prevent entry into the cross-presentation pathway. In the absence of direct presentation, this evasion mechanism leads to a complete ablation of the T(CD8+ response and a potential replicative advantage for the virus. Such mechanisms of viral modulation of antigen presentation

  5. Carcinoma-associated antigens

    International Nuclear Information System (INIS)

    Bartorelli, A.; Accinni, R.

    1981-01-01

    This invention relates to novel antigens associated with breast carcinoma, anti-sera specific to said antigens, 125 I-labeled forms of said antigens and methods of detecting said antigens in serum or plasma. The invention also relates to a diagnostic kit containing standardised antigens or antisera or marked forms thereof for the detection of said antigens in human blood, serum or plasma. (author)

  6. Identification of Surface Exposed Elementary Body Antigens of ...

    African Journals Online (AJOL)

    This study sought to identify the surface exposed antigenic components of Cowdria ruminantium elementary body (EB) by biotin labeling, determine effect of reducing and non-reducing conditions and heat on the mobility of these antigens and their reactivity to antibodies from immunized animals by Western blotting.

  7. Immunity to tumour antigens.

    Science.gov (United States)

    Li, Geng; Ali, Selman A; McArdle, Stephanie E B; Mian, Shahid; Ahmad, Murrium; Miles, Amanda; Rees, Robert C

    2005-01-01

    During the last decade, a large number of human tumour antigens have been identified. These antigens are classified as tumour-specific shared antigens, tissue-specific differentiation antigens, overexpressed antigens, tumour antigens resulting from mutations, viral antigens and fusion proteins. Antigens recognised by effectors of immune system are potential targets for antigen-specific cancer immunotherapy. However, most tumour antigens are self-proteins and are generally of low immunogenicity and the immune response elicited towards these tumour antigens is not always effective. Strategies to induce and enhance the tumour antigen-specific response are needed. This review will summarise the approaches to discovery of tumour antigens, the current status of tumour antigens, and their potential application to cancer treatment.

  8. Effective antigen presentation to helper T cells by human eosinophils.

    Science.gov (United States)

    Farhan, Ruhaifah K; Vickers, Mark A; Ghaemmaghami, Amir M; Hall, Andrew M; Barker, Robert N; Walsh, Garry M

    2016-12-01

    Although eosinophils are inflammatory cells, there is increasing attention on their immunomodulatory roles. For example, murine eosinophils can present antigen to CD4 + T helper (Th) cells, but it remains unclear whether human eosinophils also have this ability. This study determined whether human eosinophils present a range of antigens, including allergens, to activate Th cells, and characterized their expression of MHC class II and co-stimulatory molecules required for effective presentation. Human peripheral blood eosinophils purified from non-allergic donors were pulsed with the antigens house dust mite extract (HDM), Timothy Grass extract (TG) or Mycobacterium tuberculosis purified protein derivative (PPD), before co-culture with autologous CD4 + Th cells. Proliferative and cytokine responses were measured, with eosinophil expression of HLA-DR/DP/DQ and the co-stimulatory molecules CD40, CD80 and CD86 determined by flow cytometry. Eosinophils pulsed with HDM, TG or PPD drove Th cell proliferation, with the response strength dependent on antigen concentration. The cytokine responses varied with donor and antigen, and were not biased towards any particular Th subset, often including combinations of pro- and anti-inflammatory cytokines. Eosinophils up-regulated surface expression of HLA-DR/DP/DQ, CD80, CD86 and CD40 in culture, increases that were sustained over 5 days when incubated with antigens, including HDM, or the major allergens it contains, Der p I or Der p II. Human eosinophils can, therefore, act as effective antigen-presenting cells to stimulate varied Th cell responses against a panel of antigens including HDM, TG or PPD, an ability that may help to determine the development of allergic disease. © 2016 John Wiley & Sons Ltd.

  9. Epithelium Expressing the E7 Oncoprotein of HPV16 Attracts Immune-Modulatory Dendritic Cells to the Skin and Suppresses Their Antigen-Processing Capacity.

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    Janin Chandra

    Full Text Available Antigen presenting cells (APCs in skin can promote either antigen-specific effector functions or antigen tolerance, and thus determine clearance or persistence of cutaneous viral infections. Human papillomavirus (HPV infections can persist in squamous epithelium in immunocompetent individuals, and some persisting HPV infections, particularly with HPV16, promote malignant epithelial transformation. Here, we investigate whether local expression of the HPV16 protein most associated with malignant transformation, HPV16-E7, affects the phenotype and function of APC subsets in the skin. We demonstrate an expanded population of Langerhans cells in HPV16-E7 transgenic skin with distinct cell surface markers which express immune-modulatory enzymes and cytokines not expressed by cells from non transgenic skin. Furthermore, HPV16-E7 transgene expression in keratinocytes attracts new APC subsets to the epidermis. In vivo migration and transport of antigen to the draining lymph node by these APCs is markedly enhanced in HPV16-E7 expressing skin, whereas antigen-processing, as measured by proteolytic cleavage of DQ-OVA and activation of T cells in vivo by APCs, is significantly impaired. These data suggest that local expression of HPV16-E7 in keratinocytes can contribute to persisting infection with this oncogenic virus, by altering the phenotype and function of local APCs.

  10. Can resting B cells present antigen to T cells

    International Nuclear Information System (INIS)

    Ashwell, J.D.; DeFranco, A.L.; Paul, W.E.; Schwartz, R.H.

    1985-01-01

    Antigen stimulation of T lymphocytes can occur only in the presence of an antigen-presenting cell (APC). An ever-increasing number of cell types have been found to act as APCs; these include macrophages, splenic and lymph node dendritic cells, and Langerhans cells of the skin. Although activated B lymphocytes and B cell lymphomas are known to serve as APCs, it has been generally believed that resting B cells cannot perform this function. However, in recent studies the authors have found that resting B cells can indeed present soluble antigen to T cell clones as well as to antigen-primed T cells. The previous difficulty in demonstrating this activity can be explained by the finding that, in contrast to macrophages and dendritic cells, the antigen-presenting ability of resting B cells is very radiosensitive. Macrophages are usually irradiated with 2000-3300 rads to prevent them from incorporating [ 3 H]thymidine in the T cell proliferation assay. Resting B cells, however, begin to lose presenting function at 1500 rads and have completely lost this activity at 3300 rads. It was also possible to distinguish two distinct T cell clonal phenotypes when resting B cells were used as APCs on the basis of two different assays (T cell proliferation, and B cell proliferation resulting from T cell activation). The majority of T cell clones tested were capable of both proliferating themselves and inducing the proliferation of B cells. Some T cells clones, however, could not proliferate in the presence of antigen and B cell APCs, although they were very good at inducing the proliferation of B cells

  11. Combined radioimmunoassay of HBs-antigen and anti-HBs using the Biotest combRIA-Au kit

    International Nuclear Information System (INIS)

    Kselikova, M.; Novak, J.; Urbankova, J.

    1979-01-01

    Hepatitis B antigen and antibody were determined simultaneously by radioimmunoassay using the Biotest combRIA-Au kit. The results in determining antigens and antibodies in sera with a known content of these substances by this kit nearly equal those of isolated determinations of antigen by the Abbott AUSRIA II-125 kit, and of antibody by the Abbott AUSAB kit, resp., except for a negligible number of sera with a very low content of either antigen or antibody. (author)

  12. Distribution of ABO and Rh-Hr blood group antigens, alleles and ...

    African Journals Online (AJOL)

    ABO and Rh-Hr blood group antigens represent a genetically stably determined trait with many-sided biological and clinical significance. The indigenous Ajarian population (105 subjects) was investigated for ABO Rh-Hr red cell blood group antigens. Using immunoserologic methods, seven blood group antigens (A, B, C, c, ...

  13. Somatic hypermutation of the new antigen receptor gene (NAR) in the nurse shark does not generate the repertoire: possible role in antigen-driven reactions in the absence of germinal centers.

    Science.gov (United States)

    Diaz, M; Greenberg, A S; Flajnik, M F

    1998-11-24

    The new antigen receptor (NAR) gene in the nurse shark diversifies extensively by somatic hypermutation. It is not known, however, whether NAR somatic hypermutation generates the primary repertoire (like in the sheep) or rather is used in antigen-driven immune responses. To address this issue, the sequences of NAR transmembrane (Tm) and secretory (Sec) forms, presumed to represent the primary and secondary repertoires, respectively, were examined from the peripheral blood lymphocytes of three adult nurse sharks. More than 40% of the Sec clones but fewer than 11% of Tm clones contained five mutations or more. Furthermore, more than 75% of the Tm clones had few or no mutations. Mutations in the Sec clones occurred mostly in the complementarity-determining regions (CDR) with a significant bias toward replacement substitutions in CDR1; in Tm clones there was no significant bias toward replacements and only a low level of targeting to the CDRs. Unlike the Tm clones where the replacement mutational pattern was similar to that seen for synonymous changes, Sec replacements displayed a distinct pattern of mutations. The types of mutations in NAR were similar to those found in mouse Ig genes rather than to the unusual pattern reported for shark and Xenopus Ig. Finally, an oligoclonal family of Sec clones revealed a striking trend toward acquisition of glutamic/aspartic acid, suggesting some degree of selection. These data strongly suggest that hypermutation of NAR does not generate the repertoire, but instead is involved in antigen-driven immune responses.

  14. A novel plasmid-encoded serotype conversion mechanism through addition of phosphoethanolamine to the O-antigen of Shigella flexneri.

    Directory of Open Access Journals (Sweden)

    Qiangzheng Sun

    Full Text Available Shigella flexneri is the major pathogen causing bacillary dysentery in developing countries. S. flexneri is divided into at least 16 serotypes based on the combination of antigenic determinants present in the O-antigen. All the serotypes (except for serotype 6 share a basic O-unit containing one N-acetyl-d-glucosamine and three l-rhamnose residues, whereas differences between the serotypes are conferred by phage-encoded glucosylation and/or O-acetylation. Serotype Xv is a newly emerged and the most prevalent serotype in China, which can agglutinate with both MASF IV-1 and 7,8 monoclonal antibodies. The factor responsible for the presence of MASF IV-1 (E1037 epitope has not yet been identified. In this study, we analyzed the LPS structure of serotype Xv strains and found that the MASF IV-1 positive phenotype depends on an O-antigen modification with a phosphoethanolamine (PEtN group attached at position 3 of one of the rhamnose residues. A plasmid carried gene, lpt-O (LPS phosphoethanolamine transferase for O-antigen, mediates the addition of PEtN for serotype Xv and other MASF IV-1 positive strains. These findings reveal a novel serotype conversion mechanism in S. flexneri and show the necessity of further extension of the serotype classification scheme recognizing the MASF IV-1 positive strains as distinctive subtypes.

  15. Determination of an unrelated donor pool size for human leukocyte antigen-matched platelets in Brazil

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    Carolina Bonet Bub

    2016-02-01

    Full Text Available ABSTRACT Background: Successful transfusion of platelet refractory patients is a challenge. Many potential donors are needed to sustain human leukocyte antigen matched-platelet transfusion programs because of the different types of antigens and the constant needs of these patients. For a highly mixed population such as the Brazilian population, the pool size required to provide adequate platelet support is unknown. Methods: A mathematical model was created to estimate the appropriate size of an unrelated donor pool to provide human leukocyte antigen-compatible platelet support for a Brazilian population. A group of 154 hematologic human leukocyte antigen-typed patients was used as the potential patient population and a database of 65,500 human leukocyte antigen-typed bone marrow registered donors was used as the donor population. Platelet compatibility was based on the grading system of Duquesnoy. Results: Using the mathematical model, a pool containing 31,940, 1710 and 321 donors would be necessary to match more than 80% of the patients with at least five completely compatible (no cross-reactive group, partial compatible (one cross-reactive group or less compatible (two cross-reactive group donors, respectively. Conclusion: The phenotypic diversity of the Brazilian population has probably made it more difficulty to find completely compatible donors. However, this heterogeneity seems to have facilitated finding donors when cross-reactive groups are accepted as proposed by the grading system of Duquesnoy. The results of this study may help to establish unrelated human leukocyte antigen-compatible platelet transfusions, a procedure not routinely performed in most Brazilian transfusion services.

  16. Screening Immunomodulators To Skew the Antigen-Specific Autoimmune Response.

    Science.gov (United States)

    Northrup, Laura; Sullivan, Bradley P; Hartwell, Brittany L; Garza, Aaron; Berkland, Cory

    2017-01-03

    Current therapies to treat autoimmune diseases often result in side effects such as nonspecific immunosuppression. Therapies that can induce antigen-specific immune tolerance provide an opportunity to reverse autoimmunity and mitigate the risks associated with global immunosuppression. In an effort to induce antigen-specific immune tolerance, co-administration of immunomodulators with autoantigens has been investigated in an effort to reprogram autoimmunity. To date, identifying immunomodulators that may skew the antigen-specific immune response has been ad hoc at best. To address this need, we utilized splenocytes obtained from mice with experimental autoimmune encephalomyelitis (EAE) in order to determine if certain immunomodulators may induce markers of immune tolerance following antigen rechallenge. Of the immunomodulatory compounds investigated, only dexamethasone modified the antigen-specific immune response by skewing the cytokine response and decreasing T-cell populations at a concentration corresponding to a relevant in vivo dose. Thus, antigen-educated EAE splenocytes provide an ex vivo screen for investigating compounds capable of skewing the antigen-specific immune response, and this approach could be extrapolated to antigen-educated cells from other diseases or human tissues.

  17. Development of an immunomagnetic bead-based time-resolved fluorescence immunoassay for rapid determination of levels of carcinoembryonic antigen in human serum

    Energy Technology Data Exchange (ETDEWEB)

    Hou Jingyuan; Liu Tiancai; Lin Guanfeng; Li Zhixiong; Zou Liping; Li Ming [Institute of Antibody Engineering, School of Biotechnology, Southern Medical University, Guangzhou 510515 (China); Wu Yingsong, E-mail: wg@fimmu.com [Institute of Antibody Engineering, School of Biotechnology, Southern Medical University, Guangzhou 510515 (China)

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer Magnetic beads was used as the solid phase for TRFIA. Black-Right-Pointing-Pointer The linearity range was broadened greatly compared with conventional TRFIA method. Black-Right-Pointing-Pointer The analysis time was significantly shorter compared with conventional TRFIA method. Black-Right-Pointing-Pointer This method could be developed for practical clinical detections of tumor-associated antigens. - Abstract: A novel immunoassay for the determination of tumor markers in human serum was established by combining a time-resolved fluoroimmunoassay (TRFIA) and immunomagnetic separation. Based on a sandwich-type immunoassay format, analytes in samples were captured by magnetic beads coated with one monoclonal antibody and 'sandwiched' by another monoclonal antibody labeled with europium chelates. The immunocomplex was separated and washed by exposure to a magnetic field and treatment with enhancement solution; fluorescence was then measured according to the number of europium ions dissociated. Levels of the model analyte, carcinoembryonic antigen (CEA), were determined in a linear range (1-1000 ng mL{sup -1}) with a limit of detection of 0.5 ng mL{sup -1} under optimal conditions. The reproducibility, recovery, and specificity of the immunoassay were demonstrated to be acceptable. To evaluate this novel assay for clinical applications, 239 serum samples were evaluated. Compared with the conventional TRFIA and chemiluminescence immunoassay (CLIA), the correlation coefficients of the developed immunoassay were 0.985 and 0.975, respectively. These results showed good correlation and confirmed that our method is feasible and could be used for the clinical determination of CEA (or other tumor antigens) in human serum.

  18. Advances in alfalfa mosaic virus-mediated expression of anthrax antigen in planta

    International Nuclear Information System (INIS)

    Brodzik, R.; Bandurska, K.; Deka, D.; Golovkin, M.; Koprowski, H.

    2005-01-01

    Plant viruses show great potential for production of pharmaceuticals in plants. Such viruses can harbor a small antigenic peptide(s) as a part of their coat proteins (CP) and elicit an antigen-specific immune response. Here, we report the high yield and consistency in production of recombinant alfalfa mosaic virus (AlMV) particles for specific presentation of the small loop 15 amino acid epitope from domain-4 of the Bacillus anthracis protective antigen (PA-D4s). The epitope was inserted immediately after the first 25 N-terminal amino acids of AlMV CP to retain genome activation and binding of CP to viral RNAs. Recombinant AlMV particles were efficiently produced in tobacco, easily purified for immunological analysis, and exhibited extended stability and systemic proliferation in planta. Intraperitional injections of mice with recombinant plant virus particles harboring the PA-D4s epitope elicited a distinct immune response. Western blotting and ELISA analysis showed that sera from immunized mice recognized both native PA antigen and the AlMV CP

  19. Monoclonal Antibody Production against Human Spermatozoal Surface Antigens

    Directory of Open Access Journals (Sweden)

    M Jedi-Tehrani

    2005-10-01

    Full Text Available Introduction: As monoclonal antibodies are potential tools for characterization of soluble or cellular surface antigens, use of these proteins has always been considered in infertility and reproduction research. Therefore, in this study, monoclonal antibodies against human sperm surface antigens were produced. Material and Methods: To produce specific clones against human sperm surface antigens, proteins were extracted using solubilization methods. Balb/c mice were immunized intraperitoneally with the proteins using complete Freund’s adjuvant in the first injection and incomplete Adjuvant in the following booster injections. Hybridoma cells producing ASA were cloned by limiting dilution. Results: Five stable ASA producing hybridoma clones were achieved and their antibody isotypes were determined by ELISA. All the isotypes were of IgG class. Their cross reactivity with rat and mice spermatozoa was examined but they did not have any cross reactivity. Conclusion: The produced antibodies can be used in further studies to characterize and evaluate each of the antigens present on human sperm surface and determining their role in fertilization.

  20. Radioimmunologic determination of the concentration of carcinoembryonic antigen in serum of normal individuals

    International Nuclear Information System (INIS)

    Milkov, V.; Milanov, S.

    1982-01-01

    The serum concentration of carcinoembryonic antigen (CEA) was determined by radioimmunoassay in 95 normal individuals (41 women and 54 men), 20 to 65 years of age. Depending on sex and age, the tested individuals were divided in four groups: gr. I - 27 women, 20 to 40 years of age; gr. II - 14 women, 4O to 65 years of age; gr. III -35 men, 20 to 40 years of age, and group IV - 19 men, 40 to 65 years of age. The following mean serum CEA levels were obtained in normal individuals: Group I -6.8 +- 1.07 ng/ml; group II - 9.71 +- 1.46 ng/ml; group III - 4.9 +- 0.73 ng/ml; group IV - 7.5 +- 1.5 ng/ml. The CEA levels in the serum of normal individuals varied with age and sex, but the differences were statistically insignificant (p> 0.10). Normal values fo serum CEA concentrations in normal individuals were determined. These values are meant to be used for comparison with serum CEA values in patients with malignant diseases. (author)

  1. THE LIVER OF WOODCHUCKS CHRONICALLY INFECTED WITH THE WOODCHUCK HEPATITIS VIRUS CONTAINS FOCI OF VIRUS CORE ANTIGEN NEGATIVE HEPATOCYTES WITH BOTH ALTERED AND NORMAL MORPHOLOGY

    Science.gov (United States)

    Xu, Chunxiao; Yamamoto, Toshiki; Zhou, Tianlun; Aldrich, Carol E.; Frank, Katy; Cullen, John M.; Jilbert, Allison R.; Mason, William S.

    2007-01-01

    The livers of woodchucks chronically infected with woodchuck hepatitis virus (WHV) contain foci of morphologically altered hepatocytes (FAH) with “basophilic”, “amphophilic” and “clear cell” phenotypes, which are possibly pre-neoplastic in nature. Interestingly, most fail to express detectable levels of WHV proteins and nucleic acids. We studied sections of WHV-infected liver tissue to determine if all foci of hepatocytes that failed to express detectable levels of WHV, as assessed by immunoperoxidase staining for WHV core antigen, could be classified morphologically as FAH. We found that at least half of the foci of WHV core antigen negative hepatocytes did not show clear morphological differences in either H&E or PAS (periodic acid Schiff) stained sections from surrounding hepatocytes, and were therefore not designated as FAH. In the second approach, we assayed core antigen negative foci for the presence of fetuin B, a serum protein produced by normal hepatocytes, but not by neoplastic hepatocytes in hepatocellular carcinomas. Basophilic and amphophilic FAH had reduced levels of fetuin B compared to hepatocytes present in the surrounding liver; fetuin B staining was detected in clear cell FAH but the level could not be accurately assessed because of the displacement of fetuin B to the cell periphery by accumulated glycogen. The foci of morphologically normal WHV core antigen negative hepatocytes had similar levels of fetuin B to that of the surrounding hepatocytes. The co-existence of at least four types of WHV core antigen negative foci, including those with no obvious morphologic changes, raises the possibility that the different foci arise from distinct primary events. We hypothesize that a common event is loss of the ability to express WHV, allowing these hepatocytes to escape immune mediated cell death and to undergo clonal expansion to form distinct foci. PMID:17078989

  2. Phase variable O antigen biosynthetic genes control expression of the major protective antigen and bacteriophage receptor in Vibrio cholerae O1.

    Directory of Open Access Journals (Sweden)

    Kimberley D Seed

    2012-09-01

    Full Text Available The Vibrio cholerae lipopolysaccharide O1 antigen is a major target of bacteriophages and the human immune system and is of critical importance for vaccine design. We used an O1-specific lytic bacteriophage as a tool to probe the capacity of V. cholerae to alter its O1 antigen and identified a novel mechanism by which this organism can modulate O antigen expression and exhibit intra-strain heterogeneity. We identified two phase variable genes required for O1 antigen biosynthesis, manA and wbeL. manA resides outside of the previously recognized O1 antigen biosynthetic locus, and encodes for a phosphomannose isomerase critical for the initial step in O1 antigen biosynthesis. We determined that manA and wbeL phase variants are attenuated for virulence, providing functional evidence to further support the critical role of the O1 antigen for infectivity. We provide the first report of phase variation modulating O1 antigen expression in V. cholerae, and show that the maintenance of these phase variable loci is an important means by which this facultative pathogen can generate the diverse subpopulations of cells needed for infecting the host intestinal tract and for escaping predation by an O1-specific phage.

  3. Radioimmunoassay for tumor antigen of human cervical squamous cell carcinoma

    International Nuclear Information System (INIS)

    Kato, H.; Torigoe, T.

    1977-01-01

    A heterologous antiserum for human cervical squamous cell carcinoma was prepared and specificity determined by Ouchterlony immunodiffusion and immunofluorescence studies. With this antiserum, a tumor antigen was purified from human cervical squamous cell carcinoma tissue. The specificities of the antigen and the antiserum were then re-examined by a radioimmunoassay method using 125 I-labeled purified antigen. Although normal cervical tissue extract showed a moderate cross-reactivity in the radioimmunoassay, the circulating antigen activity could not be detected in normal women or in several patients with other carcinomas, whereas 27 of 35 patients with cervical squamous cell carcinoma showed detectable serum antigen activity. All patients with advanced stages of cervical squamous cell carcinoma showed detectable antigen levels. These results indicate that there is a quantitative abnormality, at least, of this tumor antigen in patients with cervical squamous cell carcinoma and that the radioimmunoassay for the antigen is a potentially useful tool in clinical care

  4. Immunogenicity of 60 novel latency-related antigens of Mycobacterium tuberculosis.

    Science.gov (United States)

    Serra-Vidal, Mᵃdel Mar; Latorre, Irene; Franken, Kees L C M; Díaz, Jéssica; de Souza-Galvão, Maria Luiza; Casas, Irma; Maldonado, José; Milà, Cèlia; Solsona, Jordi; Jimenez-Fuentes, M Ángeles; Altet, Neus; Lacoma, Alícia; Ruiz-Manzano, Juan; Ausina, Vicente; Prat, Cristina; Ottenhoff, Tom H M; Domínguez, José

    2014-01-01

    The aim of our work here was to evaluate the immunogenicity of 60 mycobacterial antigens, some of which have not been previously assessed, notably a novel series of in vivo-expressed Mycobacterium tuberculosis (IVE-TB) antigens. We enrolled 505 subjects and separated them in individuals with and without latent tuberculosis infection (LTBI) vs. patients with active tuberculosis (TB). Following an overnight and 7 days stimulation of whole blood with purified recombinant M. tuberculosis antigens, interferon-γ (IFN-γ) levels were determined by ELISA. Several antigens could statistically significantly differentiate the groups of individuals. We obtained promising antigens from all studied antigen groups [dormancy survival regulon (DosR regulon) encoded antigens; resuscitation-promoting factors (Rpf) antigens; IVE-TB antigens; reactivation associated antigens]. Rv1733, which is a probable conserved transmembrane protein encoded in DosR regulon, turned out to be very immunogenic and able to discriminate between the three defined TB status, thus considered a candidate biomarker. Rv2389 and Rv2435n, belonging to Rpf family and IVE-TB group of antigens, respectively, also stood out as LTBI biomarkers. Although more studies are needed to support our findings, the combined use of these antigens would be an interesting approach to TB immunodiagnosis candidates.

  5. Immunogenicity of 60 novel latency-related antigens of Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Mªdel Mar eSerra Vidal

    2014-10-01

    Full Text Available The aim of our work here was to evaluate the immunogenicity of 60 mycobacterial antigens, some of which have not been previously assessed, notably a novel series of in vivo-expressed M.tuberculosis (IVE-TB antigens. We enrolled 505 subjects and separated them in individuals with and without latent tuberculosis infection (LTBI versus patients with active tuberculosis. Following an overnight and 7 day stimulation of whole blood with purified recombinant M.tb antigens, interferon-γ (IFN-γ levels were determined by ELISA. Several antigens could statistically significantly differentiate the groups of individuals. We obtained promising antigens from all studied antigen groups (DosR regulon encoded antigens; resuscitation-promoting factors (Rpf antigens; IVE-TB antigens; reactivation asociated antigens. Rv1733, which is a probable conserved transmembrane protein encoded in DosR regulon, turned out to be very immunogenic and able to discriminate between the three defined TB status, thus considered a candidate biomarker. Rv2389 and Rv2435n, belonging to Rpf family and IVE-TB group of antigens, respectively, also stood out as LTBI biomarkers. Although more studies are needed to support our findings, the combined use of these antigens would be an interesting approach to tuberculosis immunodiagnosis candidates.

  6. Evidence for glycosyl-phosphatidylinositol anchoring of Toxoplasma gondii major surface antigens

    International Nuclear Information System (INIS)

    Tomavo, S.; Schwarz, R.T.; Dubremetz, J.F.

    1989-01-01

    The four major surface antigens of Toxoplasma gondii tachyzoites (P43, P35, P30, and P22) were made water soluble by phosphatidylinositol-specific phospholipase C (PI-PLC). These antigens were biosynthetically labeled with 3 H-fatty acids, [ 3 H]ethanolamine, and [ 3 H]carbohydrates. Treatment of 3 H-fatty-acid-labeled parasite lysates with PI-PLC removed the radioactive label from these antigens. A cross-reacting determinant was exposed on these antigens after PI-PLC treatment

  7. Screening for epitope specificity directly on culture supernatants in the early phase of monoclonal antibody production by an ELISA with biotin-labeled antigen

    DEFF Research Database (Denmark)

    Andersen, Ditte C; Jensen, Charlotte H; Gregersen, Annemette

    2004-01-01

    This report describes an assay for comparison of epitope specificity in groups of monoclonal antibodies against a given antigen. The only prerequisite is the biotin-labeled antigen. One of the monoclonal antibodies is captured onto a plastic surface via a rabbit anti-mouse Ig, and the other...... preincubated with biotinylated antigen. When the two antibodies react with the same epitope subsequent binding of the biotin-labeled antigen is abolished (inhibition). In the cases where no inhibition was observed, the two antibodies were considered to react with distinct, independent epitopes. The obvious...

  8. Specificity of antigens on UV radiation-induced antigenic tumor cell variants measured in vitro and in vivo

    International Nuclear Information System (INIS)

    Hostetler, L.W.; Romerdahl, C.A.; Kripke, M.L.

    1989-01-01

    The purpose of this study was to determine whether antigenic variants cross-react immunologically with the parental tumor and whether the UVR-associated antigen unique to UVR-induced tumors is also present on the variants. Antigenic (regressor) variants and nonimmunogenic (progressor) clones derived from UV-irradiated cultures of the C3H K1735 melanoma and SF19 spontaneous fibrosarcoma cell lines were used to address these questions. In an in vivo immunization and challenge assay, the antigenic variants did not induce cross-protection among themselves, but each induced immunity against the immunizing variant, the parent tumor cells, and nonimmunogenic clones derived from UV-irradiated parent cultures. Therefore, the variants can be used to induce in mice a protective immunity that prevents the growth of the parent tumor and nonimmunogenic clones, but not other antigenic variants. In contrast, immunization with cells of the parental tumor or the nonimmunogenic clones induced no protective immunity against challenge with any of the cell lines. Utilizing the K1735 melanoma-derived cell lines in vitro, T-helper (Th) cells isolated from tumor-immunized mice were tested for cross-reactivity by their ability to collaborate with trinitrophenyl-primed B-cells in the presence of trinitrophenyl-conjugated tumor cells. Also, the cross-reactivity of cytotoxic T-lymphocytes from tumor-immunized mice was assessed by a 4-h 51Cr-release assay. Antigenic variants induced cytotoxic T-lymphocytes and Th activity that was higher than that induced by the parent tumor and nonimmunogenic clones from the UVR-exposed parent tumor and cross-reacted with the parental tumor cells and nonimmunogenic clones, but not with other antigenic variants

  9. Spleen-dependent regulation of antigenic variation in malaria parasites: Plasmodium knowlesi SICAvar expression profiles in splenic and asplenic hosts.

    Directory of Open Access Journals (Sweden)

    Stacey A Lapp

    Full Text Available Antigenic variation by malaria parasites was first described in Plasmodium knowlesi, which infects humans and macaque monkeys, and subsequently in P. falciparum, the most virulent human parasite. The schizont-infected cell agglutination (SICA variant proteins encoded by the SICAvar multigene family in P. knowlesi, and Erythrocyte Membrane Protein-1 (EMP-1 antigens encoded by the var multigene family in P. falciparum, are expressed at the surface of infected erythrocytes, are associated with virulence, and serve as determinants of naturally acquired immunity. A parental P. knowlesi clone, Pk1(A+, and a related progeny clone, Pk1(B+1+, derived by an in vivo induced variant antigen switch, were defined by the expression of distinct SICA variant protein doublets of 210/190 and 205/200 kDa, respectively. Passage of SICA[+] infected erythrocytes through splenectomized rhesus monkeys results in the SICA[-] phenotype, defined by the lack of surface expression and agglutination with variant specific antisera.We have investigated SICAvar RNA and protein expression in Pk1(A+, Pk1(B+1+, and SICA[-] parasites. The Pk1(A+ and Pk1(B+1+ parasites express different distinct SICAvar transcript and protein repertoires. By comparison, SICA[-] parasites are characterized by a vast reduction in SICAvar RNA expression, the lack of full-length SICAvar transcript signals on northern blots, and correspondingly, the absence of any SICA protein detected by mass spectrometry.SICA protein expression may be under transcriptional as well as post-transcriptional control, and we show for the first time that the spleen, an organ central to blood-stage immunity in malaria, exerts an influence on these processes. Furthermore, proteomics has enabled the first in-depth characterization of SICA[+] protein phenotypes and we show that the in vivo switch from Pk1(A+ to Pk1(B+1+ parasites resulted in a complete change in SICA profiles. These results emphasize the importance of studying

  10. Novel adenovirus encoded virus-like particles displaying the placental malaria associated VAR2CSA antigen

    DEFF Research Database (Denmark)

    Andersson, Anne-Marie C; dos Santos Marques Resende, Mafalda; Salanti, Ali

    2017-01-01

    The malaria parasite Plasmodium falciparum presents antigens on the infected erythrocyte surface that bind human receptors expressed on the vascular endothelium. The VAR2CSA mediated binding to a distinct chondroitin sulphate A (CSA) is a crucial step in the pathophysiology of placental malaria a...

  11. Age-Associated Decline in Thymic B Cell Expression of Aire and Aire-Dependent Self-Antigens

    Directory of Open Access Journals (Sweden)

    Sergio Cepeda

    2018-01-01

    Full Text Available Although autoimmune disorders are a significant source of morbidity and mortality in older individuals, the mechanisms governing age-associated increases in susceptibility remain incompletely understood. Central T cell tolerance is mediated through presentation of self-antigens by cells constituting the thymic microenvironment, including epithelial cells, dendritic cells, and B cells. Medullary thymic epithelial cells (mTECs and B cells express distinct cohorts of self-antigens, including tissue-restricted self-antigens (TRAs, such that developing T cells are tolerized to antigens from peripheral tissues. We find that expression of the TRA transcriptional regulator Aire, as well as Aire-dependent genes, declines with age in thymic B cells in mice and humans and that cell-intrinsic and cell-extrinsic mechanisms contribute to the diminished capacity of peripheral B cells to express Aire within the thymus. Our findings indicate that aging may diminish the ability of thymic B cells to tolerize T cells, revealing a potential mechanistic link between aging and autoimmunity.

  12. Dissecting antigen processing and presentation routes in dermal vaccination strategies

    NARCIS (Netherlands)

    Platteel, Anouk C M; Henri, Sandrine; Zaiss, Dietmar M; Sijts, Alice J A M

    2017-01-01

    The skin is an attractive site for vaccination due to its accessibility and presence of immune cells surveilling this barrier. However, knowledge of antigen processing and presentation upon dermal vaccination is sparse. In this study we determined antigen processing routes that lead to CD8(+) T cell

  13. Polyclonal antibodies for the detection of Trypanosoma cruzi circulating antigens.

    Directory of Open Access Journals (Sweden)

    Edith S Málaga-Machaca

    2017-11-01

    Full Text Available Detection of Trypanosoma cruzi antigens in clinical samples is considered an important diagnostic tool for Chagas disease. The production and use of polyclonal antibodies may contribute to an increase in the sensitivity of immunodiagnosis of Chagas disease.Polyclonal antibodies were raised in alpacas, rabbits, and hens immunized with trypomastigote excreted-secreted antigen, membrane proteins, trypomastigote lysate antigen and recombinant 1F8 to produce polyclonal antibodies. Western blot analysis was performed to determine specificity of the developed antibodies. An antigen capture ELISA of circulating antigens in serum, plasma and urine samples was developed using IgY polyclonal antibodies against T. cruzi membrane antigens (capture antibody and IgG from alpaca raised against TESA. A total of 33 serum, 23 plasma and 9 urine samples were analyzed using the developed test. Among serum samples, compared to serology, the antigen capture ELISA tested positive in 55% of samples. All plasma samples from serology positive subjects were positive in the antigen capture ELISA. All urine positive samples had corresponding plasma samples that were also positive when tested by the antigen capture ELISA.Polyclonal antibodies are useful for detection of circulating antigens in both the plasma and urine of infected individuals. Detection of antigens is direct evidence of the presence of the parasite, and could be a better surrogate of current infection status.

  14. Distinct uptake mechanisms but similar intracellular processing of two different toll-like receptor ligand-peptide conjugates in dendritic cells.

    Science.gov (United States)

    Khan, Selina; Bijker, Martijn S; Weterings, Jimmy J; Tanke, Hans J; Adema, Gosse J; van Hall, Thorbald; Drijfhout, Jan W; Melief, Cornelis J M; Overkleeft, Hermen S; van der Marel, Gijsbert A; Filippov, Dmitri V; van der Burg, Sjoerd H; Ossendorp, Ferry

    2007-07-20

    Covalent conjugation of Toll-like receptor ligands (TLR-L) to synthetic antigenic peptides strongly improves antigen presentation in vitro and T lymphocyte priming in vivo. These molecularly well defined TLR-L-peptide conjugates, constitute an attractive vaccination modality, sharing the peptide antigen and a defined adjuvant in one single molecule. We have analyzed the intracellular trafficking and processing of two TLR-L conjugates in dendritic cells (DCs). Long synthetic peptides containing an ovalbumin cytotoxic T-cell epitope were chemically conjugated to two different TLR-Ls the TLR2 ligand, Pam(3)CysSK(4) (Pam) or the TLR9 ligand CpG. Rapid and enhanced uptake of both types of TLR-L-conjugated peptide occurred in DCs. Moreover, TLR-L conjugation greatly enhanced antigen presentation, a process that was dependent on endosomal acidification, proteasomal cleavage, and TAP translocation. The uptake of the CpG approximately conjugate was independent of endosomally-expressed TLR9 as reported previously. Unexpectedly, we found that Pam approximately conjugated peptides were likewise internalized independently of the expression of cell surface-expressed TLR2. Further characterization of the uptake mechanisms revealed that TLR2-L employed a different uptake route than TLR9-L. Inhibition of clathrin- or caveolin-dependent endocytosis greatly reduced uptake and antigen presentation of the Pam-conjugate. In contrast, internalization and antigen presentation of CpG approximately conjugates was independent of clathrin-coated pits but partly dependent on caveolae formation. Importantly, in contrast to the TLR-independent uptake of the conjugates, TLR expression and downstream TLR signaling was required for dendritic cell maturation and for priming of naïve CD8(+) T-cells. Together, our data show that targeting to two distinct TLRs requires distinct uptake mechanism but follows similar trafficking and intracellular processing pathways leading to optimal antigen

  15. Dendritic cells and hepatocytes use distinct pathways to process protective antigen from plasmodium in vivo.

    Directory of Open Access Journals (Sweden)

    Ian A Cockburn

    2011-03-01

    Full Text Available Malaria-protective CD8+ T cells specific for the circumsporozoite (CS protein are primed by dendritic cells (DCs after sporozoite injection by infected mosquitoes. The primed cells then eliminate parasite liver stages after recognizing the CS epitopes presented by hepatocytes. To define the in vivo processing of CS by DCs and hepatocytes, we generated parasites carrying a mutant CS protein containing the H-2K(b epitope SIINFEKL, and evaluated the T cell response using transgenic and mutant mice. We determined that in both DCs and hepatocytes CS epitopes must reach the cytosol and use the TAP transporters to access the ER. Furthermore, we used endosomal mutant (3d and cytochrome c treated mice to address the role of cross-presentation in the priming and effector phases of the T cell response. We determined that in DCs, CS is cross-presented via endosomes while, conversely, in hepatocytes protein must be secreted directly into the cytosol. This suggests that the main targets of protective CD8+ T cells are parasite proteins exported to the hepatocyte cytosol. Surprisingly, however, secretion of the CS protein into hepatocytes was not dependent upon parasite-export (Pexel/VTS motifs in this protein. Together, these results indicate that the presentation of epitopes to CD8+ T cells follows distinct pathways in DCs when the immune response is induced and in hepatocytes during the effector phase.

  16. Despite disorganized synapse structure, Th2 cells maintain directional delivery of CD40L to antigen-presenting B cells.

    Science.gov (United States)

    Gardell, Jennifer L; Parker, David C

    2017-01-01

    Upon recognition of peptide displayed on MHC molecules, Th1 and Th2 cells form distinct immunological synapse structures. Th1 cells have a bull's eye synapse structure with TCR/ MHC-peptide interactions occurring central to a ring of adhesion molecules, while Th2 cells have a multifocal synapse with small clusters of TCR/MHC interactions throughout the area of T cell/antigen-presenting cell interaction. In this study, we investigated whether this structural difference in the immunological synapse affects delivery of T cell help. The immunological synapse is thought to ensure antigen-specific delivery of cytolytic granules and killing of target cells by NK cells and cytolytic T cells. In helper T cells, it has been proposed that the immunological synapse may direct delivery of other effector molecules including cytokines. CD40 ligand (CD40L) is a membrane-bound cytokine essential for antigen-specific T cell help for B cells in the antibody response. We incubated Th1 and Th2 cells overnight with a mixture of antigen-presenting and bystander B cells, and the delivery of CD40L to B cells and subsequent B cell responses were compared. Despite distinct immunological synapse structures, Th1 and Th2 cell do not differ in their ability to deliver CD40L and T cell help in an antigen-specific fashion, or in their susceptibility to inhibition of help by a blocking anti-CD40L antibody.

  17. Molecular mimics of the tumour antigen MUC1.

    Directory of Open Access Journals (Sweden)

    Tharappel C James

    Full Text Available A key requirement for the development of cancer immunotherapy is the identification of tumour-associated antigens that are differentially or exclusively expressed on the tumour and recognized by the host immune system. However, immune responses to such antigens are often muted or lacking due to the antigens being recognized as "self", and further complicated by the tumour environment and regulation of immune cells within. In an effort to circumvent the lack of immune responses to tumour antigens, we have devised a strategy to develop potential synthetic immunogens. The strategy, termed mirror image phage display, is based on the concept of molecular mimicry as demonstrated by the idiotype/anti-idiotype paradigm in the immune system. Here as 'proof of principle' we have selected molecular mimics of the well-characterised tumour associated antigen, the human mucin1 protein (MUC1 from two different peptide phage display libraries. The putative mimics were compared in structure and function to that of the native antigen. Our results demonstrate that several of the mimic peptides display T-cell stimulation activity in vitro when presented by matured dendritic cells. The mimic peptides and the native MUC1 antigenic epitopes can cross-stimulate T-cells. The data also indicate that sequence homology and/or chemical properties to the original epitope are not the sole determining factors for the observed immunostimulatory activity of the mimic peptides.

  18. Unusual monosaccharides: components of O-antigenic polysaccharides of microorganisms

    Science.gov (United States)

    Kochetkov, Nikolai K.

    1996-09-01

    The data on new monosaccharides detected in O-antigenic polysaccharides of Gram-negative bacteria have been surveyed. The results of isolation and structure determination of these unusual monosaccharides have been arranged and described systematically. The NMR spectroscopy techniques are shown to be promising for the O-antigenic polysaccharides structure determination. The information about fine structure of monosaccharides which constitute the base of important class of microbial polysaccharides, is of great significance for applied studies, first of all, the design and synthesis of biologically active substances. The bibliography includes 216 references.

  19. Comparison of serum prostate specific antigen levels and bone scintigraphy in patients with prostate carcinoma

    International Nuclear Information System (INIS)

    Bielickaite, J.; Zadeikaite, R.; Jurkiene, N. and others

    2003-01-01

    The aim of this study was to analyze the levels of serum prostate specific antigen in patients with and without bone metastases detected by means of bone scintigraphy and to determine the highest prostate specific antigen level in patients without bone metastases. The 50 patients consecutively diagnosed of prostate cancer between 1999 and 2001 in our institution made up the study population. Prostate specific antigen plasmatic levels were determined and bone scintigraphy was performed (whole body study after 99mTc-methyl-diphosphonate administration) in all the patients. In patients with positive bone scans (n=23), the mean prostate specific antigen level was 71.4±35.2 ng/ml and was significantly (p<0.00005) higher than in 14 patients with negative bone scans (mean prostate specific antigen level was 10.1±10.5 ng/ml). Suspicious lesions were found in 13 patients and their mean prostate specific antigen level was 8.5±7.7 ng/ml. Regarding prostate specific antigen levels, no statistically significant differences were found between patients with suspicious lessons and normal bone scans. The highest determined prostate specific antigen level in patients without bone metastases was 18 ng/ml. The bone scintigraphy should be performed in all patients with prostate specific antigen level above 18 ng/ml, but it is of limited value in patients with prostate specific antigen level below 18 ng/ml. (author)

  20. The prevalence of hepatitis B virus E antigen among Ghanaian ...

    African Journals Online (AJOL)

    We studied the prevalence of hepatitis B virus 'e' antigen (HBeAg) among individuals determined to be hepatitis B virus (HBV) surface antigen- positive and analyzed the gender/age category associated with more active HBV infection and whether alteration in the levels of alanine aminotransferase could be associated with ...

  1. Commensal oral bacteria antigens prime human dendritic cells to induce Th1, Th2 or Treg differentiation.

    Science.gov (United States)

    Kopitar, A N; Ihan Hren, N; Ihan, A

    2006-02-01

    In various immunopathologic conditions, bacterial flora induce an immune response which results in inflammatory manifestations, e.g. periapical granuloma. Dendritic cells provide the main orchestration of specific immune responses. The aim of our study was to test the capacity of distinct oral bacterial antigens (prepared from Streptococcus mitis, Propionibacterium acnes, and Bacteroides spp.) to prime human dendritic cells for stimulation of the T-lymphocyte response. To assess the T-lymphocyte response, the expression of CD25, CD69, intracellular interferon gamma (cIFN-gamma), and intracellular interleukin 4 (cIL-4) was determined. Dendritic cells were prepared from leukocyte buffy coat from healthy blood donors. Monocytes were stimulated with IL-4 and GM-CSF and dendritic cells activated with bacterial lysates. Cell suspensions contained up to 90% dendritic cells, which represented 2-12% of the initial number of mononuclear cells. Lymphocyte subsets that developed in lymphocyte cultures after 1 week of stimulation were analyzed by flow cytometry. Dendritic cells, primed with antigens of Bacteroides fragilis have shown significantly higher activation and expression of intercellular IFN-gamma by T lymphocytes compared to negative controls. The dendritic cells primed with antigens of P. acnes had no effect on T-lymphocyte activation or cytokine production; instead they induced differentiation of T lymphocytes into CD25bright cells (regulatory T cells) with a potentially inhibitory effect on immune response. Dendritic cells primed with antigens of S. mitis induced increased expression of cIL-4. We conclude that commensal oral bacteria antigens prepared from B. fragilis, S. mitis, and P. acnes prime human dendritic cells to induce Th1, Th2, and T(reg) differentiation, respectively. This may advance our understanding of immunopathologic manifestations in the oral cavity and offer new possibilities for redirecting immune responses in mucosal vaccination.

  2. Using molecular principal axes for structural comparison: determining the tertiary changes of a FAB antibody domain induced by antigenic binding

    Directory of Open Access Journals (Sweden)

    Silverman B David

    2007-11-01

    Full Text Available Abstract Background Comparison of different protein x-ray structures has previously been made in a number of different ways; for example, by visual examination, by differences in the locations of secondary structures, by explicit superposition of structural elements, e.g. α-carbon atom locations, or by procedures that utilize a common symmetry element or geometrical feature of the structures to be compared. Results A new approach is applied to determine the structural changes that an antibody protein domain experiences upon its interaction with an antigenic target. These changes are determined with the use of two different, however comparable, sets of principal axes that are obtained by diagonalizing the second-order tensors that yield the moments-of-geometry as well as an ellipsoidal characterization of domain shape, prior to and after interaction. Determination of these sets of axes for structural comparison requires no internal symmetry features of the domains, depending solely upon their representation in three-dimensional space. This representation may involve atomic, Cα, or residue centroid coordinates. The present analysis utilizes residue centroids. When the structural changes are minimal, the principal axes of the domains, prior to and after interaction, are essentially comparable and consequently may be used for structural comparison. When the differences of the axes cannot be neglected, but are nevertheless slight, a smaller relatively invariant substructure of the domains may be utilized for comparison. The procedure yields two distance metrics for structural comparison. First, the displacements of the residue centroids due to antigenic binding, referenced to the ellipsoidal principal axes, are noted. Second, changes in the ellipsoidal distances with respect to the non-interacting structure provide a direct measure of the spatial displacements of the residue centroids, towards either the interior or exterior of the domain

  3. Antigen presentation by hapten-specific B lymphocytes. II. Specificity and properties of antigen-presenting B lymphocytes, and function of immunoglobulin receptors

    International Nuclear Information System (INIS)

    Abbas, A.K.; Haber, S.; Rock, K.L.

    1985-01-01

    Studies were designed to examine the ability of hapten-binding murine B lymphocytes to present hapten-protein conjugates to protein antigen-specific, Ia-restricted T cell hybridomas. BALB/c B cells specific for TNP or FITC presented hapten-modified proteins (TNP-G1 phi, TNP-OVA, or FITC-OVA) to the relevant T cell hybridomas at concentrations below 0.1 microgram/ml. Effective presentation of the same antigens by B lymphocyte-depleted splenocytes, and of unmodified proteins by either hapten-binding B cells or Ig spleen cells, required about 10(3)-to 10(4)-fold higher concentrations of antigen. The use of two different haptens and two carrier proteins showed that this extremely efficient presentation of antigen was highly specific, with hapten specificity being a property of the B cells and carrier specificity of the responding T cells. The presentation of hapten-proteins by hapten-binding B lymphocytes was radiosensitive and was not affected by the depletion of plastic-adherent cells, suggesting that conventional APCs (macrophages or dendritic cells) are not required in this phenomenon. Antigen-pulsing and antibody-blocking experiments showed that this hapten-specific antigen presentation required initial binding of antigen to surface Ig receptors. Moreover, linked recognition of hapten and carrier determinants was required, but these recognition events could be temporally separated. Finally, an antigen-processing step was found to be necessary, and this step was disrupted by ionizing radiation. These data suggest a role for B cell surface Ig in providing a specific high-affinity receptor to allow efficient uptake or focusing of antigen for its subsequent processing and presentation to T lymphocytes

  4. Antigenic typing Polish isolates of canine parvovirus

    Energy Technology Data Exchange (ETDEWEB)

    Mizak, B. [National Veterinary Research Institute, Pulawy (Poland); Plucienniczak, A. [Polish Academy ofd Sciences. Microbiology and Virology Center, Lodz (Poland)

    1995-12-31

    Polish strains of canine parvovirus isolated between 1982 and 1993 were examined to determine the extent to which the virus has evolved antigenically and genetically over eleven years. Two CPV isolates obtained in Warsaw in 1982 and Pulawy in 1993, were examined using monoclonal antibody typing, restriction analysis and sequencing VP-2 protein gene. Five other isolates from Warsaw and Pulawy were tested with the panel of monoclonal antibodies specific to CPV-2, CPV-2a and common for canine parvovirus, feline panleukopenia virus and milk enteritis virus. Results of the studies demonstrated that all isolates tested represented CPV-2a antigenic type. Rapid antigenic strain replacement recorded by Parrish and Senda in the U.S.A and Japan was not confirmed in Poland. (author). 30 refs, 2 tabs.

  5. Antigenic typing Polish isolates of canine parvovirus

    International Nuclear Information System (INIS)

    Mizak, B.; Plucienniczak, A.

    1995-01-01

    Polish strains of canine parvovirus isolated between 1982 and 1993 were examined to determine the extent to which the virus has evolved antigenically and genetically over eleven years. Two CPV isolates obtained in Warsaw in 1982 and Pulawy in 1993, were examined using monoclonal antibody typing, restriction analysis and sequencing VP-2 protein gene. Five other isolates from Warsaw and Pulawy were tested with the panel of monoclonal antibodies specific to CPV-2, CPV-2a and common for canine parvovirus, feline panleukopenia virus and milk enteritis virus. Results of the studies demonstrated that all isolates tested represented CPV-2a antigenic type. Rapid antigenic strain replacement recorded by Parrish and Senda in the U.S.A and Japan was not confirmed in Poland. (author). 30 refs, 2 tabs

  6. Abnormal antigens in breast cancer tissues and production of monoclonal antibodies against one of these antigens

    Energy Technology Data Exchange (ETDEWEB)

    Mohammed, M E. A. [University of Khartoum, Khartoum (Sudan)

    2010-02-15

    approximately 30 KDa molecular weight. However, some tests were done to determine the specificity of the produced monoclonal antibody. In conclusion, breast cancer tissues are associated with abnormal antigens, but with low incidence rate, this strongly favors the individualization of breast cancer diagnosis and treatment. (Author)

  7. Abnormal antigens in breast cancer tissues and production of monoclonal antibodies against one of these antigens

    International Nuclear Information System (INIS)

    Mohammed, M. E. A.

    2010-02-01

    approximately 30 KDa molecular weight. However, some tests were done to determine the specificity of the produced monoclonal antibody. In conclusion, breast cancer tissues are associated with abnormal antigens, but with low incidence rate, this strongly favors the individualization of breast cancer diagnosis and treatment. (Author)

  8. Use of synthetic, crystalline, L-α-dimyristoyl lecithin in cardiolipin antigens

    Science.gov (United States)

    Reyn, Alice; Bentzon, Michael Weis

    1956-01-01

    Experiments were carried out by the authors to determine whether synthetic, crystalline, L-α-dimyristoyl lecithin could replace natural purified lecithins in the preparation of cardiolipin antigens. These experiments were designed specifically to find out whether it was possible to obtain the same serological reactions, qualitatively and quantitatively, with the test antigen as with a reference antigen containing natural lecithin, and whether the test antigen had the same keeping qualities as the reference antigen. The tests used were the quantitative complement-fixation test as modified by Mørch in 1933, and the VDRL slide flocculation test. The results showed that synthetic, crystalline, L-α-dimyristoyl lecithin could replace natural lecithin in the preparation of cardiolipin antigens, but that the antigens prepared with the synthetic lecithin were significantly less sensitive than those prepared with an equimolar amount of natural lecithin. The authors consider that further investigation is required before the use of synthetic lecithin is finally adopted. PMID:13342931

  9. Histoplasma Urinary Antigen Testing Obviates the Need for Coincident Serum Antigen Testing.

    Science.gov (United States)

    Libert, Diane; Procop, Gary W; Ansari, Mohammad Q

    2018-03-07

    Serum and urine antigen (SAg, UAg) detection are common tests for Histoplasma capsulatum. UAg detection is more widely used and reportedly has a higher sensitivity. We investigated whether SAg detection contributes meaningfully to the initial evaluation of patients with suspected histoplasmosis. We reviewed 20,285 UAg and 1,426 SAg tests ordered from 1997 to 2016 and analyzed paired UAg and SAg tests completed on the same patient within 1 week. We determined the positivity rate for each test. Of 601 paired specimens, 542 were concurrent negatives and 48 were concurrent positives (98% agreement). Medical records were available for eight of 11 pairs with discrepant results. UAg was falsely positive in six instances, truly positive once, and falsely negative once. These findings support using a single antigen detection test, rather than both UAg and SAg, as an initial screen for suspected histoplasmosis. This aligns with the current practice of most physicians.

  10. Levels of estrogen, carcinoembryonic antigen and cancer antigen of breast in breast cancer patients

    International Nuclear Information System (INIS)

    Abdelhadi, H. A.

    2005-09-01

    This study was conducted during the period from february 2004 to July 2004; with the objective of measuring the levels of estrogen (E2), carcinoembryonic antigen (CEA) and cancer antigen of breast (CA-15.3) so as to facilitate the early diagnosis of breast cancer and determine the involvement of these parameters as risk factors for breast cancer. Ninety blood samples were collected from Sudanese females, divided into two groups; control group and patient groups. The patients group was sixty Sudanese females visiting the Radio Isotope Center, Khartoum (RICK) and they were confirmed as breast cancer patient by histopathology. The levels of the above mentioned parameters were determined by using radioimmunoassay technique. The results showed that, no significant (p=0.05) difference between the levels of the estrogen in patients compared to the control, on the other hand there was non significant (p>0.05) elevation in CEA levels in the patients with breast cancer compared to the control. The level of CA15.3 was significantly (p<0.0001) higher in the breast cancer patients compared to the control.(Author)

  11. Lea blood group antigen on human platelets

    International Nuclear Information System (INIS)

    Dunstan, R.A.; Simpson, M.B.; Rosse, W.F.

    1985-01-01

    One- and two-stage radioligand assays were used to determine if human platelets possess the Lea antigen. Goat IgG anti-Lea antibody was purified by multiple adsorptions with Le(a-b-) human red blood cells, followed by affinity chromatography with synthetic Lea substance and labeling with 125 I. Human IgG anti-Lea antibody was used either in a two stage radioassay with 125 I-labeled mouse monoclonal IgG anti-human IgG as the second antibody or, alternatively, purified by Staph protein A chromatography, labeled with 125 I, and used in a one-stage radioassay. Platelets from donors of appropriate red blood cell phenotypes were incubated with the antisera, centrifuged through phthalate esters, and assayed in a gamma scintillation counter. Dose response and saturation curve analysis demonstrate the presence of Lewis a antigen on platelets from Lea+ donors. Furthermore, platelets from an Le(a-b-) donor incubated in Le (a+b-) plasma adsorb Lea antigen in a similar manner to red blood cells. The clinical significance of these antigens in platelet transfusion remains undefined

  12. Insights into the biological features of the antigenic determinants recognized by four monoclonal antibodies in redia and adult stages of the liver fluke Fasciola hepatica.

    Science.gov (United States)

    Alba, Annia; Sánchez, Jorge; Hernández, Hilda; Mosqueda, Maryani; Rodríguez, Suanel Y; Capó, Virginia; Otero, Oscar; Alfonso, Carlos; Marcet, Ricardo; Sarracent, Jorge

    2016-09-01

    Fasciola hepatica is a digenean trematode which infects a wide variety of domestic animals and also humans. Previous studies have demonstrated that four monoclonal antibodies (Mabs) against the total extract of F. hepatica redia (named as 1E4, 6G11, 4E5 and 4G11) also recognized the excretion - secretion antigens (ES Ag) of adult parasites, which is a biologically-relevant mixture of molecules with functional roles during infection and immune evasion on definitive hosts. In the present report we describe the partial characterization of the epitopes recognized by these Mabs by heat treatment, mercaptoethanol reduction, pronase proteolysis and sodium peryodate oxidation, which suggested their predominant protein and conformational nature. Also, a comparative study using immunodetection assays on crude extracts and on histological sections of both rediae and adults of F. hepatica were performed to explore the expression pattern of the antigenic determinants in these developmental stages. From these experiments it was found that the Mabs reacted most likely with the same proteins of approximately 64 and 105 kDa present on both rediae and adult's extracts. However, the 1E4, 6G11 and 4E5 Mabs also recognized other molecules of the total extract of F. hepatica adults, a fact that constitutes an evidence of the antigenic variation between both stages and points at a certain biological relevance of the recognized antigenic determinants. Immunolocalization studies on histological sections revealed that all Mabs reacted with the tegument of F. hepatica in both rediae and adults stages, while the epitopes recognized by 1E4, 6G11 and 4E5 antibodies were also preferentially localized in the intestinal caeca and in different organs of the reproductive system of adult specimens. The immunogenicity of these antigenic determinants, their conserved status among different stages of the life cycle of F. hepatica and their presence in both tegument and ES Ag of adult parasites

  13. Development of a double-antibody radioimmunoassay for detecting ovarian tumor-associated antigen fraction OCA in plasma

    International Nuclear Information System (INIS)

    Knauf, S.; Urbach, G.I.

    1978-01-01

    Ovarian tumor-associated antigen isolated from human tumor tissue was shown to have a different mobility from that of carcinoembryonic antigen (CEA) in both acrylamide gel electrophoresis and immunoelectrophoresis in agarose. The ovarian tumor antigen is composed of six species with different electrophoretic mobility in acrylamide gel electrophoresis. Three of these species were detected in Sephadex G-100 ovarian fraction OCA (from the void volume peak) and the other three species of lower apparent molecular weight were detected in fraction OCD (from the second peak). Fractions OCA and OCD did not share common antigenic determinants as determined by immunodiffusion. CEA was shown to share antigenic determinants with both OCA and OCD. A double-antibody radioimmunoassay capable of detecting nanogram quantities of plasma OCA was developed. In a preliminary study of ovarian cancer patients, OCA appeared to be a more sensitive marker for ovarian cancer than CEA. There was virtually no correlation (r 2 = 0.1) between OCA and CEA levels in these patients, as determined by radioimmunoassay

  14. Role of the Antigen Capture Pathway in the Induction of a Neutralizing Antibody Response to Anthrax Protective Antigen

    Directory of Open Access Journals (Sweden)

    Anita Verma

    2018-02-01

    Full Text Available Toxin neutralizing antibodies represent the major mode of protective immunity against a number of toxin-mediated bacterial diseases, including anthrax; however, the cellular mechanisms that lead to optimal neutralizing antibody responses remain ill defined. Here we show that the cellular binding pathway of anthrax protective antigen (PA, the binding component of anthrax toxin, determines the toxin neutralizing antibody response to this antigen. PA, which binds cellular receptors and efficiently enters antigen-presenting cells by receptor-mediated endocytosis, was found to elicit robust anti-PA IgG and toxin neutralizing antibody responses. In contrast, a receptor binding-deficient mutant of PA, which does not bind receptors and only inefficiently enters antigen-presenting cells by macropinocytosis, elicited very poor antibody responses. A chimeric protein consisting of the receptor binding-deficient PA mutant tethered to the binding subunit of cholera toxin, which efficiently enters cells using the cholera toxin receptor rather than the PA receptor, elicited an anti-PA IgG antibody response similar to that elicited by wild-type PA; however, the chimeric protein elicited a poor toxin neutralizing antibody response. Taken together, our results demonstrate that the antigen capture pathway can dictate the magnitudes of the total IgG and toxin neutralizing antibody responses to PA as well as the ratio of the two responses.

  15. Radioimmunoassay to determine the cardioembryonic and carbohydrate antigens in the diagnosis of rectal cancer recurrences and metastases

    International Nuclear Information System (INIS)

    Ozhiganov, E.L.; Kuznetsova, L.F.

    1986-01-01

    A study was made of the results of measuring the carcinoembryonic and carbohydrate antigens using a kit of reagents in 75 patients with rectal cancer recurrences and metastases. The concentration of these antigens in healthy persons was for CEA 6.4±0.71 μg/l, the carbohydrate antigen - 19.6±2.51 units/ml. In this group of patients rectal cancer local recurrence was found in 52, metastases to the liver in 19 and metastatic involvement of the liver and lungs in 4. An elevated level of the CEA was detected in 92.8% of the patients with cancer recurrence (the mean concentration was 99.9±9.29 μg/l), and in 100% of the patients with metastases (the mean concentration was 193.4±30.42 μg/l). The content of the carbohydrate antigen in cancer recurrences was raised in 21.3% of the cases only, in metastases to the liver in 31.6% and in 2 patients with metastatic liver and lung involvement. Thus, measuring the CEA content turned out to be the most specific and sensitive test for the diagnosis of rectal cancer recurrences and metastases. The use of the carbohydrate antigen for this purpose was found ineffective

  16. Antigen sensitivity is a major determinant of CD8+ T-cell polyfunctionality and HIV-suppressive activity.

    Science.gov (United States)

    Almeida, Jorge R; Sauce, Delphine; Price, David A; Papagno, Laura; Shin, So Youn; Moris, Arnaud; Larsen, Martin; Pancino, Gianfranco; Douek, Daniel C; Autran, Brigitte; Sáez-Cirión, Asier; Appay, Victor

    2009-06-18

    CD8(+) T cells are major players in the immune response against HIV. However, recent failures in the development of T cell-based vaccines against HIV-1 have emphasized the need to reassess our basic knowledge of T cell-mediated efficacy. CD8(+) T cells from HIV-1-infected patients with slow disease progression exhibit potent polyfunctionality and HIV-suppressive activity, yet the factors that unify these properties are incompletely understood. We performed a detailed study of the interplay between T-cell functional attributes using a bank of HIV-specific CD8(+) T-cell clones isolated in vitro; this approach enabled us to overcome inherent difficulties related to the in vivo heterogeneity of T-cell populations and address the underlying determinants that synthesize the qualities required for antiviral efficacy. Conclusions were supported by ex vivo analysis of HIV-specific CD8(+) T cells from infected donors. We report that attributes of CD8(+) T-cell efficacy against HIV are linked at the level of antigen sensitivity. Highly sensitive CD8(+) T cells display polyfunctional profiles and potent HIV-suppressive activity. These data provide new insights into the mechanisms underlying CD8(+) T-cell efficacy against HIV, and indicate that vaccine strategies should focus on the induction of HIV-specific T cells with high levels of antigen sensitivity to elicit potent antiviral efficacy.

  17. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

    Directory of Open Access Journals (Sweden)

    Isabel Correa

    2018-03-01

    Full Text Available Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1 specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.

  18. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells.

    Science.gov (United States)

    Correa, Isabel; Ilieva, Kristina M; Crescioli, Silvia; Lombardi, Sara; Figini, Mariangela; Cheung, Anthony; Spicer, James F; Tutt, Andrew N J; Nestle, Frank O; Karagiannis, Panagiotis; Lacy, Katie E; Karagiannis, Sophia N

    2018-01-01

    Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires.

  19. Evaluation of Antigen-Conjugated Fluorescent Beads to Identify Antigen-Specific B Cells

    Science.gov (United States)

    Correa, Isabel; Ilieva, Kristina M.; Crescioli, Silvia; Lombardi, Sara; Figini, Mariangela; Cheung, Anthony; Spicer, James F.; Tutt, Andrew N. J.; Nestle, Frank O.; Karagiannis, Panagiotis; Lacy, Katie E.; Karagiannis, Sophia N.

    2018-01-01

    Selection of single antigen-specific B cells to identify their expressed antibodies is of considerable interest for evaluating human immune responses. Here, we present a method to identify single antibody-expressing cells using antigen-conjugated fluorescent beads. To establish this, we selected Folate Receptor alpha (FRα) as a model antigen and a mouse B cell line, expressing both the soluble and the membrane-bound forms of a human/mouse chimeric antibody (MOv18 IgG1) specific for FRα, as test antibody-expressing cells. Beads were conjugated to FRα using streptavidin/avidin-biotin bridges and used to select single cells expressing the membrane-bound form of anti-FRα. Bead-bound cells were single cell-sorted and processed for single cell RNA retrotranscription and PCR to isolate antibody heavy and light chain variable regions. Variable regions were then cloned and expressed as human IgG1/k antibodies. Like the original clone, engineered antibodies from single cells recognized native FRα. To evaluate whether antigen-coated beads could identify specific antibody-expressing cells in mixed immune cell populations, human peripheral blood mononuclear cells (PBMCs) were spiked with test antibody-expressing cells. Antigen-specific cells could comprise up to 75% of cells selected with antigen-conjugated beads when the frequency of the antigen-positive cells was 1:100 or higher. In PBMC pools, beads conjugated to recombinant antigens FRα and HER2 bound antigen-specific anti-FRα MOv18 and anti-HER2 Trastuzumab antibody-expressing cells, respectively. From melanoma patient-derived B cells selected with melanoma cell line-derived protein-coated fluorescent beads, we generated a monoclonal antibody that recognized melanoma antigen-coated beads. This approach may be further developed to facilitate analysis of B cells and their antibody profiles at the single cell level and to help unravel humoral immune repertoires. PMID:29628923

  20. Genetic diversity of K-antigen gene clusters of Escherichia coli and their molecular typing using a suspension array.

    Science.gov (United States)

    Yang, Shuang; Xi, Daoyi; Jing, Fuyi; Kong, Deju; Wu, Junli; Feng, Lu; Cao, Boyang; Wang, Lei

    2018-04-01

    Capsular polysaccharides (CPSs), or K-antigens, are the major surface antigens of Escherichia coli. More than 80 serologically unique K-antigens are classified into 4 groups (Groups 1-4) of capsules. Groups 1 and 4 contain the Wzy-dependent polymerization pathway and the gene clusters are in the order galF to gnd; Groups 2 and 3 contain the ABC-transporter-dependent pathway and the gene clusters consist of 3 regions, regions 1, 2 and 3. Little is known about the variations among the gene clusters. In this study, 9 serotypes of K-antigen gene clusters (K2ab, K11, K20, K24, K38, K84, K92, K96, and K102) were sequenced and correlated with their CPS chemical structures. On the basis of sequence data, a K-antigen-specific suspension array that detects 10 distinct CPSs, including the above 9 CPSs plus K30, was developed. This is the first report to catalog the genetic features of E. coli K-antigen variations and to develop a suspension array for their molecular typing. The method has a number of advantages over traditional bacteriophage and serum agglutination methods and lays the foundation for straightforward identification and detection of additional K-antigens in the future.

  1. A Distinct Population of Microglia Supports Adult Neurogenesis in the Subventricular Zone

    DEFF Research Database (Denmark)

    Ribeiro Xavier, Anna L.; Kress, Benjamin T.; Goldman, Steven A.

    2015-01-01

    found that microglia residing in the SVZ and adjacent rostral migratory stream (RMS) comprise a morphologically and antigenically distinct phenotype of immune effectors. Whereas exhibiting characteristics of alternatively activated microglia, the SVZ/RMS microglia were clearly distinguished by their low...... STATEMENT: Microglial cells are a specialized population of macrophages in the CNS, playing key roles as immune mediators. As integral components in the CNS, the microglia stand out for using the same mechanisms, phagocytosis and cytochemokine release, to promote homeostasis, synaptic pruning, and neural...... toward olfactory bulb layers. In addition to other unique populations residing in the SVZ niche, microglia display distinct morphofunctional properties that boost neuronal progenitor survival and migration in the mammalian brain....

  2. Antigen processing and remodeling of the endosomal pathway: requirements for antigen cross-presentation.

    Science.gov (United States)

    Compeer, Ewoud Bernardus; Flinsenberg, Thijs Willem Hendrik; van der Grein, Susanna Geertje; Boes, Marianne

    2012-01-01

    Cross-presentation of endocytosed antigen as peptide/class I major histocompatibility complex complexes plays a central role in the elicitation of CD8(+) T cell clones that mediate anti-viral and anti-tumor immune responses. While it has been clear that there are specific subsets of professional antigen presenting cells capable of antigen cross-presentation, identification of mechanisms involved is still ongoing. Especially amongst dendritic cells (DC), there are specialized subsets that are highly proficient at antigen cross-presentation. We here present a focused survey on the cell biological processes in the endosomal pathway that support antigen cross-presentation. This review highlights DC-intrinsic mechanisms that facilitate the cross-presentation of endocytosed antigen, including receptor-mediated uptake, maturation-induced endosomal sorting of membrane proteins, dynamic remodeling of endosomal structures and cell surface-directed endosomal trafficking. We will conclude with the description of pathogen-induced deviation of endosomal processing, and discuss how immune evasion strategies pertaining endosomal trafficking may preclude antigen cross-presentation.

  3. Antigen processing and remodeling of the endosomal pathway: requirements for antigen cross-presentation.

    Directory of Open Access Journals (Sweden)

    Ewoud Bernardus Compeer

    2012-03-01

    Full Text Available The cross-presentation of endocytosed antigen as peptide/class I MHC complexes plays a central role in the elicitation of CD8+ T cell clones that mediate anti-viral and anti-tumor immune responses. While it has been clear that there are specific subsets of professional antigen presenting cells (APC capable of antigen cross-presentation, description of mechanisms involved is still ongoing. Especially amongst dendritic cells (DC, there are specialized subsets that are highly proficient at antigen cross-presentation. We here present a focused survey on the cell biological processes in the endosomal pathway that support antigen cross-presentation. This review highlight DC-intrinsic mechanisms that facilitate the cross-presentation of endocytosed antigen, including receptor-mediated uptake, recycling and maturation including the sorting of membrane proteins, dynamic remodeling of endosomal structures and cell-surface directed endosomal trafficking. We will conclude with description of pathogen-induced deviation of endosomal processing, and discuss how immune evasion strategies pertaining endosomal trafficking may preclude antigen cross-presentation.

  4. Cell Wall-Associated Protein Antigens of Streptococcus salivarius: Purification, Properties, and Function in Adherence

    OpenAIRE

    Weerkamp, Anton H.; Jacobs, Ton

    1982-01-01

    Three cell wall-associated protein antigens (antigens b, c, and d) were isolated from mutanolysin-solubilized cell walls of Streptococcus salivarius HB and purified to apparent homogeneity by a combination of ion-exchange chromatography, gel filtration, and immunoadsorption chromatography. Antigens b and c were also isolated from culture supernatants. Antigen b consisted of more than 80% protein and had an apparent molecular weight as determined by sodium dodecyl sulfate-polyacrylamide gel el...

  5. Protein C activity and antigen levels in childhood

    NARCIS (Netherlands)

    van Teunenbroek, A.; Peters, M.; Sturk, A.; Borm, J. J.; Breederveld, C.

    1990-01-01

    Hereditary protein C deficiency is an important risk factor for thrombosis. To enable its diagnosis shortly after birth, we determined reference values of protein C antigen and activity levels for the first 3 months of life. To establish an age-related range of protein C levels we also determined

  6. Effects of proteolytic enzymes and neuraminidase on the I and i erythrocyte antigen sites

    International Nuclear Information System (INIS)

    Doinel, C.; Ropars, C.; Salmon, C.

    1978-01-01

    Homogeneous cold agglutinins, purified and labelled with 125 I, have been used in a study of the effects of neuraminidase and proteolytic enzymes on the I and i reactivities of human adult erythrocytes. Measurements were made of antigen site numbers, equilibrium constants and thermodynamic parameters. There was enhanced reactivity after enzyme treatment as well as after the release of N-acetylneuraminic acid. Steric factors were shown to be of primary importance in the accessibility of the I and i antigenic determinant. After enzyme treatment, the antigenic structures became more homogeneous in their reaction with antibodies. The heterogeneity of binding constants observed with antigenic determinants of non-treated erythrocytes is probably due to the wide range of spatial distribution of these receptors within the membrane. (author)

  7. Deteksi Antigen pada Kriptokokosis

    Directory of Open Access Journals (Sweden)

    Robiatul Adawiyah

    2014-12-01

    , laboratory and radiologicalexaminations. Laboratory examinations performed by morphological identification, serology and PCR. Morphological examination with India ink is positive when the number of fungi is around 10 10  cells/ml. Cultur examination is performed in Sabouraud dextrose agar (SDA and niger sheed agar (NSA medium, fungi grows in 5-7 days. Antigen and antibody detection could be performed on body fluid and do not take a long time. Detection of Cr. neoformans antibody can not show positive result in acute infection, IgA still positive after 1-2 years of healing phase and IgG can be persistent. The immunocompromised person showed very complex result and inconsistent in determining the diagnosis. Polysaccharides are the most instrumental component in Cr. neoformans virulence. The component of Polysaccharide especially glucuronoxylomannan is the most important marker in thediagnosis of cryptococcosis. Antigen detection of Cr. neoformans can show positive result in acute/chronic infection, high sensitivity and specificity. Polysaccharides can be detected from 10 ng/ ml of body fluid, so in minimal level of antigen we still can diagnose cryptococcosis.Keywords: Cr. neoformans, glucuronoxylomannan, antigen

  8. Antigenic and genetic characterization of a divergent African virus, Ikoma lyssavirus.

    Science.gov (United States)

    Horton, Daniel L; Banyard, Ashley C; Marston, Denise A; Wise, Emma; Selden, David; Nunez, Alejandro; Hicks, Daniel; Lembo, Tiziana; Cleaveland, Sarah; Peel, Alison J; Kuzmin, Ivan V; Rupprecht, Charles E; Fooks, Anthony R

    2014-05-01

    In 2009, a novel lyssavirus (subsequently named Ikoma lyssavirus, IKOV) was detected in the brain of an African civet (Civettictis civetta) with clinical rabies in the Serengeti National Park of Tanzania. The degree of nucleotide divergence between the genome of IKOV and those of other lyssaviruses predicted antigenic distinction from, and lack of protection provided by, available rabies vaccines. In addition, the index case was considered likely to be an incidental spillover event, and therefore the true reservoir of IKOV remained to be identified. The advent of sensitive molecular techniques has led to a rapid increase in the discovery of novel viruses. Detecting viral sequence alone, however, only allows for prediction of phenotypic characteristics and not their measurement. In the present study we describe the in vitro and in vivo characterization of IKOV, demonstrating that it is (1) pathogenic by peripheral inoculation in an animal model, (2) antigenically distinct from current rabies vaccine strains and (3) poorly neutralized by sera from humans and animals immunized against rabies. In a laboratory mouse model, no protection was elicited by a licensed rabies vaccine. We also investigated the role of bats as reservoirs of IKOV. We found no evidence for infection among 483 individuals of at least 13 bat species sampled across sites in the Serengeti and Southern Kenya.

  9. Antigenic Relationships among Human Pathogenic Orientia tsutsugamushi Isolates from Thailand.

    Directory of Open Access Journals (Sweden)

    Sarah L James

    2016-06-01

    Full Text Available Scrub typhus is a common cause of undiagnosed febrile illness in certain tropical regions, but can be easily treated with antibiotics. The causative agent, Orientia tsutsugamushi, is antigenically variable which complicates diagnosis and efforts towards vaccine development.This study aimed to dissect the antigenic and genetic relatedness of O. tsutsugamushi strains and investigate sero-diagnostic reactivities by titrating individual patient sera against their O. tsutsugamushi isolates (whole-cell antigen preparation, in homologous and heterologous serum-isolate pairs from the same endemic region in NE Thailand. The indirect immunofluorescence assay was used to titrate Orientia tsutsugamushi isolates and human sera, and a mathematical technique, antigenic cartography, was applied to these data to visualise the antigenic differences and cross-reactivity between strains and sera. No functional or antigen-specific analyses were performed. The antigenic variation found in clinical isolates was much less pronounced than the genetic differences found in the 56kDa type-specific antigen genes. The Karp-like sera were more broadly reactive than the Gilliam-like sera.Antigenic cartography worked well with scrub typhus indirect immunofluorescence titres. The data from humoral responses suggest that a Karp-like strain would provide broader antibody cross-reactivity than a Gilliam-like strain. Although previous exposure to O. tsutsugamushi could not be ruled out, scrub typhus patient serum antibody responses were characterised by strong homologous, but weak heterologous antibody titres, with little evidence for cross-reactivity by Gilliam-like sera, but a broader response from some Karp-like sera. This work highlights the importance of antigenic variation in O. tsutsugamushi diagnosis and determination of new serotypes.

  10. Electrochemical immunoassay for the prostate specific antigen using a reduced graphene oxide functionalized with a high molecular-weight silk peptide

    International Nuclear Information System (INIS)

    Wang, Yanying; Qu, Ying; Li, Chunya; Wu, Kangbing; Liu, Guishen; Hou, Xiaodong; Huang, Yina; Wu, Wangze

    2015-01-01

    High molecular-weight silk peptide (SP) was used to functionalize the surface of nanosheets of reduced graphene oxide (rGO). The SP-rGO nanocomposite was then mixed with mouse anti-human prostate specific antigen monoclonal antibody (anti-PSA) and coated onto a glassy carbon electrode to fabricate an immunosensor. By using the hexacyanoferrate redox system as electroactive probe, the immunosensor was characterized by voltammetry and electrochemical impedance spectroscopy. The peak current, measured at the potential of 0.24 V (vs. SCE), is distinctly reduced after binding prostate specific antigen (PSA). Response (measured by differential pulse voltammetry) is linearly related to PSA concentration in the range from 0.1 to 5.0 ng · mL −1 and from 5.0 to 80.0 ng∙mL −1 , and the detection limit is 53 pg∙mL −1 (at an SNR of 3). The immunosensor was successfully applied to the determination of PSA in clinical serum samples, and the results were found to agree well with those obtained with an enzyme-linked immunosorbent assay. (author)

  11. Two distinct forms of Chlamydia psittaci associated with disease and infertility in Phascolarctos cinereus (koala).

    Science.gov (United States)

    Girjes, A A; Hugall, A F; Timms, P; Lavin, M F

    1988-01-01

    While several diseases associated with Chlamydia psittaci infection have been reported in Phascolarctos cinereus (koala), it is still unclear whether one or more chlamydial strains are responsible. In this study, we provide evidence, obtained by restriction enzyme and gene probe analysis, that two quite distinct strains of C. psittaci infect koalas; one strain was isolated from the conjunctivae, and the other was isolated from the urogenital tract and the rectum. A gene probe, pFEN207, containing the coding sequence for an enzyme involved in the biosynthesis of the chlamydial genus-specific lipopolysaccharide antigen, and a separate probe, pCPML-4N, prepared from a DNA fragment of a koala-infecting strain of C. psittaci, were used to determine the patterns of hybridization in the koala-infecting strains; these patterns were found to be quite distinct from those observed with C. psittaci isolates from other animals. We also demonstrated by hybridization analysis with an avian strain plasmid that all three koala urogenital isolates contain a plasmid and that there is no evidence for the presence of a homologous plasmid in any of the ocular isolates. Images PMID:3397180

  12. Identification of a peptide binding protein that plays a role in antigen presentation

    International Nuclear Information System (INIS)

    Lakey, E.K.; Margoliash, E.; Pierce, S.K.

    1987-01-01

    The helper T-cell response to globular proteins appears, in general, to require intracellular processing of the antigen, such that a peptide fragment containing the T-cell antigenic determinant is released and transported to and held on the surface of an Ia-expressing, antigen-presenting cell. However, the molecular details underlying these phenomena are largely unknown. The means by which antigenic peptides are anchored on the antigen-presenting cell surface was investigated. A cell surface protein is identified that was isolated by it ability to bind to a 24-amino acid peptide fragment of pigeon cytochrome c, residues 81-104, containing the major antigenic determinant for B10.A mouse T cells. This peptide binding protein, purified from [ 35 S]methionine-labeled cells, appears as two discrete bands of ≅72 and 74 kDa after NaDodSO 4 /PAGE. The protein can be eluted from the peptide affinity column with equivalent concentrations of either the antigenic pigeon cytochrome c peptide or the corresponding nonantigenic peptide of mouse cytochrome c. However, it does not bind to the native cytochromes c, either of pigeon or mouse, and thus the protein appears to recognize some structure available only in the free peptides. This protein plays a role in antigen presentation. Its expression is not major histocompatibility complex-restricted in that the blocking activity of the antisera can be absorbed on spleen cells from mice of different haplotypes. This peptide binding protein can be isolated from a variety of cell types, including B cells, T cells, and fibroblasts. The anchoring of processed peptides on the cell surface by such a protein may play a role in antigen presentation

  13. CD8(+)NKT-like cells regulate the immune response by killing antigen-bearing DCs.

    Science.gov (United States)

    Wang, Chao; Liu, Xi; Li, Zhengyuan; Chai, Yijie; Jiang, Yunfeng; Wang, Qian; Ji, Yewei; Zhu, Zhongli; Wan, Ying; Yuan, Zhenglong; Chang, Zhijie; Zhang, Minghui

    2015-09-15

    CD1d-dependent NKT cells have been extensively studied; however, the function of CD8(+)NKT-like cells, which are CD1d-independent T cells with NK markers, remains unknown. Here, we report that CD1d-independent CD8(+)NKT-like cells, which express both T cell markers (TCRβ and CD3) and NK cell receptors (NK1.1, CD49b and NKG2D), are activated and significantly expanded in mice immunized with GFP-expressing dendritic cells. Distinct from CD1d-dependent NKT cells, CD8(+)NKT-like cells possess a diverse repertoire of TCRs and secrete high levels of IFN-gamma but not IL-4. CD8(+)NKT-like cell development is normal in CD1d(-/-) mice, which suggests that CD8(+)NKT-like cells undergo a unique development pathway that differs from iNKT cells. Further functional analyses show that CD8(+)NKT-like cells suppress T-cell responses through elimination of dendritic cells in an antigen-specific manner. Adoptive transfer of antigen-specific CD8(+)NKT-like cells into RIP-OVA mice prevented subsequent development of diabetes in the animals induced by activated OT-I CD8 T cells. Our study suggests that CD8(+)NKT-like cells can function as antigen-specific suppressive cells to regulate the immune response through killing antigen-bearing DCs. Antigen-specific down regulation may provide an active and precise method for constraining an excessive immune response and avoiding bypass suppression of necessary immune responses to other antigens.

  14. CD8+NKT-like cells regulate the immune response by killing antigen-bearing DCs

    Science.gov (United States)

    Wang, Chao; Liu, Xi; Li, Zhengyuan; Chai, Yijie; Jiang, Yunfeng; Wang, Qian; Ji, Yewei; Zhu, Zhongli; Wan, Ying; Yuan, Zhenglong; Chang, Zhijie; Zhang, Minghui

    2015-01-01

    CD1d-dependent NKT cells have been extensively studied; however, the function of CD8+NKT-like cells, which are CD1d-independent T cells with NK markers, remains unknown. Here, we report that CD1d-independent CD8+NKT-like cells, which express both T cell markers (TCRβ and CD3) and NK cell receptors (NK1.1, CD49b and NKG2D), are activated and significantly expanded in mice immunized with GFP-expressing dendritic cells. Distinct from CD1d-dependent NKT cells, CD8+NKT-like cells possess a diverse repertoire of TCRs and secrete high levels of IFN-gamma but not IL-4. CD8+NKT-like cell development is normal in CD1d−/− mice, which suggests that CD8+NKT-like cells undergo a unique development pathway that differs from iNKT cells. Further functional analyses show that CD8+NKT-like cells suppress T-cell responses through elimination of dendritic cells in an antigen-specific manner. Adoptive transfer of antigen-specific CD8+NKT-like cells into RIP-OVA mice prevented subsequent development of diabetes in the animals induced by activated OT-I CD8 T cells. Our study suggests that CD8+NKT-like cells can function as antigen-specific suppressive cells to regulate the immune response through killing antigen-bearing DCs. Antigen-specific down regulation may provide an active and precise method for constraining an excessive immune response and avoiding bypass suppression of necessary immune responses to other antigens. PMID:26369936

  15. Levels of estrogen, carcinoembryonic antigen and cancer antigen of breast in Sudanese female with breast cancer

    International Nuclear Information System (INIS)

    Abdelhadi, H. A.; Sirelkhatim, D. A.; Eltayeb, E. A.; Ahmed, W. A.; Elhussein, B.

    2006-12-01

    This study was conducted during the period from february 2004 to july 2004; with the objective of measuring the levels of estrogen (E2), carcinoembryonic antigen (CEA) and cancer antigen of breast (CA-15.3) so as to facilitate the early diagnosis of breast cancer and to determine the involvement of these parameters as risk factors for breast cancer. Ninety blood samples were collected from Sudanese females, divided into two groups; control group and patients groups. The patients group was sixty Sudanese females visiting the Radio Isotope Center, Khartoum (RICK) and they were confirmed as breast cancer patients by histopathology. The levels of the above mentioned parameters were determined by using radioimmunoassay technique. The results showed that , no significant (P=0.05) difference between the levels of the estrogen in patients compared to the control, on the other hand, there was non-significant (p<0.05) elevation in CEA levels in the patients with breast cancer compared to the control. The levels of CA 15.3 was significantly (p<0.0001) higher in the breast cancer patients compared to the control.(Author)

  16. The translocon protein Sec61 mediates antigen transport from endosomes in the cytosol for cross-presentation to CD8(+) T cells.

    Science.gov (United States)

    Zehner, Matthias; Marschall, Andrea L; Bos, Erik; Schloetel, Jan-Gero; Kreer, Christoph; Fehrenschild, Dagmar; Limmer, Andreas; Ossendorp, Ferry; Lang, Thorsten; Koster, Abraham J; Dübel, Stefan; Burgdorf, Sven

    2015-05-19

    The molecular mechanisms regulating antigen translocation into the cytosol for cross-presentation are under controversial debate, mainly because direct data is lacking. Here, we have provided direct evidence that the activity of the endoplasmic reticulum (ER) translocon protein Sec61 is essential for endosome-to-cytosol translocation. We generated a Sec61-specific intrabody, a crucial tool that trapped Sec61 in the ER and prevented its recruitment into endosomes without influencing Sec61 activity and antigen presentation in the ER. Expression of this ER intrabody inhibited antigen translocation and cross-presentation, demonstrating that endosomal Sec61 indeed mediates antigen transport across endosomal membranes. Moreover, we showed that the recruitment of Sec61 toward endosomes, and hence antigen translocation and cross-presentation, is dependent on dendritic cell activation by Toll-like receptor (TLR) ligands. These data shed light on a long-lasting question regarding antigen cross-presentation and point out a role of the ER-associated degradation machinery in compartments distinct from the ER. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Increased serum anti-mycobacterial antibody titers in rheumatoid arthritis patients: Is there any specific antigenic target?

    International Nuclear Information System (INIS)

    Cetin, Emel S.; Aksoy, Ali M

    2007-01-01

    Objective was to investigate the presence of immunoreactivity against mycobacterial antigens in the sera of patients with rheumatoid arthritis (Ra) and to detect the target of the immune reaction. This study was carried out on 60 patients with RA, and 25 patients with no joint diseases in the laboratory of Clinical Microbiology Department of Ankara University Medical Faculty, Ankara, Turkey between July 2003 to January 2004. Secreted and cellular antigens of Mycobacterium tuberculosis (M. tuberculosis) H37Rv and Mycobacterium bovis (M. bovis) were isolated and purified by high performance liquid chromatography to antigenic fractions. The immunoreactivity of patient and control sera against these antigens were determined by enzyme-linked immunosorbent assay (ELISA). Immunoreactivity against mycobacterial antigens in RA patients were significantly higher than controls. Significant difference between patients and controls has been determined with M. bovis Bacillus Calmette Guerin (BCG) culture fluid and sonicate antigens, but not with M. tuberculosis H37Rv. This suggests that the antigen triggering immune response in patients with RA may belong to or mainly expressed on M. bovis BCG. The ELISA results showed significant difference between RA patients and controls with all antigenic fractions. Presence of increased immunoreactivity against mycobacterial antigens in the sera of patients with RA was detected. When statistical analysis was considered, we cannot put forward any antigenic fraction alone as the one responsible for the increased reactivity. (author)

  18. Determination of the frequency of the most immunogenic Rhesus antigens among Saudi donors in King Abdulaziz Medical City ? Riyadh

    OpenAIRE

    Elsayid, Mohieldin; Al Qahtani, Faris Saeed; Al Qarni, Abdulaziz Mohammed; Almajed, Faisal; Al Saqri, Faisal; Qureshi, Shoeb

    2017-01-01

    Background: The Rhesus (Rh) blood group system is one of the most polymorphic and immunogenic systems known in humans, because of its immunogenicity along with ABO grouping, RhD antigen testing was made mandatory before issuing a compatible blood. At present, there are five major antigens, i.e., D, C, E, c, and e in Rh blood group system. Aims: The aim of this study is to provide essential data about the distribution of the major Rh antigens and the most common phenotype among the Saudi popul...

  19. TCRs Used in Cancer Gene Therapy Cross-React with MART-1/Melan-A Tumor Antigens via Distinct Mechanisms

    Energy Technology Data Exchange (ETDEWEB)

    Borbulevych, Oleg Y.; Santhanagopolan, Sujatha M.; Hossain, Moushumi; Baker, Brian M. (Notre)

    2013-09-18

    T cells engineered to express TCRs specific for tumor Ags can drive cancer regression. The first TCRs used in cancer gene therapy, DMF4 and DMF5, recognize two structurally distinct peptide epitopes of the melanoma-associated MART-1/Melan-A protein, both presented by the class I MHC protein HLA-A*0201. To help understand the mechanisms of TCR cross-reactivity and provide a foundation for the further development of immunotherapy, we determined the crystallographic structures of DMF4 and DMF5 in complex with both of the MART-1/Melan-A epitopes. The two TCRs use different mechanisms to accommodate the two ligands. Although DMF4 binds the two with a different orientation, altering its position over the peptide/MHC, DMF5 binds them both identically. The simpler mode of cross-reactivity by DMF5 is associated with higher affinity toward both ligands, consistent with the superior functional avidity of DMF5. More generally, the observation of two diverging mechanisms of cross-reactivity with the same Ags and the finding that TCR-binding orientation can be determined by peptide alone extend our understanding of the mechanisms underlying TCR cross-reactivity.

  20. Prevalence of Weak D Antigen In Western Indian Population

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    Tanvi Sadaria

    2015-12-01

    Full Text Available Introduction: Discovery of Rh antigens in 1939 by Landsteiner and Weiner was the revolutionary stage in blood banking. Of these antigens, D, which decides Rh positivity or negativity, is the most antigenic. A problem is encountered when an individual has a weakened expression of D (Du, i.e., fewer numbers of D antigens on red cell membrane. Aims and Objectives: To know the prevalence of weak D in Indian population because incidence varies in different population. To determine the risk of alloimmunization among Rh D negative patients who receives the blood of weak D positive donors. Material and Methods: Rh grouping of 38,962 donors who came to The Department of Immunohematology and Blood Transfusion of Civil Hospital, Ahmedabad from 1st January 2013 to 30th September 2014 was done using the DIAGAST (Automated Grouping. The samples that tested negative for D antigen were further analysed for weak D (Du by indirect antiglobulin test using blend of Ig G and Ig M Anti D. This was done using Column agglutination method in ID card (gel card. Results: The total number of donors studied was 38,962. Out of these 3360(8.6% were tested Rh D negative. All Rh D negative donors were tested for weak D (Du. 22 (0.056% of total donors and 0.65% of Rh negative donors turned out to be weak D (Du positive. Conclusion: The prevalence of weak D (Du in Western Indian population is 0.056 %, So the risk of alloimmunization in our setting due to weak D (Du antigen is marginal. But, testing of weak D antigen is necessary in blood bank because weak D antigen is immunogenic and can produce alloimmunization if transfused to Rh D negative subjects.

  1. Phenotypic and functional profiling of CD4 T cell compartment in distinct populations of healthy adults with different antigenic exposure.

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    Sophie Roetynck

    Full Text Available Multiparameter flow cytometry has revealed extensive phenotypic and functional heterogeneity of CD4 T cell responses in mice and humans, emphasizing the importance of assessing multiple aspects of the immune response in correlation with infection or vaccination outcome. The aim of this study was to establish and validate reliable and feasible flow cytometry assays, which will allow us to characterize CD4 T cell population in humans in field studies more fully.We developed polychromatic flow cytometry antibody panels for immunophenotyping the major CD4 T cell subsets as well as broadly characterizing the functional profiles of the CD4 T cells in peripheral blood. We then validated these assays by conducting a pilot study comparing CD4 T cell responses in distinct populations of healthy adults living in either rural or urban Kenya. This study revealed that the expression profile of CD4 T cell activation and memory markers differed significantly between African and European donors but was similar amongst African individuals from either rural or urban areas. Adults from rural Kenya had, however, higher frequencies and greater polyfunctionality among cytokine producing CD4 T cells compared to both urban populations, particularly for "Th1" type of response. Finally, endemic exposure to malaria in rural Kenya may have influenced the expansion of few discrete CD4 T cell populations with specific functional signatures.These findings suggest that environmentally driven T cell activation does not drive the dysfunction of CD4 T cells but is rather associated with greater magnitude and quality of CD4 T cell response, indicating that the level or type of microbial exposure and antigenic experience may influence and shape the functionality of CD4 T cell compartment. Our data confirm that it is possible and mandatory to assess multiple functional attributes of CD4 T cell response in the context of infection.

  2. Antigen-specific murine T cell clones produce soluble interleukin 2 receptor on stimulation with specific antigens

    International Nuclear Information System (INIS)

    Wagner, D.K.; York-Jolley, J.; Malek, T.R.; Berzofsky, J.A.; Nelson, D.L.

    1986-01-01

    In this study, monoclonal antibodies were used to the murine IL 2 receptor (IL 2R) termed 3C7 and 7D4, which bind to different epitopes on the murine IL 2R, to develop an ELISA to measure soluble murine IL 2R. Surprisingly, stimulated murine spleen cells not only expressed cell-associated IL 2R, but also produced a considerable level of cellfree IL 2R in the culture supernatant fluid. To assess the fine specificity of this response, myoglobin-immune murine T cell clones were stimulated with appropriate or inappropriate antigen and syngeneic or allogeneic presenting cells. Proliferation, measured by [ 3 H] thymidine incorporation, and levels of soluble IL 2R were determined at day 4. The production of soluble IL2R displayed the same epitope fine specificity, genetic restriction, and antigen dose-response as the proliferative response. Indeed, in some cases there was sharper discrimination of epitope specificity and genetic restriction with the soluble IL 2R levels. There was also reproducible clone-to-clone variation in the amount of soluble receptor produced in response to antigen among 12 T cell clones and lines tested. In time course experiments, proliferation was greatest at day 3, whereas soluble IL 2R levels continued to rise in subsequent days. To the authors' knowledge, this is the first demonstration of release of secretion of soluble IL 2R by murine T cells, and the first demonstration of the fine specificity and genetic restriction of the induction of soluble IL 2R by specific antigen

  3. The O-antigen structure of bacterium Comamonas aquatica CJG.

    Science.gov (United States)

    Wang, Xiqian; Kondakova, Anna N; Zhu, Yutong; Knirel, Yuriy A; Han, Aidong

    2017-11-01

    Genus Comamonas is a group of bacteria that are able to degrade a variety of environmental waste. Comamonas aquatica CJG (C. aquatica) in this genus is able to absorb low-density lipoprotein but not high-density lipoprotein of human serum. Using 1 H and 13 C NMR spectroscopy, we found that the O-polysaccharide (O-antigen) of this bacterium is comprised of a disaccharide repeat (O-unit) of d-glucose and 2-O-acetyl-l-rhamnose, which is shared by Serratia marcescens O6. The O-antigen gene cluster of C. aquatica, which is located between coaX and tnp4 genes, contains rhamnose synthesis genes, glycosyl and acetyl transferase genes, and ATP-binding cassette transporter genes, and therefore is consistent with the O-antigen structure determined here.

  4. Mucosal-Associated Invariant T Cells: New Insights into Antigen Recognition and Activation

    Directory of Open Access Journals (Sweden)

    Xingxing Xiao

    2017-11-01

    Full Text Available Mucosal-associated invariant T (MAIT cells, a novel subpopulation of innate-like T cells that express an invariant T cell receptor (TCRα chain and a diverse TCRβ chain, can recognize a distinct set of small molecules, vitamin B metabolites, derived from some bacteria, fungi but not viruses, in the context of an evolutionarily conserved major histocompatibility complex-related molecule 1 (MR1. This implies that MAIT cells may play unique and important roles in host immunity. Although viral antigens are not recognized by this limited TCR repertoire, MAIT cells are known to be activated in a TCR-independent mechanism during some viral infections, such as hepatitis C virus and influenza virus. In this article, we will review recent works in MAIT cell antigen recognition, activation and the role MAIT cells may play in the process of bacterial and viral infections and pathogenesis of non-infectious diseases.

  5. A negative feedback modulator of antigen processing evolved from a frameshift in the cowpox virus genome.

    Directory of Open Access Journals (Sweden)

    Jiacheng Lin

    2014-12-01

    Full Text Available Coevolution of viruses and their hosts represents a dynamic molecular battle between the immune system and viral factors that mediate immune evasion. After the abandonment of smallpox vaccination, cowpox virus infections are an emerging zoonotic health threat, especially for immunocompromised patients. Here we delineate the mechanistic basis of how cowpox viral CPXV012 interferes with MHC class I antigen processing. This type II membrane protein inhibits the coreTAP complex at the step after peptide binding and peptide-induced conformational change, in blocking ATP binding and hydrolysis. Distinct from other immune evasion mechanisms, TAP inhibition is mediated by a short ER-lumenal fragment of CPXV012, which results from a frameshift in the cowpox virus genome. Tethered to the ER membrane, this fragment mimics a high ER-lumenal peptide concentration, thus provoking a trans-inhibition of antigen translocation as supply for MHC I loading. These findings illuminate the evolution of viral immune modulators and the basis of a fine-balanced regulation of antigen processing.

  6. Systems Analysis Reveals High Genetic and Antigen-Driven Predetermination of Antibody Repertoires throughout B Cell Development

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    Victor Greiff

    2017-05-01

    Full Text Available Antibody repertoire diversity and plasticity is crucial for broad protective immunity. Repertoires change in size and diversity across multiple B cell developmental stages and in response to antigen exposure. However, we still lack fundamental quantitative understanding of the extent to which repertoire diversity is predetermined. Therefore, we implemented a systems immunology framework for quantifying repertoire predetermination on three distinct levels: (1 B cell development (pre-B cell, naive B cell, plasma cell, (2 antigen exposure (three structurally different proteins, and (3 four antibody repertoire components (V-gene usage, clonal expansion, clonal diversity, repertoire size extracted from antibody repertoire sequencing data (400 million reads. Across all three levels, we detected a dynamic balance of high genetic (e.g., >90% for V-gene usage and clonal expansion in naive B cells and antigen-driven (e.g., 40% for clonal diversity in plasma cells predetermination and stochastic variation. Our study has implications for the prediction and manipulation of humoral immunity.

  7. Intra-uterine exposure of horses to Sarcocystis spp. antigens

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    A.M. Antonello

    2016-04-01

    Full Text Available The aim of this study was to examine the intra-uterine exposure to Sarcocystis spp. antigens, determining the number of foals with detectable concentrations of antibodies against these agents in the serum, before colostrum ingestion and collect data about exposure of horses to the parasite. Serum samples were collected from 195 thoroughbred mares and their newborns in two farms from southern Brazil. Parasite specific antibody responses to Sarcocystis antigens were detected using the indirect immunofluorescent antibody test (IFAT and immunoblot analysis. In 84.1% (159/189 of the pregnant mares and in 7.4% (14/189 of foals we detected antibodies anti-Sarcocystis spp. by IFAT. All samples seropositive from foals were also positive in their respective mares. Serum samples of seropositive foals by IFAT, showed no reactivity on the immunoblot, having as antigens S. neurona merozoites. In conclusion, the intra-uterine exposure to Sarcocystis spp. antigens in horses was demonstrated, with occurrence not only in mares, but also in their foals, before colostrum ingestion these occurrences were reduced.

  8. Distinct neutrophil subpopulations phenotype by flow cytometry in myelodysplastic syndromes.

    Science.gov (United States)

    Vikentiou, Myrofora; Psarra, Katerina; Kapsimali, Violetta; Liapis, Konstantinos; Michael, Michalis; Tsionos, Konstantinos; Lianidou, Evi; Papasteriades, Chryssa

    2009-03-01

    The cardinal feature of myelodysplastic syndromes (MDS) is dysplasia involving one or more myeloid cell lineages. In the present study, we used 4-color flow cytometric analysis to investigate dysgranulopoiesis in bone marrow specimens from 65 patients with MDS. The antigen expression patterns of total neutrophil granulocytes (TNG) and of the two distinct neutrophil granulocytic subpopulations (NGSs), NGS-1 (dimmer CD45 expression) and NGS-2 (stronger CD45 expression) identified on the side scatter (SS) vs. CD45-intensity plot, were studied. The neutrophil granulocytes from patients with MDS showed characteristic antigen expression aberrancies which were more pronounced in NGS-2 subpopulation. Studying separately the NGS-2 subpopulation with the CD16/MPO/LF combination, the low CD16(+)/MPO(+) and low CD16(+)/LF(+) percentages seemed to discriminate between lower-risk and higher-risk patients with MDS in most occasions. Furthermore, a detailed assessment of the NGS-1 and NGS-2 immunophenotypic patterns revealed early dysplastic changes, not otherwise observed by standard TNG analysis, especially in cases of lower-risk MDS.

  9. RAPID DETECTION OF PNEUMOCOCCAL ANTIGEN IN PLEURAL FLUID OF PATIENTS WITH COMMUNITY ACQUIRED PNEUMONIA

    NARCIS (Netherlands)

    BOERSMA, WG; LOWENBERG, A; HOLLOWAY, Y; KUTTSCHRUTTER, H; SNIJDER, JAM; KOETER, GH

    Background Detection of pneumococcal antigen may help to increase the rate of diagnosis of pneumococcal pneumonia. This study was designed to determine the value of rapid detection of pneumococcal antigen in pleural fluid from patients with community acquired pneumonia. Methods Thoracentesis was

  10. Enhancement of MHC-I antigen presentation via architectural control of pH-responsive, endosomolytic polymer nanoparticles.

    Science.gov (United States)

    Wilson, John T; Postma, Almar; Keller, Salka; Convertine, Anthony J; Moad, Graeme; Rizzardo, Ezio; Meagher, Laurence; Chiefari, John; Stayton, Patrick S

    2015-03-01

    Protein-based vaccines offer a number of important advantages over organism-based vaccines but generally elicit poor CD8(+) T cell responses. We have previously demonstrated that pH-responsive, endosomolytic polymers can enhance protein antigen delivery to major histocompatibility complex class I (MHC-I) antigen presentation pathways thereby augmenting CD8(+) T cell responses following immunization. Here, we describe a new family of nanocarriers for protein antigen delivery assembled using architecturally distinct pH-responsive polymers. Reversible addition-fragmentation chain transfer (RAFT) polymerization was used to synthesize linear, hyperbranched, and core-crosslinked copolymers of 2-(N,N-diethylamino)ethyl methacrylate (DEAEMA) and butyl methacrylate (BMA) that were subsequently chain extended with a hydrophilic N,N-dimethylacrylamide (DMA) segment copolymerized with thiol-reactive pyridyl disulfide (PDS) groups. In aqueous solution, polymer chains assembled into 25 nm micellar nanoparticles and enabled efficient and reducible conjugation of a thiolated protein antigen, ovalbumin. Polymers demonstrated pH-dependent membrane-destabilizing activity in an erythrocyte lysis assay, with the hyperbranched and cross-linked polymer architectures exhibiting significantly higher hemolysis at pH ≤ 7.0 than the linear diblock. Antigen delivery with the hyperbranched and cross-linked polymer architecture enhanced in vitro MHC-I antigen presentation relative to free antigen, whereas the linear construct did not have a discernible effect. The hyperbranched system elicited a four- to fivefold increase in MHC-I presentation relative to the cross-linked architecture, demonstrating the superior capacity of the hyperbranched architecture in enhancing MHC-I presentation. This work demonstrates that the architecture of pH-responsive, endosomolytic polymers can have dramatic effects on intracellular antigen delivery, and offers a promising strategy for enhancing CD8(+) T cell

  11. Expression of Immunoglobulin Receptors with Distinctive Features Indicating Antigen Selection by Marginal Zone B Cells from Human Spleen

    Science.gov (United States)

    Colombo, Monica; Cutrona, Giovanna; Reverberi, Daniele; Bruno, Silvia; Ghiotto, Fabio; Tenca, Claudya; Stamatopoulos, Kostas; Hadzidimitriou, Anastasia; Ceccarelli, Jenny; Salvi, Sandra; Boccardo, Simona; Calevo, Maria Grazia; De Santanna, Amleto; Truini, Mauro; Fais, Franco; Ferrarini, Manlio

    2013-01-01

    Marginal zone (MZ) B cells, identified as surface (s)IgMhighsIgDlowCD23low/−CD21+CD38− B cells, were purified from human spleens, and the features of their V(D)J gene rearrangements were investigated and compared with those of germinal center (GC), follicular mantle (FM) and switched memory (SM) B cells. Most MZ B cells were CD27+ and exhibited somatic hypermutations (SHM), although to a lower extent than SM B cells. Moreover, among MZ B-cell rearrangements, recurrent sequences were observed, some of which displayed intraclonal diversification. The same diversifying sequences were detected in very low numbers in GC and FM B cells and only when a highly sensitive, gene-specific polymerase chain reaction was used. This result indicates that MZ B cells could expand and diversify in situ and also suggested the presence of a number of activation-induced cytidine deaminase (AID)-expressing B cells in the MZ. The notion of antigen-driven expansion/selection in situ is further supported by the VH CDR3 features of MZ B cells with highly conserved amino acids at specific positions and by the finding of shared (“stereotyped”) sequences in two different spleens. Collectively, the data are consistent with the notion that MZ B cells are a special subset selected by in situ antigenic stimuli. PMID:23877718

  12. Antigenic evaluation of a recombinant baculovirus-expressed Sarcocystis neurona SAG1 antigen.

    Science.gov (United States)

    Gupta, G D; Lakritz, J; Saville, W J; Livingston, R S; Dubey, J P; Middleton, J R; Marsh, A E

    2004-10-01

    Sarcocystis neurona is the primary parasite associated with equine protozoal myeloencephalitis (EPM). This is a commonly diagnosed neurological disorder in the Americas that infects the central nervous system of horses. Current serologic assays utilize culture-derived parasites as antigen. This method requires large numbers of parasites to be grown in culture, which is labor intensive and time consuming. Also, a culture-derived whole-parasite preparation contains conserved antigens that could cross-react with antibodies against other Sarcocystis species and members of Sarcocystidae such as Neospora spp., Hammondia spp., and Toxoplasma gondii. Therefore, there is a need to develop an improved method for the detection of S. neurona-specific antibodies. The sera of infected horses react strongly to surface antigen 1 (SnSAG1), an approximately 29-kDa protein, in immunoblot analysis, suggesting that it is an immunodominant antigen. The SnSAG1 gene of S. neurona was cloned, and recombinant S. neurona SAG1 protein (rSnSAG1-Bac) was expressed with the use of a baculovirus system. By immunoblot analysis, the rSnSAG1-Bac antigen detected antibodies to S. neurona from naturally infected and experimentally inoculated equids, cats, rabbit, mice, and skunk. This is the first report of a baculovirus-expressed recombinant S. neurona antigen being used to detect anti-S. neurona antibodies in a variety of host species.

  13. Remarkably similar antigen receptors among a subset of patients with chronic lymphocytic leukemia

    Science.gov (United States)

    Ghiotto, Fabio; Fais, Franco; Valetto, Angelo; Albesiano, Emilia; Hashimoto, Shiori; Dono, Mariella; Ikematsu, Hideyuki; Allen, Steven L.; Kolitz, Jonathan; Rai, Kanti R.; Nardini, Marco; Tramontano, Anna; Ferrarini, Manlio; Chiorazzi, Nicholas

    2004-01-01

    Studies of B cell antigen receptors (BCRs) expressed by leukemic lymphocytes from patients with B cell chronic lymphocytic leukemia (B-CLL) suggest that B lymphocytes with some level of BCR structural restriction become transformed. While analyzing rearranged VHDJH and VLJL genes of 25 non–IgM-producing B-CLL cases, we found five IgG+ cases that display strikingly similar BCRs (use of the same H- and L-chain V gene segments with unique, shared heavy chain third complementarity-determining region [HCDR3] and light chain third complementarity-determining region [LCDR3] motifs). These H- and L-chain characteristics were not identified in other B-CLL cases or in normal B lymphocytes whose sequences are available in the public databases. Three-dimensional modeling studies suggest that these BCRs could bind the same antigenic epitope. The structural features of the B-CLL BCRs resemble those of mAb’s reactive with carbohydrate determinants of bacterial capsules or viral coats and with certain autoantigens. These findings suggest that the B lymphocytes that gave rise to these IgG+ B-CLL cells were selected for this unique BCR structure. This selection could have occurred because the precursors of the B-CLL cells were chosen for their antigen-binding capabilities by antigen(s) of restricted nature and structure, or because the precursors derived from a B cell subpopulation with limited BCR heterogeneity, or both. PMID:15057307

  14. Potential radioimmunoassay system for detection of Hanganutziu-Deicher type heterophile antigen(s) and antibodies in tissues and fluids

    Energy Technology Data Exchange (ETDEWEB)

    Mukuria, J C; Naiki, Masaharu; Hashimoto, Masato; Nishiura, Katsumi; Okabe, Masahiro; Kato, Shiro

    1985-06-12

    A relatively simple, specific and sensitive radioimmunoassay system has been developed for the detection of heterophile Hanganutziu-Deicher (H-D) antigen(s) and antibodies. The SVI-labeled H-D antigen-active molecule used for the assay is a bovine erythrocyte major glycoprotein previously found to have a strong H-D antigen potency. Different H-D antigen-active molecules were compared for heterophile H-D antigen potency. Eight different lung cancer tissues were assayed for H-D antigen. The sera from the 8 lung cancer patients were also screened by ELISA and RIA in an attmept to correlate expression of H-D antigen on tissues with elevation of H-D antibodies.

  15. Comparative characteristic of the methods of protein antigens epitope mapping

    Directory of Open Access Journals (Sweden)

    O. Yu. Galkin

    2014-08-01

    Full Text Available Comparative analysis of experimental methods of epitope mapping of protein antigens has been carried out. The vast majority of known techniques are involved in immunochemical study of the interaction of protein molecules or peptides with antibodies of corresponding specifici­ty. The most effective and widely applicable metho­dological techniques are those that use synthetic and genetically engineered peptides. Over the past 30 years, these groups of methods have travelled a notable evolutionary path up to the maximum automation and the detection of antigenic determinants of various types (linear and conformational epitopes, and mimotopes. Most of epitope searching algorithms were integrated into a computer program, which greatly facilitates the analysis of experimental data and makes it possible to create spatial models. It is possible to use comparative epitope mapping for solving the applied problems; this less time-consuming method is based on the analysis of competition between different antibodies interactions with the same antigen. The physical method of antigenic structure study is X-ray analysis of antigen-antibody complexes, which may be applied only to crystallizing­ proteins, and nuclear magnetic resonance.

  16. A compound chimeric antigen receptor strategy for targeting multiple myeloma.

    Science.gov (United States)

    Chen, K H; Wada, M; Pinz, K G; Liu, H; Shuai, X; Chen, X; Yan, L E; Petrov, J C; Salman, H; Senzel, L; Leung, E L H; Jiang, X; Ma, Y

    2018-02-01

    Current clinical outcomes using chimeric-antigen receptors (CARs) against multiple myeloma show promise in the eradication of bulk disease. However, these anti-BCMA (CD269) CARs observe relapse as a common phenomenon after treatment due to the reemergence of either antigen-positive or -negative cells. Hence, the development of improvements in CAR design to target antigen loss and increase effector cell persistency represents a critical need. Here, we report on the anti-tumor activity of a CAR T-cell possessing two complete and independent CAR receptors against the multiple myeloma antigens BCMA and CS1. We determined that the resulting compound CAR (cCAR) T-cell possesses consistent, potent and directed cytotoxicity against each target antigen population. Using multiple mouse models of myeloma and mixed cell populations, we are further able to show superior in vivo survival by directed cytotoxicity against multiple populations compared to a single-expressing CAR T-cell. These findings indicate that compound targeting of BCMA and CS1 on myeloma cells can potentially be an effective strategy for augmenting the response against myeloma bulk disease and for initiation of broader coverage CAR therapy.

  17. Screening for epitope specificity directly on culture supernatants in the early phase of monoclonal antibody production by an ELISA with biotin-labeled antigen.

    Science.gov (United States)

    Andersen, Ditte C; Jensen, Charlotte H; Gregersen, Annemette; Brandt, Jette; Kliem, Anette; Skjødt, Karsten; Koch, Claus; Teisner, Børge

    2004-01-01

    This report describes an assay for comparison of epitope specificity in groups of monoclonal antibodies against a given antigen. The only prerequisite is the biotin-labeled antigen. One of the monoclonal antibodies is captured onto a plastic surface via a rabbit anti-mouse Ig, and the other preincubated with biotinylated antigen. When the two antibodies react with the same epitope subsequent binding of the biotin-labeled antigen is abolished (inhibition). In the cases where no inhibition was observed, the two antibodies were considered to react with distinct, independent epitopes. The obvious advantages using this assay, are that it can be performed directly on culture supernatants in the early phase of monoclonal antibody production, and also works for antigens with repetitive epitopes. Moreover, the bonus effect, i.e., a signal in excess of the reference signal when sets of monoclonal antibodies with different epitope specificity are compared, gives a relative measure of affinity.

  18. Targeting tumor antigens to secreted membrane vesicles in vivo induces efficient antitumor immune responses.

    Science.gov (United States)

    Zeelenberg, Ingrid S; Ostrowski, Matias; Krumeich, Sophie; Bobrie, Angélique; Jancic, Carolina; Boissonnas, Alexandre; Delcayre, Alain; Le Pecq, Jean-Bernard; Combadière, Béhazine; Amigorena, Sebastian; Théry, Clotilde

    2008-02-15

    Expression of non-self antigens by tumors can induce activation of T cells in vivo, although this activation can lead to either immunity or tolerance. CD8+ T-cell activation can be direct (if the tumor expresses MHC class I molecules) or indirect (after the capture and cross-presentation of tumor antigens by dendritic cells). The modes of tumor antigen capture by dendritic cells in vivo remain unclear. Here we examine the immunogenicity of the same model antigen secreted by live tumors either in association with membrane vesicles (exosomes) or as a soluble protein. We have artificially addressed the antigen to secreted vesicles by coupling it to the factor VIII-like C1C2 domain of milk fat globule epidermal growth factor-factor VIII (MFG-E8)/lactadherin. We show that murine fibrosarcoma tumor cells that secrete vesicle-bound antigen grow slower than tumors that secrete soluble antigen in immunocompetent, but not in immunodeficient, host mice. This growth difference is due to the induction of a more potent antigen-specific antitumor immune response in vivo by the vesicle-bound than by the soluble antigen. Finally, in vivo secretion of the vesicle-bound antigen either by tumors or by vaccination with naked DNA protects against soluble antigen-secreting tumors. We conclude that the mode of secretion can determine the immunogenicity of tumor antigens and that manipulation of the mode of antigen secretion may be used to optimize antitumor vaccination protocols.

  19. Mini-review: Can non-human leucocyte antigen genes determine susceptibility to severe dengue syndromes?

    Science.gov (United States)

    Ng, Dorothy; Ghosh, Aparna; Jit, Mark; Seneviratne, Suranjith L

    2017-09-01

    Dengue viral infections are endemic or epidemic in virtually all tropical countries. Among individuals infected with the dengue virus, severe dengue syndromes (i.e., dengue haemorrhagic fever and dengue shock syndromes) tend to affect only some and this may be due to a combination of host genetic susceptibility and viral factors. In this review article we analyse and discuss the present knowledge of non-human leucocyte antigen host genetic susceptibility to severe dengue syndromes. The relevance of genetic polymorphisms in the pathways of antigen recognition, uptake, processing and presentation, activation of interferon α responses, mast cell and complement activation and T cell activation and dengue disease severity has been reviewed and analysed. © The Author(s) 2018. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Effect of ionizing radiation on the antigenic composition of typhoid bacteria

    International Nuclear Information System (INIS)

    Sinilova, N.G.; Nikolaeva, L.A.; Tumanyan, M.A.

    1978-01-01

    Changes in the antigenic composition of typhoid bacteria occurring during the exposure of microbial suspension to different doses of gamma radiation ( 60 Co) ranging between 0.5 and 3.0 Mrad were studied. Immunoelectrophoresis in agar was used to determine the antigenic composition of different samples of irradiated bacteria. The antigenic composition of bacteria irradiated with doses up to 2.5 Mrad was found to be similar to that of non-irradiated bacteria. Antigens demonstrated by means of Vi, H and O antisera are preserved in these bacteria. However, all irradiated bacteria in general slightly differ from non-irradiated bacteria; this is manifest in a different configuration and position of the precipitation lines in the cathodic part of the immunophoreograms. The content of the component migrating rapidly towards the cathode, evidently the O antigen in the R form, in the irradiated bacteria increases with the dose of radiation. No new serologically active substances, non-existent in non-irradiated bacteria, were found to appear in the process of irradiation. (author)

  1. Equine infectious anemia virus-infected dendritic cells retain antigen presentation capability

    International Nuclear Information System (INIS)

    Rivera, Julie A.; McGuire, Travis C.

    2005-01-01

    To determine if equine monocyte-derived dendritic cells (DC) were susceptible to equine infectious anemia virus (EIAV) infection, ex vivo-generated DC were infected with virus in vitro. EIAV antigen was detected by immunofluorescence 3 days post-infection with maximum antigen being detected on day 4, whereas there was no antigen detected in DC incubated with the same amount of heat-inactivated EIAV. No cytolytic activity was observed after EIAV WSU5 infection of DC. These monocyte-derived DC were more effective than macrophages and B cells in stimulating allogenic T lymphocytes. Both infected macrophages and DC stimulated similar levels of memory CTL responses in mixtures of CD8+ and CD4+ cells as detected with 51 Cr-release assays indicating that EIAV infection of DC did not alter antigen presentation. However, EIAV-infected DC were more effective than infected macrophages when used to stimulate memory CTL in isolated CD8+ cells. The maintenance of antigen processing and presenting function by EIAV-infected DC in vitro suggests that this function is maintained during in vivo infection

  2. SENSITIVITY AND SPECIFICITY OF URINARY BLADDER CANCER ANTIGEN FOR DIAGNOSIS OF BLADDER TUMOR;A COMPARATIVE STUDY WITH URINARY CYTOLOGY

    Directory of Open Access Journals (Sweden)

    K. Radkhah

    2005-06-01

    Full Text Available Cystoscopy and urinary cytology are currently the basis for diagnosis and ‎follow-up of bladder tumors. Research to find a sensitive and specific tumor ‎marker for diagnosis of bladder tumor is actively underway, however, due to low sensitivity ‎and high cost of cytology. This cross-sectional study was performed in 65 patients to evaluate whether urinary bladder ‎cancer (UBC antigen level can predict the presence of active bladder tumor. In patients with ‎inactive tumor, UBC antigen level was determined in addition to standard cystoscopy ‎and cytology for follow-up. Patients with active tumor were ‎subjected to standard treatment and UBC antigen level determination. UBC antigen ‎ levels were measured by ELISA, using monoclonal antibodies ‎specific for UBC antigen. As a control group, UBC antigen level ‎was also determined in 65 persons who had been referred for urinalysis for other reasons. ‎UBC antigen level more than 1 μg/L which was regarded as ‎positive was found in 49.4% of the patients. In control group, 96.9% had UBC antigen < 1μg/L‎. Mean UBC antigen level in patients was ‎3.77 μg/L while it was 0.508 μg/L in controls (P < 0.0001. Sensitivity of ‎UBC antigen was 53.3% and its specificity was 40%. Sensitivity and specificity of urinary cytology was 17.3% and 88.2%, respectively. This difference was statistically ‎significant (P < 0.001. UBC antigen is more sensitive than urinary cytology, although cytology still ‎retains its priority in specificity. It is not yet recommended to replace UBC antigen for ‎cytology due to its low specificity and not favorable sensitivity.

  3. Quantitative and qualitative characterization of human cancer-associated serum glycoprotein antigens expressing epitopes consisting of sialyl or sialyl-fucosyl type 1 chain.

    Science.gov (United States)

    Kannagi, R; Kitahara, A; Itai, S; Zenita, K; Shigeta, K; Tachikawa, T; Noda, A; Hirano, H; Abe, M; Shin, S

    1988-07-01

    The levels of carbohydrate antigens having epitopes consisting of type 1 chain (R----Gal beta 1----GlcNAc beta 1----3Gal beta 1----R) in the sera of patients with various malignant and nonmalignant disorders have been investigated with the use of three monoclonal antibodies, N-19-9, FH-7, and FH-9. Serum levels of 2----3 sialylated Lea antigen and 2----6 sialylated Lea antigen, defined respectively by antibodies N-19-9 and FH-7, were found to be frequently high in patients with cancer of the digestive system, particularly pancreatic cancer. High levels of 2----3,2----6 disialylated Lc4 antigen, defined by antibody FH-9, were less frequent in cancer patients when compared with the other two antigens. In patients with nonmalignant disorders, especially renal and autoimmune diseases, serum levels of the two type 1 chain antigens defined by FH-7 and FH-9 were more frequently high than that defined by N-19-9. Molecular weights and other general biochemical characteristics of serum mucin carrying the type 1 chain determinants were not significantly different in cancer patients as compared with patients with nonmalignant disorders. However, the degree of glycosylation of the antigen, as assessed by its solubility in perchloric acid, showed significant differences; i.e., the mucin antigen carrying 2----6 sialylated Lea determinant in the sera of patients with nonmalignant disorders had the highest carbohydrate/protein ratio, followed by the mucin carrying the same determinant in the sera of cancer patients. Mucin antigen carrying 2----3 sialylated Lea antigen or 2----3, 2----6 disialylated Lc4 antigen in cancer patients had the lowest carbohydrate/protein ratio among the four groups tested. Thus, the carbohydrate/protein ratio in the type 1 chain mucin antigens in sera of normal subjects is higher than that in sera of cancer patients (P less than 0.05). This finding is in contrast to previous findings on the mucin antigens carrying the type 2 chain determinant (R. Kannagi

  4. Prostate specific antigen in the diagnosis and treatment of adenocarcinoma of the prostate. III. Radiation treated patients

    International Nuclear Information System (INIS)

    Stamey, T.A.; Kabalin, J.N.; Ferrari, M.

    1989-01-01

    Serum prostate specific antigen was determined (Yang polyclonal radioimmunoassay) in 183 men after radiation therapy for adenocarcinoma of the prostate. A total of 163 men had received 7,000 rad external beam radiotherapy and 20 had been implanted with iodine-125 seeds. Only 11 per cent of these 183 patients had undetectable prostate specific antigen levels at a mean interval of 5 years since completion of radiotherapy. Prostate specific antigen levels after radiotherapy were directly related to initial clinical stage and Gleason score before treatment. Multiple prostate specific antigen determinations were performed with time in 124 of 183 patients. During year 1 after radiotherapy prostate specific antigen levels were decreasing in 82 per cent of the patients but only 8 per cent continued to decrease beyond year 1. Of 80 patients observed greater than 1 year after completion of radiotherapy 51 per cent had increasing values and 41 per cent had stable values. Increasing prostate specific antigen values after radiotherapy were correlated with progression to metastastic disease and residual cancer on prostate biopsy. Total serum acid phosphatase levels were poorly related to prostate specific antigen levels, were less effective in discriminating patients with metastatic disease and provided no additional information beyond that provided by prostate specific antigen

  5. Memory control by the B cell antigen receptor.

    Science.gov (United States)

    Engels, Niklas; Wienands, Jürgen

    2018-05-01

    The generation of memory B cells (MBCs) that have undergone immunoglobulin class switching from IgM, which dominates primary antibody responses, to other immunoglobulin isoforms is a hallmark of immune memory. Hence, humoral immunological memory is characterized by the presence of serum immunoglobulins of IgG subtypes known as the γ-globulin fraction of blood plasma proteins. These antibodies reflect the antigen experience of B lymphocytes and their repeated triggering. In fact, efficient protection against a previously encountered pathogen is critically linked to the production of pathogen-specific IgG molecules even in those cases where the primary immune response required cellular immunity, for example, T cell-mediated clearance of intracellular pathogens such as viruses. Besides IgG, also IgA and IgE can provide humoral immunity depending on the microbe's nature and infection route. The molecular mechanisms underlying the preponderance of switched immunoglobulin isotypes during memory antibody responses are a matter of active and controversial debate. Here, we summarize the phenotypic characteristics of distinct MBC subpopulations and discuss the decisive roles of different B cell antigen receptor isotypes for the functional traits of class-switched B cell populations. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. Oncogenic cancer/testis antigens

    DEFF Research Database (Denmark)

    Gjerstorff, Morten F; Andersen, Mads H; Ditzel, Henrik J

    2015-01-01

    Recent developments have set the stage for immunotherapy as a supplement to conventional cancer treatment. Consequently, a significant effort is required to further improve efficacy and specificity, particularly the identification of optimal therapeutic targets for clinical testing. Cancer....../testis antigens are immunogenic, highly cancer-specific, and frequently expressed in various types of cancer, which make them promising candidate targets for cancer immunotherapy, including cancer vaccination and adoptive T-cell transfer with chimeric T-cell receptors. Our current understanding of tumor...... immunology and immune escape suggests that targeting oncogenic antigens may be beneficial, meaning that identification of cancer/testis antigens with oncogenic properties is of high priority. Recent work from our lab and others provide evidence that many cancer/testis antigens, in fact, have oncogenic...

  7. MERS-CoV and H5N1 influenza virus antagonize antigen presentation by altering the epigenetic landscape

    Energy Technology Data Exchange (ETDEWEB)

    Menachery, Vineet D.; Schafer, Alexandra; Burnum-Johnson, Kristin E.; Mitchell, Hugh D.; Eisfeld-Fenney, Amie J.; Walters, Kevin B.; Nicora, Carrie D.; Purvine, Samuel O.; Casey, Cameron P.; Monroe, Matthew E.; Weitz, Karl K.; Stratton, Kelly G.; Webb-Robertson, Bobbie-Jo M.; Gralinski, Lisa; Metz, Thomas O.; Smith, Richard D.; Waters, Katrina M.; Sims, Amy C.; Kawaoka, Yoshihiro; Baric, Ralph

    2018-01-16

    Convergent evolution dictates that diverse groups of viruses will target both similar and distinct host pathways in order to manipulate the immune response and improve infection. In this study, we sought to leverage this uneven viral antagonism to identify critical host factors that govern disease outcome. Utilizing a systems based approach, we examined differential regulation of IFNγ dependent genes following infection with highly pathogenic viruses including influenza (H5N1-VN1203, H1N1-CA04) and coronaviruses (SARS-CoV, MERS-CoV). Categorizing by function, we observed down regulation of genes associated with antigen presentation following both H5N1-VN1203 and MERS-CoV infection. Further examination revealed global down regulation of antigen presentation genes and was confirmed by proteomics for both H5N1-VN1203 and MERS-CoV infection. Importantly, epigenetic analysis suggested that DNA methylation rather than histone modification plays a crucial role in MERS-CoV mediated antagonism of antigen presentation genes; in contrast, H5N1-VN1203 likely utilizes a combination of epigenetic mechanisms to target antigen presentation. Together, the results indicate a common approach utilized by H5N1-VN1203 and MERS-CoV to modulate antigen presentation and the host adaptive immune response.

  8. Radioimmunoassays of hidden viral antigens

    International Nuclear Information System (INIS)

    Neurath, A.R.; Strick, N.; Baker, L.; Krugman, S.

    1982-01-01

    Antigens corresponding to infectious agents may be present in biological specimens only in a cryptic form bound to antibodies and, thus, may elude detection. We describe a solid-phase technique for separation of antigens from antibodies. Immune complexes are precipitated from serum by polyethylene glycol, dissociated with NaSCN, and adsorbed onto nitrocellulose or polystyrene supports. Antigens remain topographically separated from antibodies after removal of NaSCN and can be detected with radiolabeled antibodies. Genomes from viruses immobilized on nitrocellulose can be identified by nucleic acid hybridization. Nanogram quantities of sequestered hepatitis B surface and core antigens and picogram amounts of hepatitis B virus DNA were detected. Antibody-bound adenovirus, herpesvirus, and measles virus antigens were discerned by the procedure

  9. Antigen uptake and expression of antigen presentation-related immune genes in flounder (Paralichthys olivaceus) after vaccination with an inactivated Edwardsiella tarda immersion vaccine, following hyperosmotic treatment.

    Science.gov (United States)

    Gao, Yingli; Tang, Xiaoqian; Sheng, Xiuzhen; Xing, Jing; Zhan, Wenbin

    2016-08-01

    Antigen uptake is a critical process for activation of the immune system, and therefore the ability to enhance antigen uptake is a primary consideration in the development of an immersion vaccination of fish. In the present work, flounders (Paralichthys olivaceus) were immersed in three hyperosmotic solutions with 40, 50 and 60‰ salinities, then transferred into seawater of normal salinity (i.e. 30‰) containing formalin-inactivated Edwardsiella tarda for 30 min. The antigen uptake in vaccinated flounder was determined using an absolute quantitative PCR (qPCR). The results showed significantly higher antigen uptake in the tissues of flounders immersed in solutions with 50‰ and 60‰ salinity compared to the control group directly immersed in vaccine (DI) (P immersed in the 50‰ salinity solution, whereas there was no significant difference in antigen uptake between the 40‰ salinity group and the DI group (P > 0.05). A rapid and significant increase in antigen uptake was detected in the mucosal-associated tissues including the gill, skin and intestine (P immersion, which was significantly higher than the levels of uptake measured in the other tissues (P immersion (hpi). The expression profiles of four antigen presentation-related immune genes (MHC Iα, MHC IIα, CD4-1 and CD8α) were investigated after immersion. These four genes showed a significantly stronger response in the immersed flounders exposed to 50‰ salinity compared with the DI group (P immersion, notably 50‰ salinity significantly enhanced antigen uptake and the expression of selected genes associated with antigen presentation, providing evidence for an enhanced immune activation of the fish's immune response by the hyperosmotic immersion treatment prior to vaccination. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Foetal antigen 2 (FA2) in the stromal reaction induced by breast carcinoma

    DEFF Research Database (Denmark)

    Rasmussen, H B; Teisner, B; Andersen, J A

    1992-01-01

    An indirect immunoperoxidase technique was used to examine the distribution of foetal antigen 2 (FA2), a recently described basement membrane (BM)-associated antigen, in invasive breast carcinoma (n = 34), fibroadenoma (n = 5) and normal breast tissue (n = 5), and to compare its distribution...... with that of laminin and collagen type IV. In normal breast tissue, FA2 was detected in the intralobular stroma as a broad band around acini and ducts, but was not present in the interlobular stroma. In areas of carcinoma in situ, FA2 was present diffusely around and in close contact with the glandular elements......, the staining being more intense than that found around normal glandular structures. Two distinct patterns of FA2 distribution were found in adenocarcinomas of the breast. In the fibroblast reaction type, fibroblast staining dominated, whilst in the stromal reaction type, intense and extensive staining...

  11. Leukemia-associated antigens in man.

    Science.gov (United States)

    Brown, G; Capellaro, D; Greaves, M

    1975-12-01

    Rabbit antisera raised against acute lymphoblastic leukemia (ALL) cells were used to distinguish ALL from other leukemias, to identify rare leukemia cells in the bone marrow of patients in remission, and to define human leukemia-associated antigens. Antibody binding was studied with the use of immunofluorescence reagents and the analytic capacity of the Fluorescence Activated Cell Sorter-1 (FACS-1). The results indicated that most non-T-cell ALL have three leukemia-associated antigens on their surface which are absent from normal lymphoid cells: 1) an antigen shared with myelocytes, myeloblastic leukemia cells, and fetal liver (hematopoietic) cells; 2) an antigen shared with a subset of intermediate normoblasts in normal bone marrow and fetal liver; and 3) an antigen found thus far only on non-T-cell ALL and in some acute undifferentiated leukemias, which we therefore regard as a strong candidate for a leukemia-specific antigen. These antigens are absent from a subgroup of ALL patients in which the lymphoblasta express T-cell surface markers. Preliminary studies on the bone marrow samples of patients in remission indicated that rare leukemia cells were present in some samples. The implications of these findings with respect to the heterogeneity and cell origin(s) of ALL, its diagnosis, and its potential monitoring during treatment were discussed.

  12. Anvendelse af prostataspecifikt antigen. En oversigt

    DEFF Research Database (Denmark)

    Brasso, K; Skaarup, P; Roosen, Jens Ulrik

    1998-01-01

    Since it was first introduced, measurement of prostate specific antigen has gained increasing interest, and prostate specific antigen is regarded as being the best tumour marker available. The antigen lacks cancer specificity, limiting the usefulness in early diagnosis, The use of prostate specific...... antigen in early diagnosis, staging, and in monitoring patients with prostate cancer is reviewed....

  13. Purification and properties of extracellular polysaccharide (EPS) antigens produced by different mould species.

    Science.gov (United States)

    Notermans, S; Wieten, G; Engel, H W; Rombouts, F M; Hoogerhout, P; van Boom, J H

    1987-02-01

    Extracellular polysaccharide (EPS) antigens produced by different mould species were purified and partially characterized. Purification included (NH4)2SO4 treatment, Sepharose CL-4B column chromatography and Con A-sepharose chromatography. The EPS of Penicillium digitatum, Mucor racemosus and Cladosporium cladosporioides showed high antigenic capacities. Immunologically the EPS were partially genus-specific, but cross-reactivity was observed. The EPS antigens produced by species of Penicillium, Aspergillus repens and Geotrichum candidum lost their immunological activity upon heating (100 degrees C) at pH 1.8, while the EPS antigen of M. racemosus, Rhizopus oligosporus and C. cladosporioides were stable under the same conditions. The dominant monosaccharides present in the EPS antigen were mannose, galactose and glucose. The EPS obtained from cultures of M. racemosus and R. oligosporus also contained rhamnose. In the EPS produced by Penicillium spp. and A. repens the galactose residues were determined to be immunodominant.

  14. The molecular bases of δ/αβ T cell-mediated antigen recognition.

    Science.gov (United States)

    Pellicci, Daniel G; Uldrich, Adam P; Le Nours, Jérôme; Ross, Fiona; Chabrol, Eric; Eckle, Sidonia B G; de Boer, Renate; Lim, Ricky T; McPherson, Kirsty; Besra, Gurdyal; Howell, Amy R; Moretta, Lorenzo; McCluskey, James; Heemskerk, Mirjam H M; Gras, Stephanie; Rossjohn, Jamie; Godfrey, Dale I

    2014-12-15

    αβ and γδ T cells are disparate T cell lineages that can respond to distinct antigens (Ags) via the use of the αβ and γδ T cell Ag receptors (TCRs), respectively. Here we characterize a population of human T cells, which we term δ/αβ T cells, expressing TCRs comprised of a TCR-δ variable gene (Vδ1) fused to joining α and constant α domains, paired with an array of TCR-β chains. We demonstrate that these cells, which represent ∼50% of all Vδ1(+) human T cells, can recognize peptide- and lipid-based Ags presented by human leukocyte antigen (HLA) and CD1d, respectively. Similar to type I natural killer T (NKT) cells, CD1d-lipid Ag-reactive δ/αβ T cells recognized α-galactosylceramide (α-GalCer); however, their fine specificity for other lipid Ags presented by CD1d, such as α-glucosylceramide, was distinct from type I NKT cells. Thus, δ/αβTCRs contribute new patterns of Ag specificity to the human immune system. Furthermore, we provide the molecular bases of how δ/αβTCRs bind to their targets, with the Vδ1-encoded region providing a major contribution to δ/αβTCR binding. Our findings highlight how components from αβ and γδTCR gene loci can recombine to confer Ag specificity, thus expanding our understanding of T cell biology and TCR diversity. © 2014 Pellicci et al.

  15. The molecular bases of δ/αβ T cell–mediated antigen recognition

    Science.gov (United States)

    Pellicci, Daniel G.; Uldrich, Adam P.; Le Nours, Jérôme; Ross, Fiona; Chabrol, Eric; Eckle, Sidonia B.G.; de Boer, Renate; Lim, Ricky T.; McPherson, Kirsty; Besra, Gurdyal; Howell, Amy R.; Moretta, Lorenzo; McCluskey, James; Heemskerk, Mirjam H.M.; Gras, Stephanie

    2014-01-01

    αβ and γδ T cells are disparate T cell lineages that can respond to distinct antigens (Ags) via the use of the αβ and γδ T cell Ag receptors (TCRs), respectively. Here we characterize a population of human T cells, which we term δ/αβ T cells, expressing TCRs comprised of a TCR-δ variable gene (Vδ1) fused to joining α and constant α domains, paired with an array of TCR-β chains. We demonstrate that these cells, which represent ∼50% of all Vδ1+ human T cells, can recognize peptide- and lipid-based Ags presented by human leukocyte antigen (HLA) and CD1d, respectively. Similar to type I natural killer T (NKT) cells, CD1d-lipid Ag-reactive δ/αβ T cells recognized α-galactosylceramide (α-GalCer); however, their fine specificity for other lipid Ags presented by CD1d, such as α-glucosylceramide, was distinct from type I NKT cells. Thus, δ/αβTCRs contribute new patterns of Ag specificity to the human immune system. Furthermore, we provide the molecular bases of how δ/αβTCRs bind to their targets, with the Vδ1-encoded region providing a major contribution to δ/αβTCR binding. Our findings highlight how components from αβ and γδTCR gene loci can recombine to confer Ag specificity, thus expanding our understanding of T cell biology and TCR diversity. PMID:25452463

  16. Comparative analysis of rabbit hemorrhagic disease virus (RHDV) and new RHDV2 virus antigenicity, using specific virus-like particles.

    Science.gov (United States)

    Bárcena, Juan; Guerra, Beatriz; Angulo, Iván; González, Julia; Valcárcel, Félix; Mata, Carlos P; Castón, José R; Blanco, Esther; Alejo, Alí

    2015-09-24

    In 2010 a new Lagovirus related to rabbit haemorrhagic disease virus (RHDV) emerged in France and has since rapidly spread throughout domestic and wild rabbit populations of several European countries. The new virus, termed RHDV2, exhibits distinctive genetic, antigenic and pathogenic features. Notably, RHDV2 kills rabbits previously vaccinated with RHDV vaccines. Here we report for the first time the generation and characterization of RHDV2-specific virus-like particles (VLPs). Our results further confirmed the differential antigenic properties exhibited by RHDV and RHDV2, highlighting the need of using RHDV2-specific diagnostic assays to monitor the spread of this new virus.

  17. A polyvalent vaccine for high-risk prostate patients: "are more antigens better?"

    DEFF Research Database (Denmark)

    Slovin, Susan F; Ragupathi, Govind; Fernandez, Celina

    2007-01-01

    vaccine of synthetic "self" antigens broke immunologic tolerance against two or more antigens in all 30 vaccinated patients, was safe, but antibody titers against several of the antigens were lower than those seen in individual monovalent trials. No impact on PSA slope was detected. We address......We have shown the immunogenicity and safety of synthetic carbohydrate vaccines when conjugated to the carrier keyhole limpet hemocyanin (KLH) and given with the adjuvant, QS-21, in patients with biochemically relapsed prostate cancer. To determine whether immune response could be further enhanced...... and mixed with QS-21. Eight vaccinations were administered over 13 months. All 30 patients had significant elevations in antibody titers to at least two of the six antigens; 22 patients had increased reactivity with FACS. These serologic responses were lower than that seen previously in patients treated...

  18. Epstein-Barr virus early antigen diffuse (EBV-EA/D)-directed immunoglobulin A antibodies in systemic lupus erythematosus patients

    DEFF Research Database (Denmark)

    Draborg, A H; Jørgensen, J M; Müller, H

    2012-01-01

    We sought to determine whether the serological response towards lytic cycle antigens of Epstein-Barr virus (EBV) is altered in systemic lupus erythematosus (SLE) patients.......We sought to determine whether the serological response towards lytic cycle antigens of Epstein-Barr virus (EBV) is altered in systemic lupus erythematosus (SLE) patients....

  19. Comparison of antigen-specific T-cell responses of tuberculosis patients using complex or single antigens of Mycobacterium tuberculosis

    DEFF Research Database (Denmark)

    Mustafa, A S; Amoudy, H A; Wiker, H G

    1998-01-01

    GroES, rPstS, rGroEL and rDnaK) antigens of Mycobacterium tuberculosis. The responses of PBMC to these defined antigens were compared with the corresponding results obtained with complex antigens, such as whole-cell M. tuberculosis, M. tuberculosis culture filtrate (MT-CF) and cell wall antigens, as well...... as the vaccine strain, Mycobacterium bovis bacillus Calmette-Guerin (BCG). In addition, M. tuberculosis and MT-CF-induced T-cell lines were tested in the same assays against the panel of purified and complex antigens. The compiled data from PBMC and T-cell lines tested for antigen-induced proliferation and IFN...

  20. Identification of antigenic Sarcoptes scabiei proteins for use in a diagnostic test and of non-antigenic proteins that may be immunomodulatory.

    Directory of Open Access Journals (Sweden)

    Marjorie S Morgan

    2017-06-01

    Full Text Available Scabies, caused by the mite, Sarcoptes scabiei, infects millions of humans, and many wild and domestic mammals. Scabies mites burrow in the lower stratum corneum of the epidermis of the skin and are the source of substances that are antigenic or modulate aspects of the protective response of the host. Ordinary scabies is a difficult disease to diagnose.The goal of this project was to identify S. scabiei proteins that may be candidate antigens for use in a diagnostic test or may be used by the mite to modulate the host's protective response.An aqueous extract of S. scabiei was separated by 2-dimensional electrophoresis and proteins were identified by mass spectrometry. A parallel immunoblot was probed with serum from patients with ordinary scabies to identify IgM and/or IgG-binding antigens. The genes coding for 23 selected proteins were cloned into E. coli and the expressed recombinant proteins were screened with serum from patients with confirmed ordinary scabies.We identified 50 different proteins produced by S. scabiei, 34 of which were not previously identified, and determined that 66% were recognized by patient IgM and/or IgG. Fourteen proteins were screened for use in a diagnostic test but none possessed enough sensitivity and specificity to be useful. Six of the 9 proteins selected for the possibility that they may be immunomodulatory were not recognized by antibodies in patient serum.Thirty-three proteins that bound IgM and/or IgG from the serum of patients with ordinary scabies were identified. None of the 14 tested were useful for inclusion in a diagnostic test. The identities of 16 proteins that are not recognized as antigens by infected patients were also determined. These could be among the molecules that are responsible for this mite's ability to modulate its host's innate and adaptive immune responses.

  1. The role of CD4 in antigen-independent activation of isolated single T lymphocytes

    DEFF Research Database (Denmark)

    Kelso, A; Owens, T

    1988-01-01

    The membrane molecule CD4 (L3T4) is thought to facilitate activation of Class II H-2-restricted T cells by binding to Ia determinants on antigen-presenting cells. Recent reports suggest that CD4 can also contribute to antigen-independent activation by anti-T cell receptor (TCR) antibodies. An ass...

  2. Identification of toxocara canis antigens by Western blot in experimentally infected rabbits

    Directory of Open Access Journals (Sweden)

    MORALES Olga Lucía

    2002-01-01

    Full Text Available Toxocariasis is a frequent helminthiasis that can cause visceral and ocular damage in humans specially in children. The identification of specific antigens of Toxocara canis is important in order to develop better diagnostic techniques. Ten rabbits were infected orally with a dose of 5000 Toxocara canis embryonated eggs. Rabbits were bled periodically and an ELISA assay was performed to determine levels of specific Toxocara IgG antibodies. ELISA detected antibodies at day 15 after infection. Western blot (WB assay was performed using excretory/secretory antigens (E/S of T. canis second stage larvae. Different antigen concentrations were evaluated: 150, 200, 250 and 300 µg/mL. The concentration of 250 µg/mL was retained for analysis. Rabbit sera were diluted 1:100. Secondary antibody was used at a dilution of 1:1000. Results of WB indicated that in the first month after infection specific antibodies against the 200 KDa, 116 KDa, 92 KDa and 35 KDa antigens were detected; antibodies against the 92 KDa, 80 KDa, 66 KDa, 45 KDa, 31 KDa and 28 KDa antigens appeared later. All positive sera in the ELISA test were also positive in WB. Two antigen bands, 92 KDa and 35 KDa, were identified since the beginning and throughout the course of infection. These antigens merit further evaluation as candidates for use in diagnosis.

  3. Antibody Responses to Prostate-Associated Antigens in Patients with Prostatitis and Prostate Cancer

    Science.gov (United States)

    Maricque, Brett B.; Eickhoff, Jens C.; McNeel, Douglas G.

    2010-01-01

    Background An important focus of tumor immunotherapy has been the identification of appropriate antigenic targets. Serum-based screening approaches have led to the discovery of hundreds of tumor-associated antigens recognized by IgG. Our efforts to identify immunologically recognized proteins in prostate cancer have yielded a multitude of antigens, however prioritizing these antigens as targets for evaluation in immunotherapies has been challenging. In this report, we set out to determine whether the evaluation of multiple antigenic targets would allow the identification of a subset of antigens that are common immunologic targets in patients with prostate cancer. Methods Using a phage immunoblot approach, we evaluated IgG responses in patients with prostate cancer (n=126), patients with chronic prostatitis (n=45), and men without prostate disease (n=53). Results We found that patients with prostate cancer or prostatitis have IgG specific for multiple common antigens. A subset of 23 proteins was identified to which IgG were detected in 38% of patients with prostate cancer and 33% patients with prostatitis versus 6% of controls (pprostate and prostate cancer, and suggest that IgG responses to a panel of commonly recognized prostate antigens could be potentially used in the identification of patients at risk for prostate cancer or as a tool to identify immune responses elicited to prostate tissue. PMID:20632317

  4. Specific binding of antigen-antibody in physiological environments: Measurement, force characteristics and analysis

    Science.gov (United States)

    Gu, Xin; Zhou, Jun; Zhou, Lu; Xie, Shusen; Petti, Lucia; Wang, Shaomin; Wang, Fuyan

    2018-05-01

    The specific recognition of the antigen by the antibody is the crucial step in immunoassays. Measurement and analysis of the specific recognition, including the ways in which it is influenced by external factors are of paramount significance for the quality of the immunoassays. Using prostate-specific antigen (PSA)/anti-PSA antibody and α-fetoprotein (AFP) /anti-AFP antibody as examples, we have proposed a novel solution for measuring the binding forces between the antigens and their corresponding antibodies in different physiological environments by combining laminar flow control technology and optical tweezers technology. On the basis of the experimental results, the different binding forces of PSA/anti-PSA antibody and AFP/anti-AFP antibody in the same phosphate-buffered saline (PBS) environments are analysed by comparing the affinity constant of the two antibodies and the number of antigenic determinants of the two antigens. In different electrolyte environments, the changes of the binding force of antigens-antibodies are explained by the polyelectrolyte effect and hydrophobic interaction. Furthermore, in different pH environments, the changes of binding forces of antigens-antibodies are attributed to the role of the denaturation of protein. The study aims to recognise the antigen-antibody immune mechanism, thus ensuring further understanding of the biological functions of tumour markers, and it promises to be very useful for the clinical diagnosis of early-stage cancer.

  5. Antigenic and genetic evolution of contemporary swine H1 influenza viruses in the United States.

    Science.gov (United States)

    Rajao, Daniela S; Anderson, Tavis K; Kitikoon, Pravina; Stratton, Jered; Lewis, Nicola S; Vincent, Amy L

    2018-05-01

    Several lineages of influenza A viruses (IAV) currently circulate in North American pigs. Genetic diversity is further increased by transmission of IAV between swine and humans and subsequent evolution. Here, we characterized the genetic and antigenic evolution of contemporary swine H1N1 and H1N2 viruses representing clusters H1-α (1A.1), H1-β (1A.2), H1pdm (1A.3.3.2), H1-γ (1A.3.3.3), H1-δ1 (1B.2.2), and H1-δ2 (1B.2.1) currently circulating in pigs in the United States. The δ1-viruses diversified into two new genetic clades, H1-δ1a (1B.2.2.1) and H1-δ1b (1B.2.2.2), which were also antigenically distinct from the earlier H1-δ1-viruses. Further characterization revealed that a few key amino acid changes were associated with antigenic divergence in these groups. The continued genetic and antigenic evolution of contemporary H1 viruses might lead to loss of vaccine cross-protection that could lead to significant economic impact to the swine industry, and represents a challenge to public health initiatives that attempt to minimize swine-to-human IAV transmission. Published by Elsevier Inc.

  6. Analysis of a cDNA clone expressing a human autoimmune antigen: full-length sequence of the U2 small nuclear RNA-associated B antigen

    International Nuclear Information System (INIS)

    Habets, W.J.; Sillekens, P.T.G.; Hoet, M.H.; Schalken, J.A.; Roebroek, A.J.M.; Leunissen, J.A.M.; Van de Ven, W.J.M.; Van Venrooij, W.J.

    1987-01-01

    A U2 small nuclear RNA-associated protein, designated B'', was recently identified as the target antigen for autoimmune sera from certain patients with systemic lupus erythematosus and other rheumatic diseases. Such antibodies enabled them to isolate cDNA clone λHB''-1 from a phage λgt11 expression library. This clone appeared to code for the B'' protein as established by in vitro translation of hybrid-selected mRNA. The identity of clone λHB''-1 was further confirmed by partial peptide mapping and analysis of the reactivity of the recombinant antigen with monospecific and monoclonal antibodies. Analysis of the nucleotide sequence of the 1015-base-pair cDNA insert of clone λHB''-1 revealed a large open reading frame of 800 nucleotides containing the coding sequence for a polypeptide of 25,457 daltons. In vitro transcription of the λHB''-1 cDNA insert and subsequent translation resulted in a protein product with the molecular size of the B'' protein. These data demonstrate that clone λHB''-1 contains the complete coding sequence of this antigen. The deduced polypeptide sequence contains three very hydrophilic regions that might constitute RNA binding sites and/or antigenic determinants. These findings might have implications both for the understanding of the pathogenesis of rheumatic diseases as well as for the elucidation of the biological function of autoimmune antigens

  7. Expression of endogenous ALV antigens and susceptibility to subgroup E ALV in three strains of chickens (Endogenous avian C. type virus)

    International Nuclear Information System (INIS)

    Robinson, H.L.; Lamoreux, W.F.

    1976-01-01

    Cells from three strains of Kimber Farms chickens, K16, K18, and K28, have been characterized for the expression of endogenous avian C-type virus (ALV) antigens and for susceptibility to subgroup E ALV. In K16 the coordinate dominant expression of chick helper factor (chf ), the type specific antigen of endogenous subgroup E ALV, and the group specific (gs) antigen of ALV was observed. The expression of chf and gs antigen in K16 chf + gs + cells was similar to that observed in SPAFAS and H and N gs + cells. In K18 chf but not gs antigen was expressed. The expression of chf in K18 chf + gs + cells was distinct from that observed for the chf + gs + pedigrees but similar to that found in SPAFAS chf + gs - helper extremely high (h/sub E/) cells. Cells from K16 and K18 chickens were uniformly resistant to subgroup E ALV. In cells from K28 chickens, susceptibility to subgroup B virus correlated with susceptibility to subgroup E virus. The efficiency of plating of subgroup E virus on susceptible K28 cells with 10 3 --10 4 -fold lower on chf + cells than on chf - cells

  8. Characterization of Antigen-Specific B Cells Using Nominal Antigen-Coated Flow-Beads

    Science.gov (United States)

    Akl, Ahmed; Lepetit, Maud; Crochette, Romain; Giral, Magali; Lepourry, Julie; Pallier, Annaick; Castagnet, Stéphanie; Dugast, Emilie; Guillot-Gueguen, Cécile; Jacq-Foucher, Marylène; Saulquin, Xavier; Cesbron, Anne; Laplaud, David; Nicot, Arnaud; Brouard, Sophie; Soulillou, Jean-Paul

    2013-01-01

    In order to characterize the reactivity of B cells against nominal antigens, a method based on the coupling of antigens onto the surface of fluorescent core polystyrene beads was developed. We first demonstrate that murine B cells with a human MOG-specific BCR are able to interact with MOG-coated beads and do not recognize beads coated with human albumin or pp65. B cells purified from human healthy volunteer blood or immunized individuals were tested for their ability to interact with various nominal antigens, including viral, vaccine, self and alloantigens, chosen for their usefulness in studying a variety of pathological processes. A substantial amount of B cells binding self-antigen MOG-coated beads can be detected in normal blood. Furthermore, greater frequencies of B cell against anti-Tetanic Toxin or anti-EBNA1 were observed in primed individuals. This method can reveal increased frequencies of anti-HLA committed B cells in patients with circulating anti-HLA antibodies compared to unsensitized patients and normal individuals. Of interest, those specific CD19 cells were preferentially identified within CD27−IgD+ (i-e naïve) subset. These observations suggest that a broad range of medical situations could benefit from a tool that allows the detection, the quantification and the characterization of antigen-specific blood B cells. PMID:24386360

  9. Making antigen of trypanosoma evansi and its examination by catt (card agglutination test) method

    International Nuclear Information System (INIS)

    Arifin, M; Irtisam; Estikoma Dyah; Yulia Ernawati; Boky, J.T

    1998-01-01

    An experiment was carried out for making antigen (Ag) of T. evansi by using experimental animals rats and guinea pigs for for developing parasites, and in cattle for making immune serum and normal serum. The parasites were irradiated by gamma rays ( 60 Co) at a dose of 300 Gy. The determination of antigen was done by using Card Agglutination Test (CATT) to the field samples serum of cattle. The results obtained showed that antigen was good enough potently and stable at minus 70 0 C storage during five months. (author)

  10. MicroRNA transfection and AGO-bound CLIP-seq data sets reveal distinct determinants of miRNA action

    DEFF Research Database (Denmark)

    Wen, Jiayu; Parker, Brian J; Jacobsen, Anders

    2011-01-01

    the predictive effect of target flanking features. We observe distinct target determinants between expression-based and CLIP-based data. Target flanking features such as flanking region conservation are an important AGO-binding determinant-we hypothesize that CLIP experiments have a preference for strongly bound......Microarray expression analyses following miRNA transfection/inhibition and, more recently, Argonaute cross-linked immunoprecipitation (CLIP)-seq assays have been used to detect miRNA target sites. CLIP and expression approaches measure differing stages of miRNA functioning-initial binding of the mi...... miRNP-target interactions involving adjacent RNA-binding proteins that increase the strength of cross-linking. In contrast, seed-related features are major determinants in expression-based studies, but less so for CLIP-seq studies, and increased miRNA concentrations typical of transfection studies...

  11. Facts on the fragmentation of antigens in presenting cells, on the association of antigen fragments with MHC molecules in cell-free systems, and speculation on the cell biology of antigen processing

    DEFF Research Database (Denmark)

    Werdelin, O; Mouritsen, S; Petersen, B L

    1988-01-01

    The processing of a protein antigen is a multi-step event taking place in antigen-presenting cells. Processing is a prerequisite for the recognition of most antigens by T lymphocytes. The antigen is ingested by endocytosis, transported to an acid cellular compartment and subjected to proteolytic...... fragmentation. Some of the antigen fragments bind to MHC class II molecules and are transported to the surface of the antigen-presenting cell where the actual presentation to T lymphocytes occurs. The nature of the processed antigen, how and where it is derived and subsequently becomes associated with MHC...... molecules are the questions discussed in this review. To us, the entire concept of processing has appeal not only because it explains some hitherto well-established, but poorly understood, phenomena such as the fact that T lymphocytes focus their attention entirely upon antigens on other cells. It has...

  12. Natural micropolymorphism in human leukocyte antigens provides a basis for genetic control of antigen recognition

    Energy Technology Data Exchange (ETDEWEB)

    Archbold, Julia K.; Macdonald, Whitney A.; Gras, Stephanie; Ely, Lauren K.; Miles, John J.; Bell, Melissa J.; Brennan, Rebekah M.; Beddoe, Travis; Wilce, Matthew C.J.; Clements, Craig S.; Purcell, Anthony W.; McCluskey, James; Burrows, Scott R.; Rossjohn, Jamie; (Monash); (Queensland Inst. of Med. Rsrch.); (Melbourne)

    2009-07-10

    Human leukocyte antigen (HLA) gene polymorphism plays a critical role in protective immunity, disease susceptibility, autoimmunity, and drug hypersensitivity, yet the basis of how HLA polymorphism influences T cell receptor (TCR) recognition is unclear. We examined how a natural micropolymorphism in HLA-B44, an important and large HLA allelic family, affected antigen recognition. T cell-mediated immunity to an Epstein-Barr virus determinant (EENLLDFVRF) is enhanced when HLA-B*4405 was the presenting allotype compared with HLA-B*4402 or HLA-B*4403, each of which differ by just one amino acid. The micropolymorphism in these HLA-B44 allotypes altered the mode of binding and dynamics of the bound viral epitope. The structure of the TCR-HLA-B*4405EENLLDFVRF complex revealed that peptide flexibility was a critical parameter in enabling preferential engagement with HLA-B*4405 in comparison to HLA-B*4402/03. Accordingly, major histocompatibility complex (MHC) polymorphism can alter the dynamics of the peptide-MHC landscape, resulting in fine-tuning of T cell responses between closely related allotypes.

  13. Natural selection promotes antigenic evolvability.

    Science.gov (United States)

    Graves, Christopher J; Ros, Vera I D; Stevenson, Brian; Sniegowski, Paul D; Brisson, Dustin

    2013-01-01

    The hypothesis that evolvability - the capacity to evolve by natural selection - is itself the object of natural selection is highly intriguing but remains controversial due in large part to a paucity of direct experimental evidence. The antigenic variation mechanisms of microbial pathogens provide an experimentally tractable system to test whether natural selection has favored mechanisms that increase evolvability. Many antigenic variation systems consist of paralogous unexpressed 'cassettes' that recombine into an expression site to rapidly alter the expressed protein. Importantly, the magnitude of antigenic change is a function of the genetic diversity among the unexpressed cassettes. Thus, evidence that selection favors among-cassette diversity is direct evidence that natural selection promotes antigenic evolvability. We used the Lyme disease bacterium, Borrelia burgdorferi, as a model to test the prediction that natural selection favors amino acid diversity among unexpressed vls cassettes and thereby promotes evolvability in a primary surface antigen, VlsE. The hypothesis that diversity among vls cassettes is favored by natural selection was supported in each B. burgdorferi strain analyzed using both classical (dN/dS ratios) and Bayesian population genetic analyses of genetic sequence data. This hypothesis was also supported by the conservation of highly mutable tandem-repeat structures across B. burgdorferi strains despite a near complete absence of sequence conservation. Diversification among vls cassettes due to natural selection and mutable repeat structures promotes long-term antigenic evolvability of VlsE. These findings provide a direct demonstration that molecular mechanisms that enhance evolvability of surface antigens are an evolutionary adaptation. The molecular evolutionary processes identified here can serve as a model for the evolution of antigenic evolvability in many pathogens which utilize similar strategies to establish chronic infections.

  14. Natural selection promotes antigenic evolvability.

    Directory of Open Access Journals (Sweden)

    Christopher J Graves

    Full Text Available The hypothesis that evolvability - the capacity to evolve by natural selection - is itself the object of natural selection is highly intriguing but remains controversial due in large part to a paucity of direct experimental evidence. The antigenic variation mechanisms of microbial pathogens provide an experimentally tractable system to test whether natural selection has favored mechanisms that increase evolvability. Many antigenic variation systems consist of paralogous unexpressed 'cassettes' that recombine into an expression site to rapidly alter the expressed protein. Importantly, the magnitude of antigenic change is a function of the genetic diversity among the unexpressed cassettes. Thus, evidence that selection favors among-cassette diversity is direct evidence that natural selection promotes antigenic evolvability. We used the Lyme disease bacterium, Borrelia burgdorferi, as a model to test the prediction that natural selection favors amino acid diversity among unexpressed vls cassettes and thereby promotes evolvability in a primary surface antigen, VlsE. The hypothesis that diversity among vls cassettes is favored by natural selection was supported in each B. burgdorferi strain analyzed using both classical (dN/dS ratios and Bayesian population genetic analyses of genetic sequence data. This hypothesis was also supported by the conservation of highly mutable tandem-repeat structures across B. burgdorferi strains despite a near complete absence of sequence conservation. Diversification among vls cassettes due to natural selection and mutable repeat structures promotes long-term antigenic evolvability of VlsE. These findings provide a direct demonstration that molecular mechanisms that enhance evolvability of surface antigens are an evolutionary adaptation. The molecular evolutionary processes identified here can serve as a model for the evolution of antigenic evolvability in many pathogens which utilize similar strategies to establish

  15. Concepts and applications for influenza antigenic cartography

    Science.gov (United States)

    Cai, Zhipeng; Zhang, Tong; Wan, Xiu-Feng

    2011-01-01

    Influenza antigenic cartography projects influenza antigens into a two or three dimensional map based on immunological datasets, such as hemagglutination inhibition and microneutralization assays. A robust antigenic cartography can facilitate influenza vaccine strain selection since the antigenic map can simplify data interpretation through intuitive antigenic map. However, antigenic cartography construction is not trivial due to the challenging features embedded in the immunological data, such as data incompleteness, high noises, and low reactors. To overcome these challenges, we developed a computational method, temporal Matrix Completion-Multidimensional Scaling (MC-MDS), by adapting the low rank MC concept from the movie recommendation system in Netflix and the MDS method from geographic cartography construction. The application on H3N2 and 2009 pandemic H1N1 influenza A viruses demonstrates that temporal MC-MDS is effective and efficient in constructing influenza antigenic cartography. The web sever is available at http://sysbio.cvm.msstate.edu/AntigenMap. PMID:21761589

  16. Prostate specific antigen, digital rectal examination, transrectal ultrasound: how accurate are they in determining prostate carcinoma?

    International Nuclear Information System (INIS)

    Gomez, John Anthony M.; Pagdanganan, Ernest Jerome A.; Caedo, Florencio Gerardo O.; Magsino, Benjamin C.; Rivera, Eduardo Ll.; Songco, Jaime S.D.

    1998-01-01

    Prostate cancer is an increasing problem. It is the most frequent malignancy in men past the age of 65 years. In the Philippines, 10-20% of males operated for prostatic obstruction had prostate cancer. The potential for cure is optimized by early detection and treatment of organ confined disease. Digital rectal examination, serum prostatic specific antigen and transrectal ultrasound of the prostate have been advocated individually and collectively to determine prostatic cancer. Our study involved forty-nine males who underwent all three screening modalities. Results of the study showed a statistically significant association between the presence of a nodule and occurrence of prostate cancer, a statistically significant association between hardness in consistency and cancer, a statistically significant difference in mean weight between those with Ca and BPH; a statistically significant difference in mean PSA levels between those with Ca and with BPH; statistically significant association between abnormal PSA levels and Ca; and a statistically significant association between a composite positive result and cancer. On the other hand, there was no statistically significant difference in mean age between those with cancer and those with BPH; there is no statistically significant association between the presence of prostatism and whether the patient has Ca or BPH; and there is no statistically significant difference in the mean duration between those with cancer and those with BPH. The study advocates the use of DRE, serum PSA in determining prostatic Ca as well as TRUS for determining occult carcinoma. (Author)

  17. Dog Erythrocyte Antigen 1 (DEA 1): Mode of Inheritance and Initial Characterization

    Science.gov (United States)

    Polak, Klaudia; Acierno, Michelle; Raj, Karthik; Mizukami, Keijiro; Siegel, Don L.; Giger, Urs

    2015-01-01

    Background The Dog Erythrocyte Antigen (DEA) 1 blood group system remains poorly defined. Objectives The purpose of the study was to determine the DEA 1 mode of inheritance and to characterize the DEA 1 antigen and alloantibodies. Animals Canine research colony families, clinic canine patients, and DEA 1.2+ blood bank dogs were studied. Methods Canine blood was typed by flow cytometry and immunochromatographic strips using anti-DEA 1 monoclonal antibodies. Gel column experiments with polyclonal and immunoblotting with monoclonal anti-DEA 1 antibodies were performed to analyze select samples. Cross-reactivity of human typing reagents against canine RBCs and one monoclonal anti-DEA 1 antibody against human RBC panels was assessed. Results Typing of 12 families comprising 144 dogs indicated an autosomal dominant inheritance with ≥4 alleles: DEA 1− (0) and DEA 1+ weak (1+), intermediate (2+) and strong (3+ and 4+). Samples from 6 dogs previously typed as DEA 1.2+ were typed as DEA 1+ or DEA 1− using monoclonal antibodies. Human typing reagents produced varied reactions in tube agglutination experiments against DEA 1+ and DEA 1− RBCs. Polypeptide bands were not detected on immunoblots using a monoclonal anti-DEA 1 antibody, therefore the anti-DEA 1 antibody may be specific for conformational epitopes lost during denaturation. Conclusions The autosomal dominant inheritance of DEA 1 with ≥4 alleles indicates a complex blood group system; the antigenicity of each DEA 1+ type will need to be determined. The biochemical nature of the DEA 1 antigen(s) appears different from human blood group systems tested. PMID:26291052

  18. Effects of Co60 gamma radiation on the immunogenic and antigenic properties of Bothrops jararacussu venom

    International Nuclear Information System (INIS)

    Spencer, Patrick J.; Nascimento, Nanci do; Rogero, Jose R.

    1997-01-01

    Ionizing radiation has been successfully employed to attenuate animals toxins and venoms for immunizing antisera producing animals. However, the radiation effects on antigenicity and immunogenecity have not yet been elucidated. In the present work, we investigated the effects of gamma rays on the antigenic and immunogenicity have not yet been elucidated. In the present work, we investigated the effects of gamma rays on the antigenic and immunogenic behaviour of Bothrops jararacussu venon. Venom samples (2mg/ml in 150 mM NaCl) were irradiated with 500, 1000 and 2000 Gy of 60 Co gamma rays. These samples were submitted to antigen capture ELISA on plates coated with commercial bothropic antiserum. Results suggest a loss of reactivity of the 1000 and 2000 Gy irradiated samples. Antibodies against native and 2000 Gy irradiated venoms were produced in rabbits. Both sera able to bind native venom with a slightly higher titer for anti-irradiated serum. These data suggest that radiation promoted structural modification on the antigen molecules. However since the antibodies produced against irradiated antivenom were able to recognize native venom, there must have been preservation of some antigenic determinants. It has already been demosntrated that irradiation of proteins leads to structural modifications and unfolding of the molecules. Our data suggest that irradiation led to conformational epitopes destruction with preservation of linear epitopes and that the response against irradiated venom may be attributed to these linear antigenic determinants. (author). 8 refs., 3 figs

  19. Duffy blood group antigens: structure, serological properties and function

    Directory of Open Access Journals (Sweden)

    Ewa Łukasik

    2016-03-01

    Full Text Available Duffy (Fy blood group antigens are located on seven-transmembrane glycoprotein expressed on erythrocytes and endothelial cells, which acts as atypical chemokine receptor (ACKR1 and malarial receptor. The biological role of the Duffy glycoprotein has not been explained yet. It is suggested that Duffy protein modulate the intensity of the inflammatory response. The Duffy blood group system consists of two major antigens, Fya and Fyb, encoded by two codominant alleles designated FY*A and FY*B which differ by a single nucleotide polymorphism (SNP at position 125G>A of the FY gene that results in Gly42Asp amino acid change in the Fya and Fyb antigens, respectively. The presence of antigen Fya and/or Fyb on the erythrocytes determine three Duffy-positive phenotypes: Fy(a+b-, Fy(a-b+ and Fy(a+b+, identified in Caucasian population. The Duffy-negative phenotype Fy(a-b-, frequent in Africans, but very rare in Caucasians, is defined by the homozygous state of FY*B-33 alleles. The FY*B-33 allele is associated with a SNP -33T>C in the promoter region of the FY gene, which suppresses erythroid expression of this gene without affecting its expression in other tissues. The FY*X allele, found in Caucasians, is correlated with weak expression of Fyb antigen. Fyx antigen differs from the native Fyb by the Arg89Cys and Ala100Thr amino acid substitutions due to SNPs: 265C>T and 298G>A in FY*B allele. The frequency of the FY alleles shows marked geographic disparities, the FY*B-33 allele is predominant in Africans, the FY*B in Caucasians, while the FY*A allele is dominant in Asians and it is the most prevalent allele globally.

  20. Quantification of HLA-DM-Dependent Major Histocompatibility Complex of Class II Immunopeptidomes by the Peptide Landscape Antigenic Epitope Alignment Utility

    Directory of Open Access Journals (Sweden)

    Miguel Álvaro-Benito

    2018-05-01

    Full Text Available The major histocompatibility complex of class II (MHCII immunopeptidome represents the repertoire of antigenic peptides with the potential to activate CD4+ T cells. An understanding of how the relative abundance of specific antigenic epitopes affects the outcome of T cell responses is an important aspect of adaptive immunity and offers a venue to more rationally tailor T cell activation in the context of disease. Recent advances in mass spectrometric instrumentation, computational power, labeling strategies, and software analysis have enabled an increasing number of stratified studies on HLA ligandomes, in the context of both basic and translational research. A key challenge in the case of MHCII immunopeptidomes, often determined for different samples at distinct conditions, is to derive quantitative information on consensus epitopes from antigenic peptides of variable lengths. Here, we present the design and benchmarking of a new algorithm [peptide landscape antigenic epitope alignment utility (PLAtEAU] allowing the identification and label-free quantification (LFQ of shared consensus epitopes arising from series of nested peptides. The algorithm simplifies the complexity of the dataset while allowing the identification of nested peptides within relatively short segments of protein sequences. Moreover, we apply this algorithm to the comparison of the ligandomes of cell lines with two different expression levels of the peptide-exchange catalyst HLA-DM. Direct comparison of LFQ intensities determined at the peptide level is inconclusive, as most of the peptides are not significantly enriched due to poor sampling. Applying the PLAtEAU algorithm for grouping of the peptides into consensus epitopes shows that more than half of the total number of epitopes is preferentially and significantly enriched for each condition. This simplification and deconvolution of the complex and ambiguous peptide-level dataset highlights the value of the PLAt

  1. An MHC-restricted antibody-based chimeric antigen receptor requires TCR-like affinity to maintain antigen specificity

    Directory of Open Access Journals (Sweden)

    Marcela V Maus

    2016-01-01

    Full Text Available Chimeric antigen receptors (CARs are synthetic receptors that usually redirect T cells to surface antigens independent of human leukocyte antigen (HLA. Here, we investigated a T cell receptor-like CAR based on an antibody that recognizes HLA-A*0201 presenting a peptide epitope derived from the cancer-testis antigen NY-ESO-1. We hypothesized that this CAR would efficiently redirect transduced T cells in an HLA-restricted, antigen-specific manner. However, we found that despite the specificity of the soluble Fab, the same antibody in the form of a CAR caused moderate lysis of HLA-A2 expressing targets independent of antigen owing to T cell avidity. We hypothesized that lowering the affinity of the CAR for HLA-A2 would improve its specificity. We undertook a rational approach of mutating residues that, in the crystal structure, were predicted to stabilize binding to HLA-A2. We found that one mutation (DN lowered the affinity of the Fab to T cell receptor-range and restored the epitope specificity of the CAR. DN CAR T cells lysed native tumor targets in vitro, and, in a xenogeneic mouse model implanted with two human melanoma lines (A2+/NYESO+ and A2+/NYESO−, DN CAR T cells specifically migrated to, and delayed progression of, only the HLA-A2+/NY-ESO-1+ melanoma. Thus, although maintaining MHC-restricted antigen specificity required T cell receptor-like affinity that decreased potency, there is exciting potential for CARs to expand their repertoire to include a broad range of intracellular antigens.

  2. Shedding of the immunodominant P20 surface antigen of Eimeria bovis sporozoites.

    OpenAIRE

    Speer, C A; Whitmire, W M

    1989-01-01

    P20 is an immunodominant surface antigen of Eimeria bovis sporozoites. As parasites underwent merogony within cultured bovine monocytes and Madin-Darby bovine kidney (MDBK) cells, P20 appeared to be shed gradually by meronts and was absent in type 1 and 2 first-generation merozoites. Meronts of E. bovis appeared to shed P20 into the parasitophorous vacuole of bovine monocytes, whereas MDBK cells evidently released P20 into the culture medium or destroyed its antigenic determinant.

  3. Ultraviolet light-induced suppression of antigen presentation

    International Nuclear Information System (INIS)

    Spellman, C.W.; Tomasi, T.B.

    1983-01-01

    Ultraviolet (UV) light irradiation of animals results in the development of specific T suppressor cells that inhibit antitumor immune responses. It is thought that suppression may arise as a consequence of altered antigen presentation by UV-irradiated epidermal cells. This hypothesis is based on evidence demonstrating that specific lymphoid tissues from UV-irradiated hosts exhibit impaired antigen-presenting function and that animals cannot be contact sensitized when antigens are applied to a UV-irradiated skin site. Langerhans cells of the skin are likely candidates as targets of UV-induced defects in antigen presentation as they bear Fc and C3b receptors, express Ia antigens, are of bone marrow origin, and are capable of presenting antigen in vitro. We speculate on the possible clinical usefulness of UV-induced tolerance to specific antigens such as those encountered in monoclonal antibody therapy and tissue transplantation

  4. Suppressive versus augmenting effect of the same pretreatment regimen in two murine tumor systems with distinct effector mechanisms

    International Nuclear Information System (INIS)

    Fujiwara, Hiromi; Hamaoka, Toshiyuki; Kitagawa, Masayasu

    1978-01-01

    The effect of presensitization with x-irradiated tumor cells on the development of host's immune resistance against the tumor-associated transplantation antigens (TATA) was investigated in two syngeneic tumor systems with distinct effector mechanisms. When X5563 plasmacytoma, to which immune resistance was mediated exclusively by killer T lymphocytes, was intravenously inoculated into syngeneic C3H/He mice with lower number after 7000 R x-irradiation, the mice failed to exhibit any protective immunity against the subsequent challenge with viable tumor cells. Moreover, these mice lost their capability to develop any immune resistance even after an appropriate immunization procedure. The immunodepression induced by such a pretreatment regimen was specific for X5563 tumor. While no suppressor cell activity was detected in the above pretreated mice, serum factor(s) from these mice was virtually responsible for this suppression. When the serum factor mediating this tumor-specific suppression was fractionated on the Sephadex G-200 column, the suppressive activity was found in albumin-corresponding fraction, free of any immunoglobulin component. In contrast, in MM102 mammary tumor system, in which immune resistance is solely mediated by tumor-specific antibody, the pretreatment with x-irradiated MM102 cells augmented the induction of anti-tumor immunity. These results indicate that while tumor antigens given in the form of x-irradiated tumor cells suppress the induction of killer T cell-mediated immunity in one system, the same presensitization regimen of tumor antigens augments the antibody-mediated immunity in another system, thus giving a divergent effect on the distinct effector mechanisms of syngeneic tumor immunity. (author)

  5. Binding of hydrophobic antigens to surfaces

    DEFF Research Database (Denmark)

    2017-01-01

    A first aspect of the present invention is a method of detecting antibodies comprising the steps of: i) providing a first group of beads comprising a surface modified with C1-C10 alkyl groups comprising amine, ammonium, ether and/or hydroxyl groups, ii) contacting said first group of beads......-antigen-antibody conjugates, and v) detecting said bead-antigen-antibody conjugates. Further aspects include an antibody detection kit, a bead-antigen conjugate and a composition comprising at least two different groups of bead-antigen-conjugates....

  6. Humoral immunity provides resident intestinal eosinophils access to luminal antigen via eosinophil-expressed low affinity Fc gamma receptors

    Science.gov (United States)

    Smith, Kalmia M.; Rahman, Raiann S.; Spencer, Lisa A.

    2016-01-01

    Eosinophils are native to the healthy gastrointestinal tract, and are associated with inflammatory diseases likely triggered by exposure to food allergens (e.g. food allergies and eosinophilic gastrointestinal disorders). In models of allergic respiratory diseases and in vitro studies, direct antigen engagement elicits eosinophil effector functions including degranulation and antigen presentation. However, it was not known whether intestinal tissue eosinophils that are separated from luminal food antigens by a columnar epithelium might similarly engage food antigens. Using an intestinal ligated loop model in mice, here we determined that resident intestinal eosinophils acquire antigen from the lumen of antigen-sensitized but not naïve mice in vivo. Antigen acquisition was immunoglobulin-dependent; intestinal eosinophils were unable to acquire antigen in sensitized immunoglobulin-deficient mice, and passive immunization with immune serum or antigen-specific IgG was sufficient to enable intestinal eosinophils in otherwise naïve mice to acquire antigen in vivo. Intestinal eosinophils expressed low affinity IgG receptors, and the activating receptor FcγRIII was necessary for immunoglobulin-mediated acquisition of antigens by isolated intestinal eosinophils in vitro. Our combined data suggest that intestinal eosinophils acquire lumen-derived food antigens in sensitized mice via FcγRIII antigen focusing, and may therefore participate in antigen-driven secondary immune responses to oral antigens. PMID:27683752

  7. Human antibody recognition of antigenic site IV on Pneumovirus fusion proteins.

    Science.gov (United States)

    Mousa, Jarrod J; Binshtein, Elad; Human, Stacey; Fong, Rachel H; Alvarado, Gabriela; Doranz, Benjamin J; Moore, Martin L; Ohi, Melanie D; Crowe, James E

    2018-02-01

    Respiratory syncytial virus (RSV) is a major human pathogen that infects the majority of children by two years of age. The RSV fusion (F) protein is a primary target of human antibodies, and it has several antigenic regions capable of inducing neutralizing antibodies. Antigenic site IV is preserved in both the pre-fusion and post-fusion conformations of RSV F. Antibodies to antigenic site IV have been described that bind and neutralize both RSV and human metapneumovirus (hMPV). To explore the diversity of binding modes at antigenic site IV, we generated a panel of four new human monoclonal antibodies (mAbs) and competition-binding suggested the mAbs bind at antigenic site IV. Mutagenesis experiments revealed that binding and neutralization of two mAbs (3M3 and 6F18) depended on arginine (R) residue R429. We discovered two R429-independent mAbs (17E10 and 2N6) at this site that neutralized an RSV R429A mutant strain, and one of these mAbs (17E10) neutralized both RSV and hMPV. To determine the mechanism of cross-reactivity, we performed competition-binding, recombinant protein mutagenesis, peptide binding, and electron microscopy experiments. It was determined that the human cross-reactive mAb 17E10 binds to RSV F with a binding pose similar to 101F, which may be indicative of cross-reactivity with hMPV F. The data presented provide new concepts in RSV immune recognition and vaccine design, as we describe the novel idea that binding pose may influence mAb cross-reactivity between RSV and hMPV. Characterization of the site IV epitope bound by human antibodies may inform the design of a pan-Pneumovirus vaccine.

  8. Molecular cloning of cDNA for the human tumor-associated antigen CO-029 and identification of related transmembrane antigens

    International Nuclear Information System (INIS)

    Szala, S.; Kasai, Yasushi; Steplewski, Z.; Rodeck, U.; Koprowski, H.; Linnenbach, A.J.

    1990-01-01

    The human tumor-associated antigen CO-029 is a monoclonal antibody-defined cell surface glycoprotein of 27-34 kDa. By using the high-efficiency COS cell expression system, a full-length cDNA clone for CO-029 was isolated. When transiently expressed in COS cells, the cDNA clone directed the synthesis of an antigen reactive to monoclonal antibody CO-029 in mixed hemadsorption and immunoblot assays. Sequence analysis revealed that CO-029 belongs to a family of cell surface antigens that includes the melanoma-associated antigen ME491, the leukocyte cell surface antigen CD37, and the Sm23 antigen of the parasitic helminth Schistosoma mansoni. CO-029 and ME491 antigen expression and the effect of their corresponding monoclonal antibodies on cell growth were compared in human tumor cell lines of various histologic origins

  9. Synthesis and evaluation of artificial antigens for astragaloside IV

    Directory of Open Access Journals (Sweden)

    Sheng-lan Yu

    Full Text Available The objective of this study was to produce artificial antigens for astragaloside IV that could be used to prepare antibodies against astragaloside IV screened in Radix astragali (Astragalus membranaceus (Fisch Bunge, Fabaceae and its preparations, using an indirect ELISA. Astragaloside IV was coupled to carrier proteins, bovine serum albumin and ovalbumin using the sodium periodate method and was then evaluated using SDS-PAGE, MALDI-TOF MS and animal immunizations. The coupling ratio of astragaloside IV to bovine serum albumin ratio was determined to be thirteen, and the indirect ELISA demonstrated that three groups of mice immunized with astragaloside IV-bovine serum albumin produced anti-astragaloside IV- bovine serum albumin-specific antibody, with a minimum serum titer of 1:9600. A method for synthesizing highly immunogenic astragaloside IV artificial antigens was successfully developed thus indicating its feasibility in the establishment of a fast immunoassay for astragaloside IV content determination in Radix astragali and its products.

  10. Comprehensive Analysis of the Activation and Proliferation Kinetics and Effector Functions of Human Lymphocytes, and Antigen Presentation Capacity of Antigen-Presenting Cells in Xenogeneic Graft-Versus-Host Disease.

    Science.gov (United States)

    Kawasaki, Yasufumi; Sato, Kazuya; Hayakawa, Hiroko; Takayama, Norihito; Nakano, Hirofumi; Ito, Ryoji; Mashima, Kiyomi; Oh, Iekuni; Minakata, Daisuke; Yamasaki, Ryoko; Morita, Kaoru; Ashizawa, Masahiro; Yamamoto, Chihiro; Hatano, Kaoru; Fujiwara, Shin-Ichiro; Ohmine, Ken; Muroi, Kazuo; Kanda, Yoshinobu

    2018-04-17

    Xenogeneic graft-versus-host disease (GVHD) models in highly immunodeficient mice are currently being used worldwide to investigate human immune responses against foreign antigens in vivo. However, the individual roles of CD4 + and CD8 + T cells, and donor/host hematopoietic and nonhematopoietic antigen-presenting cells (APCs) in the induction and development of GVHD have not been fully investigated. In the present study, we comprehensively investigated the immune responses of human T cells and the antigen presentation capacity of donor/host hematopoietic and nonhematopoietic APCs in xenogeneic GVHD models using nonobese diabetic/Shi-scid-IL2rg null mice. CD4 + T cells and, to a lesser extent, CD8 + T cells individually mediated potentially lethal GVHD. In addition to inflammatory cytokine production, CD4 + T cells also supported the activation and proliferation of CD8 + T cells. Using bone marrow chimeras, we demonstrated that host hematopoietic, but not nonhematopoietic, APCs play a critical role in the development of CD4 + T cell-mediated GVHD. During early GVHD, we detected 2 distinct populations in memory CD4 + T cells. One population was highly activated and proliferated in major histocompatibility complex antigen (MHC) +/+ mice but not in MHC -/- mice, indicating alloreactive T cells. The other population showed a less activated and slowly proliferative status regardless of host MHC expression, and was associated with higher susceptibility to apoptosis, indicating nonalloreactive T cells in homeostasis-driven proliferation. These observations are clinically relevant to donor T cell response after allogeneic hematopoietic stem cell transplantation. Our findings provide a better understanding of the immunobiology of humanized mice and support the development of novel options for the prevention and treatment for GVHD. Copyright © 2018. Published by Elsevier Inc.

  11. Deteksi Antigen pada Kriptokokosis

    Directory of Open Access Journals (Sweden)

    Robiatul  Adawiyah

    2014-12-01

    number of fungi is around 10 10 cells/ml. Cultur examination is performed in Sabouraud dextrose agar (SDA and niger sheed agar (NSA medium, fungi grows in 5­7 days. Antigen and antibody detection could be performed on result in acute infection, IgA still positive after 1­2 years of healing phase and IgG can be persistent. The immunocompromised person showed very complex result and inconsistent in determining the diagnosis. Polysaccharides are the most instrumental component in Cr. neoformans virulence. The component of Polysaccharide especially glucuronoxylomannan is the most important marker in the diagnosis of cryptococcosis. Antigen detection of Cr. neoformans can show positive result in acute/ Keywords: Cr. neoformans, glucuronoxylomannan, antigen

  12. Allergens in Hymenoptera venom. XXV: The amino acid sequences of antigen 5 molecules and the structural basis of antigenic cross-reactivity.

    Science.gov (United States)

    Hoffman, D R

    1993-11-01

    The complete amino acid sequences have been determined by solid-phase protein sequencing for eight different vespid venom antigen 5 molecules. These include five species of yellow jackets, Vespula squamosa, V. flavopilosa, V. germanica, V. pensylvanica and V. vidua, representing all three species groups; two variants from the European hornet, Vespa crabro; and a species of paper wasp, Polistes fuscatus, from a second subgenus. The new sequences were compared with the seven previously published sequences from yellow jackets, hornets, and wasps, and to that of Solenopsis invicta 3 allergen from imported fire ant venom. These comparisons provided structural evidence to support the observed high degree of cross-reactivity among the antigens of the common group of yellow jackets and among those of the two common North American subgenera of paper wasps studied. The antigen 5 of V. squamosa and of V. vidua were significantly different from those of the vulgaris group. Common features that could generate immunologic cross-reactivity were seen among the antigen 5 molecules of hornets of both genera and among those of yellow jackets, hornets, and paper wasps. The imported fire ant allergen has only minimal conserved areas in common with the vespid allergens, which explains the lack of observed IgE cross-reactivity. These results provide the structural basis for the cross-reactivity patterns observed in clinical practice and suggest that the commercial extracts of yellow jacket and paper wasp could be prepared with fewer carefully selected species.

  13. Predictive value of different prostate-specific antigen-based markers in men with baseline total prostate-specific antigen <2.0 ng/mL.

    Science.gov (United States)

    Fujizuka, Yuji; Ito, Kazuto; Oki, Ryo; Suzuki, Rie; Sekine, Yoshitaka; Koike, Hidekazu; Matsui, Hiroshi; Shibata, Yasuhiro; Suzuki, Kazuhiro

    2017-08-01

    To investigate the predictive value of various molecular forms of prostate-specific antigen in men with baseline prostate-specific antigen baseline prostate-specific antigen level baseline prostate-specific antigen- and age-adjusted men who did not develop prostate cancer. Serum prostate-specific antigen, free prostate-specific antigen, and [-2] proenzyme prostate-specific antigen were measured at baseline and last screening visit. The predictive impact of baseline prostate-specific antigen- and [-2] proenzyme prostate-specific antigen-related indices on developing prostate cancer was investigated. The predictive impact of those indices at last screening visit and velocities from baseline to final screening on tumor aggressiveness were also investigated. The baseline free to total prostate-specific antigen ratio was a significant predictor of prostate cancer development. The odds ratio was 6.08 in the lowest quintile baseline free to total prostate-specific antigen ratio subgroup. No serum indices at diagnosis were associated with tumor aggressiveness. The Prostate Health Index velocity and [-2] proenzyme prostate-specific antigen/free prostate-specific antigen velocity significantly increased in patients with higher risk D'Amico risk groups and higher Gleason scores. Free to total prostate-specific antigen ratio in men with low baseline prostate-specific antigen levels seems to predict the risk of developing prostate cancer, and it could be useful for a more effective individualized screening system. Longitudinal changes in [-2] proenzyme prostate-specific antigen-related indices seem to correlate with tumor aggressiveness, and they could be used as prognostic tool before treatment and during active surveillance. © 2017 The Japanese Urological Association.

  14. The extended family of CD1d-restricted T cells: sifting through a mixed bag of TCRs, antigens and functions

    Directory of Open Access Journals (Sweden)

    Elodie eMacho-Fernandez

    2015-07-01

    Full Text Available Natural killer T (NKT cells comprise a family of specialized T cells that recognize lipid antigens presented by CD1d. Based on their T cell receptor (TCR usage and antigen-specificities, CD1d-restricted NKT cells have been divided into two main subsets: type I NKT cells that use a canonical invariant TCR α-chain and recognize α-galactosylceramide (α-GalCer, and type II NKT cells that use a more diverse αβ TCR repertoire and do not recognize α-GalCer. In addition, α-GalCer-reactive NKT cells that use non-canonical αβ TCRs and CD1d-restricted T cells that use γδ or δ/αβ TCRs have recently been identified, revealing further diversity among CD1d-restricted T cells. Importantly, in addition to their distinct antigen specificities, functional differences are beginning to emerge between the different members of the CD1d-restricted T cell family. In this review, while using type I NKT cells as comparison, we will focus on type II NKT cells and the other non-invariant CD1d-restricted T cell subsets, and discuss our current understanding of the antigens they recognize, the formation of stimulatory CD1d/antigen complexes, the modes of TCR-mediated antigen recognition, and the mechanisms and consequences of their activation that underlie their function in antimicrobial responses, antitumor immunity, and autoimmunity.

  15. The Extended Family of CD1d-Restricted NKT Cells: Sifting through a Mixed Bag of TCRs, Antigens, and Functions.

    Science.gov (United States)

    Macho-Fernandez, Elodie; Brigl, Manfred

    2015-01-01

    Natural killer T (NKT) cells comprise a family of specialized T cells that recognize lipid antigens presented by CD1d. Based on their T cell receptor (TCR) usage and antigen specificities, CD1d-restricted NKT cells have been divided into two main subsets: type I NKT cells that use a canonical invariant TCR α-chain and recognize α-galactosylceramide (α-GalCer), and type II NKT cells that use a more diverse αβ TCR repertoire and do not recognize α-GalCer. In addition, α-GalCer-reactive NKT cells that use non-canonical αβ TCRs and CD1d-restricted T cells that use γδ or δ/αβ TCRs have recently been identified, revealing further diversity among CD1d-restricted T cells. Importantly, in addition to their distinct antigen specificities, functional differences are beginning to emerge between the different members of the CD1d-restricted T cell family. In this review, while using type I NKT cells as comparison, we will focus on type II NKT cells and the other non-invariant CD1d-restricted T cell subsets, and discuss our current understanding of the antigens they recognize, the formation of stimulatory CD1d/antigen complexes, the modes of TCR-mediated antigen recognition, and the mechanisms and consequences of their activation that underlie their function in antimicrobial responses, anti-tumor immunity, and autoimmunity.

  16. Identification of antigenic Sarcoptes scabiei proteins for use in a diagnostic test and of non-antigenic proteins that may be immunomodulatory

    Science.gov (United States)

    Morgan, Marjorie S.; Rider, S. Dean; Arlian, Larry G.

    2017-01-01

    Background Scabies, caused by the mite, Sarcoptes scabiei, infects millions of humans, and many wild and domestic mammals. Scabies mites burrow in the lower stratum corneum of the epidermis of the skin and are the source of substances that are antigenic or modulate aspects of the protective response of the host. Ordinary scabies is a difficult disease to diagnose. Objective The goal of this project was to identify S. scabiei proteins that may be candidate antigens for use in a diagnostic test or may be used by the mite to modulate the host’s protective response. Methods An aqueous extract of S. scabiei was separated by 2-dimensional electrophoresis and proteins were identified by mass spectrometry. A parallel immunoblot was probed with serum from patients with ordinary scabies to identify IgM and/or IgG-binding antigens. The genes coding for 23 selected proteins were cloned into E. coli and the expressed recombinant proteins were screened with serum from patients with confirmed ordinary scabies. Results We identified 50 different proteins produced by S. scabiei, 34 of which were not previously identified, and determined that 66% were recognized by patient IgM and/or IgG. Fourteen proteins were screened for use in a diagnostic test but none possessed enough sensitivity and specificity to be useful. Six of the 9 proteins selected for the possibility that they may be immunomodulatory were not recognized by antibodies in patient serum. Conclusions Thirty-three proteins that bound IgM and/or IgG from the serum of patients with ordinary scabies were identified. None of the 14 tested were useful for inclusion in a diagnostic test. The identities of 16 proteins that are not recognized as antigens by infected patients were also determined. These could be among the molecules that are responsible for this mite’s ability to modulate its host’s innate and adaptive immune responses. PMID:28604804

  17. Understanding the immunogenicity and antigenicity of nanomaterials: Past, present and future

    International Nuclear Information System (INIS)

    Ilinskaya, Anna N.; Dobrovolskaia, Marina A.

    2016-01-01

    Nanoparticle immunogenicity and antigenicity have been under investigation for many years. During the past decade, significant progress has been made in understanding what makes a nanoparticle immunogenic, how immune cells respond to nanoparticles, what consequences of nanoparticle-specific antibody formation exist and how they challenge the application of nanoparticles for drug delivery. Moreover, it has been recognized that accidental contamination of therapeutic protein formulations with nanosized particulate materials may contribute to the immunogenicity of this type of biotechnology products. While the immunological properties of engineered nanomaterials and their application as vaccine carriers and adjuvants have been given substantial consideration in the current literature, little attention has been paid to nanoparticle immuno- and antigenicity. To fill in this gap, we herein provide an overview of this subject to highlight the current state of the field, review past and present research, and discuss future research directions. - Highlights: • Most engineered nanomaterials are not immunogenic per se. • Generation of nanoparticle-specific antibody can be T-cell dependent or independent. • Antibodies can be generated to particle core, terminal groups or surface coatings. • Engineered and accidental nanomaterials have distinct contribution to immunogenicity. • Tunable physicochemical properties make each nanoparticle unique.

  18. Maximum-Entropy Models of Sequenced Immune Repertoires Predict Antigen-Antibody Affinity.

    Directory of Open Access Journals (Sweden)

    Lorenzo Asti

    2016-04-01

    Full Text Available The immune system has developed a number of distinct complex mechanisms to shape and control the antibody repertoire. One of these mechanisms, the affinity maturation process, works in an evolutionary-like fashion: after binding to a foreign molecule, the antibody-producing B-cells exhibit a high-frequency mutation rate in the genome region that codes for the antibody active site. Eventually, cells that produce antibodies with higher affinity for their cognate antigen are selected and clonally expanded. Here, we propose a new statistical approach based on maximum entropy modeling in which a scoring function related to the binding affinity of antibodies against a specific antigen is inferred from a sample of sequences of the immune repertoire of an individual. We use our inference strategy to infer a statistical model on a data set obtained by sequencing a fairly large portion of the immune repertoire of an HIV-1 infected patient. The Pearson correlation coefficient between our scoring function and the IC50 neutralization titer measured on 30 different antibodies of known sequence is as high as 0.77 (p-value 10-6, outperforming other sequence- and structure-based models.

  19. Understanding the immunogenicity and antigenicity of nanomaterials: Past, present and future

    Energy Technology Data Exchange (ETDEWEB)

    Ilinskaya, Anna N.; Dobrovolskaia, Marina A., E-mail: marina@mail.nih.gov

    2016-05-15

    Nanoparticle immunogenicity and antigenicity have been under investigation for many years. During the past decade, significant progress has been made in understanding what makes a nanoparticle immunogenic, how immune cells respond to nanoparticles, what consequences of nanoparticle-specific antibody formation exist and how they challenge the application of nanoparticles for drug delivery. Moreover, it has been recognized that accidental contamination of therapeutic protein formulations with nanosized particulate materials may contribute to the immunogenicity of this type of biotechnology products. While the immunological properties of engineered nanomaterials and their application as vaccine carriers and adjuvants have been given substantial consideration in the current literature, little attention has been paid to nanoparticle immuno- and antigenicity. To fill in this gap, we herein provide an overview of this subject to highlight the current state of the field, review past and present research, and discuss future research directions. - Highlights: • Most engineered nanomaterials are not immunogenic per se. • Generation of nanoparticle-specific antibody can be T-cell dependent or independent. • Antibodies can be generated to particle core, terminal groups or surface coatings. • Engineered and accidental nanomaterials have distinct contribution to immunogenicity. • Tunable physicochemical properties make each nanoparticle unique.

  20. A method for visualizing surface-exposed and internal PfEMP1 adhesion antigens in Plasmodium falciparum infected erythrocytes

    Directory of Open Access Journals (Sweden)

    Arnot David E

    2008-06-01

    Full Text Available Abstract Background The insertion of parasite antigens into the host erythrocyte membrane and the structure and distribution of Plasmodium falciparum adhesion receptors on that membrane are poorly understood. Laser scanning confocal microscopy (LSCM and a novel labelling and fixation method have been used to obtain high resolution immuno-fluorescent images of erythrocyte surface PfEMP1 and internal antigens which allow analysis of the accumulation of PfEMP1 on the erythrocyte membrane during asexual development. Methods A novel staining technique has been developed which permits distinction between erythrocyte surface PfEMP1 and intracellular PfEMP1, in parasites whose nuclear material is exceptionally well resolved. Primary antibody detection by fluorescence is carried out on the live parasitized erythrocyte. The surface labelled cells are then fixed using paraformaldehyde and permeabilized with a non-ionic detergent to permit access of antibodies to internal parasite antigens. Differentiation between surface and internal antigens is achieved using antibodies labelled with different fluorochromes and confocal microscopy Results Surface exposed PfEMP1 is first detectable by antibodies at the trophozoite stage of intracellular parasite development although the improved detection method indicates that there are differences between different laboratory isolates in the kinetics of accumulation of surface-exposed PfEMP1. Conclusion A sensitive method for labelling surface and internal PfEMP1 with up to three different fluorochromes has been developed for laser scanning confocal optical microscopy and the analysis of the developmental expression of malaria adhesion antigens.

  1. Selective elution of HLA antigens and beta 2-microglobulin from human platelets by chloroquine diphosphate

    International Nuclear Information System (INIS)

    Kao, K.J.

    1988-01-01

    To determine whether chloroquine can specifically elute HLA antigens and beta 2-microglobulin (beta 2-M) from the platelet surface, quantitative immunofluorescence flow cytometry and monoclonal antibodies were used to show that HLA antigens and beta 2-M were proportionally eluted from the platelet surface without affecting the membrane glycoproteins IIb and IIIa. Second, an autoradiogram of electrophoresed I-125-labeled platelets showed that only beta 2-M but not other I-125-labeled membrane proteins could be eluted. Although HLA antigens were poorly labeled by I-125 and could not be detected on the autoradiogram, the eluted HLA antigens could be detected by anti-HLA monoclonal antibody and immunoblotting techniques. No loss of plasma membrane integrity was observed by transmission electron microscopy after chloroquine treatment of platelets. The results indicate that chloroquine selectively elutes HLA antigens and their noncovalently associated beta 2-M without affecting other integral platelet membrane proteins

  2. Tissue distribution of histo-blood group antigens

    DEFF Research Database (Denmark)

    Ravn, V; Dabelsteen, Erik

    2000-01-01

    carrier carbohydrate chains. Histo-blood group antigens are found in most epithelial tissues. Meanwhile, several factors influence the type, the amount, and the histological distribution of histoblood group antigens, i.e. the ABO, Lewis, and saliva-secretor type of the individual, and the cell- and tissue......The introduction of immunohistochemical techniques and monoclonal antibodies to specific carbohydrate epitopes has made it possible to study in detail the tissue distribution of histo-blood group antigens and related carbohydrate structures. The present paper summarizes the available data...... concerning the histological distribution of histo-blood group antigens and their precursor structures in normal human tissues. Studies performed have concentrated on carbohydrate antigens related to the ABO, Lewis, and TTn blood group systems, i.e. histo-blood group antigens carried by type 1, 2, and 3 chain...

  3. IgM and IgD B cell receptors differentially respond to endogenous antigens and control B cell fate

    Science.gov (United States)

    Noviski, Mark; Mueller, James L; Satterthwaite, Anne; Garrett-Sinha, Lee Ann; Brombacher, Frank

    2018-01-01

    Naive B cells co-express two BCR isotypes, IgM and IgD, with identical antigen-binding domains but distinct constant regions. IgM but not IgD is downregulated on autoreactive B cells. Because these isotypes are presumed to be redundant, it is unknown how this could impose tolerance. We introduced the Nur77-eGFP reporter of BCR signaling into mice that express each BCR isotype alone. Despite signaling strongly in vitro, IgD is less sensitive than IgM to endogenous antigen in vivo and developmental fate decisions are skewed accordingly. IgD-only Lyn−/− B cells cannot generate autoantibodies and short-lived plasma cells (SLPCs) in vivo, a fate thought to be driven by intense BCR signaling induced by endogenous antigens. Similarly, IgD-only B cells generate normal germinal center, but impaired IgG1+ SLPC responses to T-dependent immunization. We propose a role for IgD in maintaining the quiescence of autoreactive B cells and restricting their differentiation into autoantibody secreting cells. PMID:29521626

  4. Antigen delivery by α2-macroglobulin enhances the cytotoxic T lymphocyte response

    Science.gov (United States)

    Bowers, Edith V.; Horvath, Jeffrey J.; Bond, Jennifer E.; Cianciolo, George J.; Pizzo, Salvatore V.

    2009-01-01

    α2M* targets antigens to APCs for rapid internalization, processing, and presentation. When used as an antigen-delivery vehicle, α2M* amplifies MHC class II presentation, as demonstrated by increased antibody titers. Recent evidence, however, suggests that α2M* encapsulation may also enhance antigen-specific CTL immunity. In this study, we demonstrate that α2M*-delivered antigen (OVA) enhances the production of specific in vitro and in vivo CTL responses. Murine splenocytes expressing a transgenic TCR specific for CTL peptide OVA257–264 (SIINFEKL) demonstrated up to 25-fold greater IFN-γ and IL-2 secretion when treated in vitro with α2M*-OVA compared with soluble OVA. The frequency of IFN-γ-producing cells was increased ∼15-fold, as measured by ELISPOT. Expansion of the OVA-specific CD8+ T cell population, as assayed by tetramer binding and [3H]thymidine incorporation, and OVA-specific cell-mediated cytotoxicity, as determined by a flow cytometric assay, were also enhanced significantly by α2M*-OVA. Furthermore, significant CTL responses were observed at antigen doses tenfold lower than those required with OVA alone. Finally, we also observed enhanced humoral and CTL responses by naïve mice following intradermal immunization with α2M*-OVA. These α2M*-OVA-immunized mice demonstrated increased protection against a s.c.-implanted, OVA-expressing tumor, as demonstrated by delayed tumor growth and prolonged animal survival. The observation that α2M*-mediated antigen delivery elicits specific CTL responses suggests the cross-presentation of antigen onto MHC class I. These results support α2M* as an effective antigen-delivery system that may be particularly useful for vaccines based on weakly immunogenic subunits or requiring dose sparing. PMID:19652028

  5. A molecular approach to immunoscintigraphy: A study of the T-antigen conformation on the surface of tumors

    International Nuclear Information System (INIS)

    Noujaim, A.; Selvaraj, S.; Suresh, M.R.; Turner, C.; McLean, G.; Willans, D.; Longenecker, B.M.; Haines, D.M.

    1987-01-01

    The role of glycoconjugates in tumor cell differentiation has been well documented. We have examined the expression of the two anomers of the Thomsen-Friedenreich antigen on the surface of human, canine and murine tumor cell membranes both in vitro and in vivo. This has been accomplished through the synthesis of the disaccharide terminal residues in both α and β configuration. Both entities were used to generate murine monoclonal antibodies which recognized the carbohydrate determinants. The determination of fine specificities of these antibodies was effected by means of cellular uptake, immunohistopathology and immunoscintigraphy. Examination of pathological specimens of human and canine tumor tissue indicated that the expressed antigen was in the β configuration. More than 89% of all human carcinomas tested expressed the antigen in the above anomeric form. The combination of synthetic antigens and monoclonal antibodies raised specifically against them provide us with invaluable tools for the study of tumor marker expression in humans and their respective animal tumor models. (orig.) [de

  6. Comparison of antigen-specific T-cell responses of tuberculosis patients using complex or single antigens of Mycobacterium tuberculosis

    DEFF Research Database (Denmark)

    Mustafa, A S; Amoudy, H A; Wiker, H G

    1998-01-01

    We have screened peripheral blood mononuclear cells (PBMC) from tuberculosis (TB) patients for proliferative reactivity and interferon-gamma (IFN-gamma) secretion against a panel of purified recombinant (r) and natural (n) culture filtrate (rESAT-6, nMPT59, nMPT64 and nMPB70) and somatic-derived (r......GroES, rPstS, rGroEL and rDnaK) antigens of Mycobacterium tuberculosis. The responses of PBMC to these defined antigens were compared with the corresponding results obtained with complex antigens, such as whole-cell M. tuberculosis, M. tuberculosis culture filtrate (MT-CF) and cell wall antigens, as well...... as the vaccine strain, Mycobacterium bovis bacillus Calmette-Guerin (BCG). In addition, M. tuberculosis and MT-CF-induced T-cell lines were tested in the same assays against the panel of purified and complex antigens. The compiled data from PBMC and T-cell lines tested for antigen-induced proliferation and IFN...

  7. Genetically Distinct Subsets within ANCA-Associated Vasculitis

    Science.gov (United States)

    Lyons, Paul A.; Rayner, Tim F.; Trivedi, Sapna; Holle, Julia U.; Watts, Richard A.; Jayne, David R.W.; Baslund, Bo; Brenchley, Paul; Bruchfeld, Annette; Chaudhry, Afzal N.; Tervaert, Jan Willem Cohen; Deloukas, Panos; Feighery, Conleth; Gross, Wolfgang L.; Guillevin, Loic; Gunnarsson, Iva; P, Lorraine Harper M.R.C; Hrušková, Zdenka; Little, Mark A.; Martorana, Davide; Neumann, Thomas; Ohlsson, Sophie; Padmanabhan, Sandosh; Pusey, Charles D.; Salama, Alan D.; Sanders, Jan-Stephan F.; Savage, Caroline O.; Segelmark, Mårten; Stegeman, Coen A.; Tesař, Vladimir; Vaglio, Augusto; Wieczorek, Stefan; Wilde, Benjamin; Zwerina, Jochen; Rees, Andrew J.; Clayton, David G.; Smith, Kenneth G.C.

    2013-01-01

    BACKGROUND Antineutrophil cytoplasmic antibody (ANCA)–associated vasculitis is a severe condition encompassing two major syndromes: granulomatosis with polyangiitis (formerly known as Wegener’s granulomatosis) and microscopic polyangiitis. Its cause is unknown, and there is debate about whether it is a single disease entity and what role ANCA plays in its pathogenesis. We investigated its genetic basis. METHODS A genomewide association study was performed in a discovery cohort of 1233 U.K. patients with ANCA-associated vasculitis and 5884 controls and was replicated in 1454 Northern European case patients and 1666 controls. Quality control, population stratification, and statistical analyses were performed according to standard criteria. RESULTS We found both major-histocompatibility-complex (MHC) and non-MHC associations with ANCA-associated vasculitis and also that granulomatosis with polyangiitis and microscopic polyangiitis were genetically distinct. The strongest genetic associations were with the antigenic specificity of ANCA, not with the clinical syndrome. Anti–proteinase 3 ANCA was associated with HLA-DP and the genes encoding α1-antitrypsin (SERPINA1) and proteinase 3 (PRTN3) (P = 6.2×10−89, P = 5.6×10−12, and P = 2.6×10−7, respectively). Anti–myeloperoxidase ANCA was associated with HLA-DQ (P = 2.1×10−8). CONCLUSIONS This study confirms that the pathogenesis of ANCA-associated vasculitis has a genetic component, shows genetic distinctions between granulomatosis with polyangiitis and microscopic polyangiitis that are associated with ANCA specificity, and suggests that the response against the autoantigen proteinase 3 is a central pathogenic feature of proteinase 3 ANCA–associated vasculitis. These data provide preliminary support for the concept that proteinase 3 ANCA–associated vasculitis and myeloperoxidase ANCA–associated vasculitis are distinct autoimmune syndromes. (Funded by the British Heart Foundation and others.) PMID

  8. Protamine-based nanoparticles as new antigen delivery systems.

    Science.gov (United States)

    González-Aramundiz, José Vicente; Peleteiro Olmedo, Mercedes; González-Fernández, África; Alonso Fernández, María José; Csaba, Noemi Stefánia

    2015-11-01

    The use of biodegradable nanoparticles as antigen delivery vehicles is an attractive approach to overcome the problems associated with the use of Alum-based classical adjuvants. Herein we report, the design and development of protamine-based nanoparticles as novel antigen delivery systems, using recombinant hepatitis B surface antigen as a model viral antigen. The nanoparticles, composed of protamine and a polysaccharide (hyaluronic acid or alginate), were obtained using a mild ionic cross-linking technique. The size and surface charge of the nanoparticles could be modulated by adjusting the ratio of the components. Prototypes with optimal physicochemical characteristics and satisfactory colloidal stability were selected for the assessment of their antigen loading capacity, antigen stability during storage and in vitro and in vivo proof-of-concept studies. In vitro studies showed that antigen-loaded nanoparticles induced the secretion of cytokines by macrophages more efficiently than the antigen in solution, thus indicating a potential adjuvant effect of the nanoparticles. Finally, in vivo studies showed the capacity of these systems to trigger efficient immune responses against the hepatitis B antigen following intramuscular administration, suggesting the potential interest of protamine-polysaccharide nanoparticles as antigen delivery systems. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Development of indirect sandwich ELISA for determination of excretory-secretory antigens of Fasciola hepatica

    Directory of Open Access Journals (Sweden)

    Libertad Alzamora-Gonzales

    2016-05-01

    Full Text Available Fasciolosis is a cosmopolitan parasitosis medical-veterinary importance caused by Fasciola hepatica, which affects sheep, goats and cattle; and it affects man accidentally causing an epidemic-endemic infection difficult to diagnose. The aim was to develop an indirect sandwich ELISA with 3 antibodies for detecting excretory-secretory antigens of Fasciola hepatica (ESFh. For the development of indirect sandwich ELISA were used, as capture antibody, mouse polyclonal antibodies anti ESFh and polyclonal antibodies rabbit anti-ESFh as detection antibody, at the concentrations of 10 and 5 µg/mL respectively. The conjugate used was mouse monoclonal anti- total immunoglobulins rabbit linked to peroxidase (1/1000. Were analized 31 sheep fecal samples, and the results were compared with those obtained by direct coproparasitological examination (DC and counterimmunoelectrophoresis (CIEP. The detection limit obtained for indirect sandwich ELISA was 100 ng/mL. The test had a 100% sensitivity, 96.6% specificity, positive and negative predictive values of 50% and 96.6% respectively, in relation to DC test. Comparing with CIEP the specificity obtained for indirect sandwich ELISA was 93.5% and a negative predictive value of 100%. We concluded that indirect sandwich ELISA designed is able to detect metabolic antigens in ovine feces samples and can be used for Fasciola hepatica diagnosis.

  10. Application of preparative disk gel electrophoresis for antigen purification from inclusion bodies.

    Science.gov (United States)

    Okegawa, Yuki; Koshino, Masanori; Okushima, Teruya; Motohashi, Ken

    2016-02-01

    Specific antibodies are a reliable tool to examine protein expression patterns and to determine the protein localizations within cells. Generally, recombinant proteins are used as antigens for specific antibody production. However, recombinant proteins from mammals and plants are often overexpressed as insoluble inclusion bodies in Escherichia coli. Solubilization of these inclusion bodies is desirable because soluble antigens are more suitable for injection into animals to be immunized. Furthermore, highly purified proteins are also required for specific antibody production. Plastidic acetyl-CoA carboxylase (ACCase: EC 6.4.1.2) from Arabidopsis thaliana, which catalyzes the formation of malonyl-CoA from acetyl-CoA in chloroplasts, formed inclusion bodies when the recombinant protein was overexpressed in E. coli. To obtain the purified protein to use as an antigen, we applied preparative disk gel electrophoresis for protein purification from inclusion bodies. This method is suitable for antigen preparation from inclusion bodies because the purified protein is recovered as a soluble fraction in electrode running buffer containing 0.1% sodium dodecyl sulfate that can be directly injected into immune animals, and it can be used for large-scale antigen preparation (several tens of milligrams). Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Calorimetric comparison of the interactions between salivary proteins and Streptococcus mutans with and without antigen I/II

    NARCIS (Netherlands)

    Xu, C.P.; Belt-Gritter, van de B.; Busscher, H.J.; Mei, van der H.C.; Norde, W.

    2007-01-01

    Antigen I/II can be found on streptococcal cell surfaces and is involved in their interaction with salivary proteins. In this paper, we determine the adsorption enthalpies of salivary proteins to Streptococcus mutans LT11 and S. mutans IB03987 with and without antigen I/II, respectively, using

  12. Calorimetric comparison of the interactions between salivary proteins and Streptococcus mutans with and without antigen I/II

    NARCIS (Netherlands)

    Xu, Chun-Ping; Belt-Gritter, van de Betsy; Busscher, Henk J.; van der Mei, Henny C.; Norde, Willem

    2007-01-01

    Antigen I/II can be found on streptococcal cell surfaces and is involved in their interaction with salivary proteins. In this paper, we determine the adsorption enthalpies of salivary proteins to Streptococcus mutans LT 11 and S. mutans IB03987 with and without antigen I/II, respectively, using

  13. THE THEORETICAL AND PRACTICAL ASPECTS OF THE RECOMBINANT ANTIGENS FOR THE DIAGNOSIS OF BOVINE LEUKEMIA PRODUCTION

    Directory of Open Access Journals (Sweden)

    Shapovalova OV

    2016-12-01

    Full Text Available Introduction. Nowadays the problem of bovine leukemia (EBL effective diagnosis in countries where EBL is registered and the disease-free areas remains actual. The main diagnostic tests are immunodiffusion reaction (AGID and enzyme-linked immunosorbent assay (ELISA, which allow the identification of infected animals by the presence of antibodies to bovine leukemia virus (BLV both in serum and in milk samples. The effectiveness of these methods depends on the quality of diagnostic test systems used and determines by the cultural and recombinant virus antigens specificity. EBL recombinant antigens have certain advantages as they are more active and cheap. Purpose of the work. The analysis of theoretical and practical approaches in the bovine leukemia virus recombinant antigens development and its diagnostic potential evaluation. The article contains data from the literature on the recombinant antigens of bovine leukemia virus construction and use. Analysis of the literature showed that the recombinant proteins are widely used in the serological diagnosis of bovine leukemia. Numerous protocols of BLV gp51 and p24 immunodominant antigens preparation has been developed in heterologous systems (Saccharomyces cerevisiae, E. coli, vaccinia virus, baculovirus. In order to obtain recombinant antigens, the BLV provirus genome regions isolated from FLK-BLV cell culture, lymphocytes or tumor cells from naturally infected cattle are typically used. For the recombinant antigens labeled by hexahistidine or Srept II purification one-step immobilized-metal affinity chromatography IMAC and highly selective Strep-Tactin affinity chromatography methods are carried out. The end products activity and specificity are studied in the immunoblotting, ELISA and AGID diagnostic reactions. The ukrainian scientists’ publications are devoted to the clone E. coli HB101-2 transformed by the recombinant plasmid containing fully functional BLV env and gag genes nucleotide

  14. Human peripheral blood monocytes display surface antigens recognized by monoclonal antinuclear antibodies

    International Nuclear Information System (INIS)

    Holers, V.M.; Kotzin, B.L.

    1985-01-01

    The authors used monoclonal anti-nuclear autoantibodies and indirect immunofluorescence to examine normal human peripheral blood mononuclear leukocytes for the presence of cell surface nuclear antigens. Only one monoclonal anti-histone antibody (MH-2) was found to bind to freshly isolated PBL, staining approximately 10% of large cells. However, after cells were placed into culture for 16-24 h, a high percentage (up to 60%) of large-sized cells were recognized by an anti-DNA (BWD-1) and several different antihistone monoclonal antibodies (BWH-1, MH-1, and MH-2). These antibodies recognize separate antigenic determinants on chromatin and histones extracted from chromatin. The histone antigen-positive cells were viable, and the monoclonal antibodies could be shown to be binding to the cell surface and not to the nucleus. Using monoclonal antibodies specific for monocytes and T cells, and complement-mediated cytotoxicity, the cells bearing histone antigens were shown to be primarily monocytes. The appearance of histone and DNA antigen-positive cells was nearly completely inhibited by the addition of low concentrations of cycloheximide at initiation of the cultures. In contrast, little effect on the percentage of positive cells was detected if cells were exposed to high doses of gamma irradiation before culture. These data further support the existence of cell surface nuclear antigens on selected cell subsets, which may provide insight into the immunopathogenesis of systemic lupus erythematosus and related autoimmune diseases

  15. Shared and non-shared antigens from three different extracts of the metacestode of Echinococcus granulosus

    Directory of Open Access Journals (Sweden)

    David Carmena

    2005-12-01

    Full Text Available Hydatid cyst fluid (HCF, somatic antigens (S-Ag and excretory-secretory products (ES-Ag of Echinococcus granulosus protoscoleces are used as the main antigenic sources for immunodiagnosis of human and dog echinococcosis. In order to determine their non-shared as well as their shared antigenic components, these extracts were studied by ELISA-inhibition and immunoblot-inhibition. Assays were carried out using homologous rabbit polyclonal antisera, human sera from individuals with surgically confirmed hydatidosis, and sera from dogs naturally infected with E. granulosus. High levels of cross-reactivity were observed for all antigenic extracts, but especially for ES-Ag and S-Ag. Canine antibodies evidenced lesser avidity for their specific antigens than antibodies from human origin. The major antigenic components shared by HCF, S-Ag, and ES-Ag have apparent molecular masses of 4-6, 20-24, 52, 80, and 100-104 kDa, including doublets of 41/45, 54/57, and 65/68 kDa. Non-shared polypeptides of each antigenic extract of E. granulosus were identified, having apparent masses of 108 and 78 kDa for HCF, of 124, 94, 83, and 75 kDa for S-Ag, and of 89, 66, 42, 39, 37, and 35 kDa for ES-Ag.

  16. A midgut lysate of the Riptortus pedestris has antibacterial activity against LPS O-antigen-deficient Burkholderia mutants.

    Science.gov (United States)

    Jang, Ho Am; Seo, Eun Sil; Seong, Min Young; Lee, Bok Luel

    2017-02-01

    Riptortus pedestris, a common pest in soybean fields, harbors a symbiont Burkholderia in a specialized posterior midgut region of insects. Every generation of second nymphs acquires new Burkholderia cells from the environment. We compared in vitro cultured Burkholderia with newly in vivo colonized Burkholderia in the host midgut using biochemical approaches. The bacterial cell envelope of in vitro cultured and in vivo Burkholderia differed in structure, as in vivo bacteria lacked lipopolysaccharide (LPS) O-antigen. The LPS O-antigen deficient bacteria had a reduced colonization rate in the host midgut compared with that of the wild-type Burkholderia. To determine why LPS O-antigen-deficient bacteria are less able to colonize the host midgut, we examined in vitro survival rates of three LPS O-antigen-deficient Burkholderia mutants and lysates of five different midgut regions. The LPS O-antigen-deficient mutants were highly susceptible when cultured with the lysate of a specific first midgut region (M1), indicating that the M1 lysate contains unidentified substance(s) capable of killing LPS O-antigen-deficient mutants. We identified a 17 kDa protein from the M1 lysate, which was enriched in the active fractions. The N-terminal sequence of the protein was determined to be a soybean Kunitz-type trypsin inhibitor. These data suggest that the 17 kDa protein, which was originated from a main soybean source of the R. pedestris host, has antibacterial activity against the LPS O-antigen deficient (rough-type) Burkholderia. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Chemoselective ligation and antigen vectorization.

    Science.gov (United States)

    Gras-Masse, H

    2001-01-01

    The interest in cocktail-lipopeptide vaccines has now been confirmed by phase I clinical trials: highly diversified B-, T-helper or cytotoxic T-cell epitopes can be combined with a lipophilic vector for the induction of B- and T-cell responses of predetermined specificity. With the goal of producing an improved vaccine that should ideally induce a multispecific response in non-selected populations, increasing the diversity of the immunizing mixture represents one of the most obvious strategies.The selective delivery of antigens to professional antigen-presenting cells represents another promising approach for the improvement of vaccine efficacy. In this context, the mannose-receptor represents an attractive entry point for the targeting to dendritic cells of antigens linked to clustered glycosides or glycomimetics. In all cases, highly complex but fully characterized molecules must be produced. To develop a modular and flexible strategy which could be generally applicable to a large set of peptide antigens, we elected to explore the potentialities of chemoselective ligation methods. The hydrazone bond was found particularly reliable and fully compatible with sulphide ligation. Hydrazone/thioether orthogonal ligation systems could be developed to account for the nature of the antigens and the solubility of the vector systems. Copyright 2001 The International Association for Biologicals.

  18. Isolation and characterization of two antigenic glycoproteins from the pollen of Prosopis juliflora.

    Science.gov (United States)

    Thakur, I S

    1991-03-01

    Two antigenically active glycoprotein fractions were isolated from crude extract of the pollen of Prosopis juliflora using DEAE-cellulose ion exchange chromatography. The glycoproteins gave single band on polyacrylamide gel electrophoresis. The molecular weight of these two glycoprotein was 20,000 and 10,000 as determined by gel filtration on Sephadex G-75. With the help of crossed immunoelectrophoresis and gel diffusion crude extract exhibited twelve and three precipitating antigens suggesting its heterogeneous nature; and the purified glycoprotein fractions however formed single precipitin band on gel diffusion test and immunoelectrophoresis. As tested by ELISA the polyclonal antisera raised in rabbit showed strong binding affinity with glycoprotein of MW 20,000. These result indicates that the two glycoprotein fractions are not antigenically identical.

  19. Antibody-antigen-adjuvant conjugates enable co-delivery of antigen and adjuvant to dendritic cells in cis but only have partial targeting specificity.

    Directory of Open Access Journals (Sweden)

    Martin Kreutz

    Full Text Available Antibody-antigen conjugates, which promote antigen-presentation by dendritic cells (DC by means of targeted delivery of antigen to particular DC subsets, represent a powerful vaccination approach. To ensure immunity rather than tolerance induction the co-administration of a suitable adjuvant is paramount. However, co-administration of unlinked adjuvant cannot ensure that all cells targeted by the antibody conjugates are appropriately activated. Furthermore, antigen-presenting cells (APC that do not present the desired antigen are equally strongly activated and could prime undesired responses against self-antigens. We, therefore, were interested in exploring targeted co-delivery of antigen and adjuvant in cis in form of antibody-antigen-adjuvant conjugates for the induction of anti-tumour immunity. In this study, we report on the assembly and characterization of conjugates consisting of DEC205-specific antibody, the model antigen ovalbumin (OVA and CpG oligodeoxynucleotides (ODN. We show that such conjugates are more potent at inducing cytotoxic T lymphocyte (CTL responses than control conjugates mixed with soluble CpG. However, our study also reveals that the nucleic acid moiety of such antibody-antigen-adjuvant conjugates alters their binding and uptake and allows delivery of the antigen and the adjuvant to cells partially independently of DEC205. Nevertheless, antibody-antigen-adjuvant conjugates are superior to antibody-free antigen-adjuvant conjugates in priming CTL responses and efficiently induce anti-tumour immunity in the murine B16 pseudo-metastasis model. A better understanding of the role of the antibody moiety is required to inform future conjugate vaccination strategies for efficient induction of anti-tumour responses.

  20. Vascular expression of endothelial antigen PAL-E indicates absence of blood-ocular barriers in the normal eye

    NARCIS (Netherlands)

    Schlingemann, R. O.; Hofman, P.; Anderson, L.; Troost, D.; van der Gaag, R.

    1997-01-01

    The endothelium-specific antigen PAL-E is expressed in capillaries and veins throughout the body with the exception of the brain, where the antigen is absent from anatomical sites with a patent blood-brain barrier. In this study we determined vascular endothelial staining for PAL-E in the normal eye

  1. Cell-surface antigens associated with dualtropic and thymotropic murine leukemia viruses inducing thymic and nonthymic lymphomas

    International Nuclear Information System (INIS)

    Haas, M.; Patch, V.

    1980-01-01

    Unique type-specific antigens were detected on cells infected with dualtropic and thymotropic viruses and x-ray-induced T cell- and B cell-malignant lymphomas of C57BL/6 mice. These findings support the contention that T cell lymphoma (TCL)-inducing and B cell lymphoma (BCL)-indcing viruses isolated from x-irradiated C57BL/6 mice are env gene recombinants in which ecotropic gene sequences have been substituted by xenotropic sequences. We found that unique antigenicities are associated with each TCL-inducing and BCL-inducing dualtropic virus, and that the thymotropic TCL-inducing virus isolates represent a separate serologic group. These virus mapping experiments indicated that many serologically different recombinant viruses can be isolated from C57BL/6 mice. It is suggested that many distinct recombinant viruses may exist in lymphomagenic C57BL/6 mice, some of which are associated with specific lyphoma induction

  2. Magnesium Presence Prevents Removal of Antigenic Nuclear-Associated Proteins from Bovine Pericardium for Heart Valve Engineering.

    Science.gov (United States)

    Dalgliesh, Ailsa J; Liu, Zhi Zhao; Griffiths, Leigh G

    2017-07-01

    Current heart valve prostheses are associated with significant complications, including aggressive immune response, limited valve life expectancy, and inability to grow in juvenile patients. Animal derived "tissue" valves undergo glutaraldehyde fixation to mask tissue antigenicity; however, chronic immunological responses and associated calcification still commonly occur. A heart valve formed from an unfixed bovine pericardium (BP) extracellular matrix (ECM) scaffold, in which antigenic burden has been eliminated or significantly reduced, has potential to overcome deficiencies of current bioprostheses. Decellularization and antigen removal methods frequently use sequential solutions extrapolated from analytical chemistry approaches to promote solubility and removal of tissue components from resultant ECM scaffolds. However, the extent to which such prefractionation strategies may inhibit removal of antigenic tissue components has not been explored. We hypothesize that presence of magnesium in prefractionation steps causes DNA precipitation and reduces removal of nuclear-associated antigenic proteins. Keeping all variables consistent bar the addition or absence of magnesium (2 mM magnesium chloride hexahydrate), residual BP ECM scaffold antigenicity and removed antigenicity were assessed, along with residual and removed DNA content, ECM morphology, scaffold composition, and recellularization potential. Furthermore, we used proteomic methods to determine the mechanism by which magnesium presence or absence affects scaffold residual antigenicity. This study demonstrates that absence of magnesium from antigen removal solutions enhances solubility and subsequent removal of antigenic nuclear-associated proteins from BP. We therefore conclude that the primary mechanism of action for magnesium removal during antigen removal processes is avoidance of DNA precipitation, facilitating solubilization and removal of nuclear-associated antigenic proteins. Future studies are

  3. Prevalence of hepatitis b virus surface antigens (HBsag) and ...

    African Journals Online (AJOL)

    The prevalences of hepatitis B virus surface antigen (HBsAg) and hepatitis C virus (HCV) antibodies were determined in 560 blood donors sera using ELISA kits (DIALAB., Austria). Forty eight (8.57%) of these were positive for hepatitis B virus infection, while 33(5.89%) were positive to hepatitis C virus antibodies. The sex ...

  4. [Search for protective antigens in Ixodes persulcatus (ixodidae) salivary gland extracts].

    Science.gov (United States)

    Shtannikov, A V; Reshetniak, T V; Repolovskaia, T V; Panfertsev, E A; Perovskaia, O N; Gutova, V P; Vasil'eva, I S; Ershova, A S; Prilipov, A G; Biketov, S F; Zeidner, N

    2010-01-01

    RT-PCR evaluation of the activity of eight Ixodes persulcatus salivary gland genes shows clear distinctions in their expression depending of the stage of tick feeding. Out of them, only Salp 10 and Salp 15 proteins may be regarded as candidates for protective antigens to develop anti-tick and anti-Borrelia vaccines. Firstly they play an important role in feeding a tick and modifying a host's immune response. Secondly, the increasing expression of the salp 10 and salp 10 genes begins at early tick feeding stages. Thirdly, the activity of these genes increases with the beginning of feeding by tens and hundreds times and keeps at this level until the third tick feeding stage is over.

  5. The role of Plasmodium falciparum variant surface antigens in protective immunity and vaccine development

    DEFF Research Database (Denmark)

    Hviid, Lars

    2010-01-01

    There is substantial immuno-epidemiological evidence that the parasite-encoded, so-called variant surface antigens (VSAs) such as PfEMP1 on the surface of infected erythrocytes (IEs) are important-in some cases probably decisive-determinants of clinical outcome of P. falciparum malaria. The evide...... of VSAs, and how vaccines based on this type of antigens fit into the current global strategy to reduce, eliminate and eventually eradicate the burden of malaria....

  6. Exposure to Sarcocystis spp. in horses from Spain determined by Western blot analysis using Sarcocystis neurona merozoites as heterologous antigen.

    Science.gov (United States)

    Arias, M; Yeargan, M; Francisco, I; Dangoudoubiyam, S; Becerra, P; Francisco, R; Sánchez-Andrade, R; Paz-Silva, A; Howe, D K

    2012-04-30

    Horses serve as an intermediate host for several species of Sarcocystis, all of which utilize canids as the definitive host. Sarcocystis spp. infection and formation of latent sarcocysts in horses often appears to be subclinical, but morbidity can occur, especially when the parasite burden is large. A serological survey was conducted to determine the presence of antibodies against Sarcocystis spp. in seemingly healthy horses from the Galicia region of Spain. Western blot analyses using Sarcocystis neurona merozoites as heterologous antigen suggested greater than 80% seroprevalance of Sarcocystis spp. in a sample set of 138 horses. The serum samples were further tested with enzyme-linked immunosorbent assays (ELISAs) based on recombinant S. neurona-specific surface antigens (rSnSAGs). As expected for horses from the Eastern Hemisphere, less than 4% of the serum samples were positive when analyzed with either the rSnSAG2 or the rSnSAG4/3 ELISAs. An additional 246 horses were tested using the rSnSAG2 ELISA, which revealed that less than 3% of the 384 samples were seropositive. Collectively, the results of this serologic study suggested that a large proportion of horses from this region of Spain are exposed to Sarcocystis spp. Furthermore, the anti-Sarcocystis seroreactivity in these European horses could be clearly distinguished from anti-S. neurona antibodies using the rSnSAG2 and rSnSAG4/3 ELISAs. Copyright © 2011 Elsevier B.V. All rights reserved.

  7. Prostate-specific antigen velocity is not better than total prostate-specific antigen in predicting prostate biopsy diagnosis.

    Science.gov (United States)

    Gorday, William; Sadrzadeh, Hossein; de Koning, Lawrence; Naugler, Christopher T

    2015-12-01

    1.) Identify whether prostate-specific antigen velocity improves the ability to predict prostate biopsy diagnosis. 2.) Test whether there is an increase in the predictive capability of models when Gleason 7 prostate cancers are separated into a 3+4 and a 4+3 group. Calgary Laboratory Services' Clinical Laboratory Information System was searched for prostate biopsies reported between January 1, 2009 and December 31, 2013. Total prostate-specific antigen tests were recorded for each patient from January 1, 2007 to the most recent test before their recorded prostate biopsy. The data set was divided into the following three groups for comparison; benign, all prostate cancer and Gleason 7-10. The Gleason grade 7-10 group was further divided into 4+3 and 3+4 Gleason 7 prostate cancers. Prostate-specific antigen velocity was calculated using four different methods found in the literature. Receiver operator curves were used to assess operational characteristics of the tests. 4622 men between the ages of 40-89 with a prostate biopsy were included for analysis. Combining prostate-specific antigen velocity with total prostate-specific antigen (AUC=0.570-0.712) resulted in small non-statistically significant changes to the area under the curve compared to the area under the curve of total prostate-specific antigen alone (AUC=0.572-0.699). There were marked increases in the area under curves when 3+4 and 4+3 Gleason 7 cancers were separated. Prostate-specific antigen velocity does not add predictive value for prostate biopsy diagnosis. The clinical significance of the prostate specific antigen test can be improved by separating Gleason 7 prostate cancers into a 3+4 and 4+3 group. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  8. Overview of Plant-Made Vaccine Antigens against Malaria

    Directory of Open Access Journals (Sweden)

    Marina Clemente

    2012-01-01

    Full Text Available This paper is an overview of vaccine antigens against malaria produced in plants. Plant-based expression systems represent an interesting production platform due to their reduced manufacturing costs and high scalability. At present, different Plasmodium antigens and expression strategies have been optimized in plants. Furthermore, malaria antigens are one of the few examples of eukaryotic proteins with vaccine value expressed in plants, making plant-derived malaria antigens an interesting model to analyze. Up to now, malaria antigen expression in plants has allowed the complete synthesis of these vaccine antigens, which have been able to induce an active immune response in mice. Therefore, plant production platforms offer wonderful prospects for improving the access to malaria vaccines.

  9. Interpretation of sequential measurements of cancer antigen 125 (CA 125), carcinoembryonic antigen (CEA), and tissue polypeptide antigen (TPA) based on analytical imprecision and biological variation in the monitoring of ovarian cancer

    DEFF Research Database (Denmark)

    Tuxen, Malgorzata K.; Sölétormos, G; Petersen, P H

    2001-01-01

    The main objective with cancer antigen 125 (CA 125), carcinoembryonic antigen (CEA), and tissue polypeptide antigen (TPA) monitoring of ovarian cancer patients is to detect an early change of disease activity with high reliability. We hypothesized that a monitoring scheme for ovarian cancer patie...

  10. Epigenetic regulation of L1CAM in endometrial carcinoma: comparison to cancer–testis (CT-X) antigens

    International Nuclear Information System (INIS)

    Schirmer, Uwe; Fiegl, Heidi; Pfeifer, Marco; Zeimet, Alain G; Müller-Holzner, Elisabeth; Bode, Peter K; Tischler, Verena; Altevogt, Peter

    2013-01-01

    L1CAM was originally identified as an adhesion molecule involved in neural development. In many human carcinomas L1CAM is over-expressed and is associated with a bad prognosis. We previously reported that L1CAM was absent in the vast majority of endometrioid endometrial carcinomas (ECs) (type 1) but was strongly expressed in the more aggressive serous and clear-cell ECs (termed type 2). The differential regulation of L1CAM in ECs is not well understood. Recent evidence suggests that it can be regulated by epigenetic mechanisms. Here we investigated the role of DNA-methylation of the L1CAM promoter for expression. We also studied the relationship to cancer testis (CT-X) antigens that co-localize with L1CAM on chromosome Xq28, a region that is often activated in human tumors. We used EC cell lines and primary tumor tissues for our analysis. For expression analysis we employed RT-PCR and Western blotting. DNA-Methylation of the L1CAM promoter was determined after bisulfite conversation and DNA sequencing. Tumor tissues were examined by immunohistochemical (IHC) staining. We demonstrate that the treatment of L1CAM low/negative expressing EC cell lines with 5 ′ -Azacytidine (5-AzaC) or knock-down of DNMT1 (DNA methyltransferase 1) as well as the HDAC (histone deacetylase) inhibitor Trichostatin A (TSA) up-regulated L1CAM at the mRNA and protein level. The L1CAM gene has two promoter regions with two distinct CpG islands. We observed that the expression of L1CAM correlated with hypermethylation in promoter 1 and 5-AzaC treatment affected the DNA-methylation pattern in this region. The CT-X antigens NY-ESO-1, MAGE-A3 and MAGE-A4 were also strongly up-regulated by 5-AzaC or knock-down of DNMT1 but did not respond to treatment with TSA. Primary EC tumor tissues showed a variable methylation pattern of the L1CAM promoter. No striking differences in promoter methylation were observed between tumor areas with L1CAM expression and those without expression. L1CAM expression

  11. Alternative haplotypes of antigen processing genes in zebrafish diverged early in vertebrate evolution

    Science.gov (United States)

    McConnell, Sean C.; Hernandez, Kyle M.; Wcisel, Dustin J.; Kettleborough, Ross N.; Stemple, Derek L.; Andrade, Jorge; de Jong, Jill L. O.

    2016-01-01

    Antigen processing and presentation genes found within the MHC are among the most highly polymorphic genes of vertebrate genomes, providing populations with diverse immune responses to a wide array of pathogens. Here, we describe transcriptome, exome, and whole-genome sequencing of clonal zebrafish, uncovering the most extensive diversity within the antigen processing and presentation genes of any species yet examined. Our CG2 clonal zebrafish assembly provides genomic context within a remarkably divergent haplotype of the core MHC region on chromosome 19 for six expressed genes not found in the zebrafish reference genome: mhc1uga, proteasome-β 9b (psmb9b), psmb8f, and previously unknown genes psmb13b, tap2d, and tap2e. We identify ancient lineages for Psmb13 within a proteasome branch previously thought to be monomorphic and provide evidence of substantial lineage diversity within each of three major trifurcations of catalytic-type proteasome subunits in vertebrates: Psmb5/Psmb8/Psmb11, Psmb6/Psmb9/Psmb12, and Psmb7/Psmb10/Psmb13. Strikingly, nearby tap2 and MHC class I genes also retain ancient sequence lineages, indicating that alternative lineages may have been preserved throughout the entire MHC pathway since early diversification of the adaptive immune system ∼500 Mya. Furthermore, polymorphisms within the three MHC pathway steps (antigen cleavage, transport, and presentation) are each predicted to alter peptide specificity. Lastly, comparative analysis shows that antigen processing gene diversity is far more extensive than previously realized (with ancient coelacanth psmb8 lineages, shark psmb13, and tap2t and psmb10 outside the teleost MHC), implying distinct immune functions and conserved roles in shaping MHC pathway evolution throughout vertebrates. PMID:27493218

  12. Crystal structure of the anti-(carcinoembryonic antigen) single-chain Fv antibody MFE-23 and a model for antigen binding based on intermolecular contacts.

    Science.gov (United States)

    Boehm, M K; Corper, A L; Wan, T; Sohi, M K; Sutton, B J; Thornton, J D; Keep, P A; Chester, K A; Begent, R H; Perkins, S J

    2000-03-01

    MFE-23 is the first single-chain Fv antibody molecule to be used in patients and is used to target colorectal cancer through its high affinity for carcinoembryonic antigen (CEA), a cell-surface member of the immunoglobulin superfamily. MFE-23 contains an N-terminal variable heavy-chain domain joined by a (Gly(4)Ser)(3) linker to a variable light-chain (V(L)) domain (kappa chain) with an 11-residue C-terminal Myc-tag. Its crystal structure was determined at 2.4 A resolution by molecular replacement with an R(cryst) of 19.0%. Five of the six antigen-binding loops, L1, L2, L3, H1 and H2, conformed to known canonical structures. The sixth loop, H3, displayed a unique structure, with a beta-hairpin loop and a bifurcated apex characterized by a buried Thr residue. In the crystal lattice, two MFE-23 molecules were associated back-to-back in a manner not seen before. The antigen-binding site displayed a large acidic region located mainly within the H2 loop and a large hydrophobic region within the H3 loop. Even though this structure is unliganded within the crystal, there is an unusually large region of contact between the H1, H2 and H3 loops and the beta-sheet of the V(L) domain of an adjacent molecule (strands DEBA) as a result of intermolecular packing. These interactions exhibited remarkably high surface and electrostatic complementarity. Of seven MFE-23 residues predicted to make contact with antigen, five participated in these lattice contacts, and this model for antigen binding is consistent with previously reported site-specific mutagenesis of MFE-23 and its effect on CEA binding.

  13. Jaffe-Campanacci syndrome, revisited: detailed clinical and molecular analyses determine whether patients have neurofibromatosis type 1, coincidental manifestations, or a distinct disorder

    NARCIS (Netherlands)

    Stewart, Douglas R.; Brems, Hilde; Gomes, Alicia G.; Ruppert, Sarah L.; Callens, Tom; Williams, Jennifer; Claes, Kathleen; Bober, Michael B.; Hachen, Rachel; Kaban, Leonard B.; Li, Hua; Lin, Angela; McDonald, Marie; Melancon, Serge; Ortenberg, June; Radtke, Heather B.; Samson, Ignace; Saul, Robert A.; Shen, Joseph; Siqveland, Elizabeth; Toler, Tomi L.; van Maarle, Merel; Wallace, Margaret; Williams, Misti; Legius, Eric; Messiaen, Ludwine

    2014-01-01

    "Jaffe-Campanacci syndrome" describes the complex of multiple nonossifying fibromas of the long bones, mandibular giant cell lesions, and café-au-lait macules in individuals without neurofibromas. We sought to determine whether Jaffe-Campanacci syndrome is a distinct genetic entity or a variant of

  14. Prostate specific antigen velocity does not aid prostate cancer detection in men with prior negative biopsy.

    Science.gov (United States)

    Vickers, Andrew J; Wolters, Tineke; Savage, Caroline J; Cronin, Angel M; O'Brien, M Frank; Roobol, Monique J; Aus, Gunnar; Scardino, Peter T; Hugosson, Jonas; Schröder, Fritz H; Lilja, Hans

    2010-09-01

    Prostate specific antigen velocity has been proposed as a marker to aid in prostate cancer detection. We determined whether prostate specific antigen velocity could predict repeat biopsy results in men with persistently increased prostate specific antigen after initial negative biopsy. We identified 1,837 men who participated in the Göteborg or Rotterdam section of the European Randomized Screening study of Prostate Cancer and who underwent 1 or more subsequent prostate biopsies after an initial negative finding. We evaluated whether prostate specific antigen velocity improved predictive accuracy beyond that of prostate specific antigen alone. Of the 2,579 repeat biopsies 363 (14%) were positive for prostate cancer, of which 44 (1.7%) were high grade (Gleason score 7 or greater). Prostate specific antigen velocity was statistically associated with cancer risk but had low predictive accuracy (AUC 0.55, p <0.001). There was some evidence that prostate specific antigen velocity improved AUC compared to prostate specific antigen for high grade cancer. However, the small increase in risk associated with high prostate specific antigen velocity (from 1.7% to 2.8% as velocity increased from 0 to 1 ng/ml per year) had questionable clinical relevance. Men with prior negative biopsy are at lower risk for prostate cancer at subsequent biopsies with high grade disease particularly rare. We found little evidence to support prostate specific antigen velocity to aid in decisions about repeat biopsy for prostate cancer. 2010 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  15. Ultra-deep sequencing reveals high prevalence and broad structural diversity of hepatitis B surface antigen mutations in a global population.

    Science.gov (United States)

    Gencay, Mikael; Hübner, Kirsten; Gohl, Peter; Seffner, Anja; Weizenegger, Michael; Neofytos, Dionysios; Batrla, Richard; Woeste, Andreas; Kim, Hyon-Suk; Westergaard, Gaston; Reinsch, Christine; Brill, Eva; Thu Thuy, Pham Thi; Hoang, Bui Huu; Sonderup, Mark; Spearman, C Wendy; Pabinger, Stephan; Gautier, Jérémie; Brancaccio, Giuseppina; Fasano, Massimo; Santantonio, Teresa; Gaeta, Giovanni B; Nauck, Markus; Kaminski, Wolfgang E

    2017-01-01

    The diversity of the hepatitis B surface antigen (HBsAg) has a significant impact on the performance of diagnostic screening tests and the clinical outcome of hepatitis B infection. Neutralizing or diagnostic antibodies against the HBsAg are directed towards its highly conserved major hydrophilic region (MHR), in particular towards its "a" determinant subdomain. Here, we explored, on a global scale, the genetic diversity of the HBsAg MHR in a large, multi-ethnic cohort of randomly selected subjects with HBV infection from four continents. A total of 1553 HBsAg positive blood samples of subjects originating from 20 different countries across Africa, America, Asia and central Europe were characterized for amino acid variation in the MHR. Using highly sensitive ultra-deep sequencing, we found 72.8% of the successfully sequenced subjects (n = 1391) demonstrated amino acid sequence variation in the HBsAg MHR. This indicates that the global variation frequency in the HBsAg MHR is threefold higher than previously reported. The majority of the amino acid mutations were found in the HBV genotypes B (28.9%) and C (25.4%). Collectively, we identified 345 distinct amino acid mutations in the MHR. Among these, we report 62 previously unknown mutations, which extends the worldwide pool of currently known HBsAg MHR mutations by 22%. Importantly, topological analysis identified the "a" determinant upstream flanking region as the structurally most diverse subdomain of the HBsAg MHR. The highest prevalence of "a" determinant region mutations was observed in subjects from Asia, followed by the African, American and European cohorts, respectively. Finally, we found that more than half (59.3%) of all HBV subjects investigated carried multiple MHR mutations. Together, this worldwide ultra-deep sequencing based genotyping study reveals that the global prevalence and structural complexity of variation in the hepatitis B surface antigen have, to date, been significantly underappreciated.

  16. Structure of antigenetic determinants in the amino-terminal region of bovine fibrinogen Aα chain

    International Nuclear Information System (INIS)

    Tanswell, P.; Reiter, H.; Timpl, R.

    1978-01-01

    A radioimmunoassay was developed for peptide F-CB1α from the amino of bovine fibrinogen Aα chain, isolated after reduction and carboxymethylation of the multichain disulfide-linked cyanogen bromide peptide F-CB1. Seven out of twelve different rabbit antisera produced against fibrinogen, peptide F-CB1 or Aα chain showed distinct binding to 125 I-labelled F-CB1α. Thrombin cleavage of F-CB1α yielded two fragments: fibrinopeptide A (residues 1-19) and the carboxy-terminal fragment Th2 (residues 20-54). Antisera could be classified into three groups according to whether they recognized antigenic determinants on fibrinopeptide A, on peptide Th2 or as they showed diminished reactions with both fragments. Only little or no cross-reaction was observed with the amino-terminal cyanogen bromide peptides of Bβ and γ chain. Proteolytic fragments of fibrinopeptide A were isolated and tested for inhibitory activity with two antisera. One antiserum contained anitbodies binding selectively to the amino-terminal sequence (residues 4-11) and did not cross-react with human fibrinopeptide A. Another antiserum showed a specific binding restricted to the carboxy-terminal sequence (residues 11-18) and cross-reacted completely with human fibrinopeptide A. These results correlate well with the primary structures of the two fibrinopeptides. The antigenic activity of the peptide fragment Th2 was localized on a 15-residue tryptic peptide derived from the central portion of the sequence. These and further data indicate that at least six different antigenic determinants are present in peptide F-CB1α. (orig.) 891 AJ [de

  17. Epitope mapping of the carcinoembryonic antigen by monoclonal antibodies and establishment of a new improved radioimmunoassay system

    International Nuclear Information System (INIS)

    Kuroki, Masahide; Arakawa, Fumiko; Matsunaga, Akira; Okamoto, Naomi; Takakura, Kyoko; Matsuoka, Yuji; Higuchi, Hiroshi.

    1987-01-01

    A comprehensive mapping of epitopes on the carcinoembryonic antigen (CEA) molecule has been achieved by analyses of the specificities of 146 monoclonal antibodies (MAbs) from more than 300 hybridomas established recently. The reactivities of MAbs were analyzed by radio-immunoassays (RIA) with highly purified preparations of CEA and related antigens including normal fecal antigen-1 (NFA-1), NFA-2 in normal adult feces, nonspecific cross-reacting antigen (NCA) in lung and NCA-2 in meconium. The MAbs could be divided into five groups: group I, 23 clones directed to the NCA-common part of the CEA molecule; group II, 31 clones directed to the normal fecal cross-reacting antigen (NFCA)-common part; group III, 46 clones directed to the NFA-1-common part; group IV, 33 clones reactive with the heterogeneous carbohydrate part; and group V, 13 clones directed to the CEA-distinctive part which seemed to be highly specific for CEA. Mutual inhibitions of CEA binding between MAbs of the individual groups revealed that at least 25 different subgroups can be defined i.e., 4, 7, 8, 4, and 2 subgroups in groups I to V, respectively. The epitopes recognized by the group IV MAbs were found to be sensitive to oxidation with periodate, while the epitopes defined by MAbs of the other groups were resistant to this treatment. A solid-phase sandwich-type RIA system for CEA was established by using 2 MAbs from groups II and III as the CEA catcher and an MAb of group V as the tracer. This assay was shown to exhibit improved cancer-specificity and accuracy in the estimation of serum CEA levels. (author)

  18. Carcinoembryonic Antigen Level in Liver Disease

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Kyoo Ok; Kim, Ki Whang; Park, Chang Yun [Yonsei University College of Medicine, Seoul (Korea, Republic of)

    1978-09-15

    Carcinoembryonic antigen was initially known as tumor specific antigen and had a potential diagnostic value in the detection of digestive tract malignancies. However, subsequent studies showed CEA and CEA-like antigen present in benign disease, particularly in liver. We had collected sera from 58 patients who had liver scan and later were diagnosed clinically and histologically as liver disease. We estimated CEA values and correlations were made with liver function tests in liver cirrhosis cases. The results: 1) The raised plasma carcinoembryonic antigen level were found in 13 (68.4%) of 19 patients cirrhosis, 5 (27.8%) of 18 patients in hepatoma, 5 (71%) of 7 patients in chronic active hepatitis, all 3 patients in liver abscesses, 2 (66.7%) of 3 patients in liver abscesses, 2 (66.7%) of 3 patients in obstructive biliary disease and none in each one patient of traumatic liver hematoma, subphrenic abscess and clonorchiasis. 2) There is no linear correlation between carcinoembryonic antigen level and liver function tests including serum bilirubin, alkaline phosphatase, SGOT and prothrombin time in liver patients.

  19. Carcinoembryonic Antigen Level in Liver Disease

    International Nuclear Information System (INIS)

    Choi, Kyoo Ok; Kim, Ki Whang; Park, Chang Yun

    1978-01-01

    Carcinoembryonic antigen was initially known as tumor specific antigen and had a potential diagnostic value in the detection of digestive tract malignancies. However, subsequent studies showed CEA and CEA-like antigen present in benign disease, particularly in liver. We had collected sera from 58 patients who had liver scan and later were diagnosed clinically and histologically as liver disease. We estimated CEA values and correlations were made with liver function tests in liver cirrhosis cases. The results: 1) The raised plasma carcinoembryonic antigen level were found in 13 (68.4%) of 19 patients cirrhosis, 5 (27.8%) of 18 patients in hepatoma, 5 (71%) of 7 patients in chronic active hepatitis, all 3 patients in liver abscesses, 2 (66.7%) of 3 patients in liver abscesses, 2 (66.7%) of 3 patients in obstructive biliary disease and none in each one patient of traumatic liver hematoma, subphrenic abscess and clonorchiasis. 2) There is no linear correlation between carcinoembryonic antigen level and liver function tests including serum bilirubin, alkaline phosphatase, SGOT and prothrombin time in liver patients.

  20. Antigenic stability of pecan [Carya illinoinensis (Wangenh.) K. Koch] proteins: effects of thermal treatments and in vitro digestion.

    Science.gov (United States)

    Venkatachalam, Mahesh; Teuber, Suzanne S; Peterson, W Rich; Roux, Kenneth H; Sathe, Shridhar K

    2006-02-22

    Rabbit polyclonal antibody-based inhibition ELISA as well as immunoblotting analyses of proteins extracted from variously processed pecans (cv. Desirable) indicate that pecan proteins are antigenically stable. Pecan antigens were more sensitive to moist heat than dry heat processing treatments. SDS-PAGE and immunoblotting analysis of the native and heat-denatured proteins that were previously subjected to in vitro simulated gastric fluid digestions indicate that stable antigenic peptides were produced. Both enzyme-to-substrate ratio and digestion time were influential in determining the stability of pecan polypeptides. The stable antigenic polypeptides may serve as useful markers in developing assays suitable for the detection of trace amounts of pecans in foods.

  1. Potential Target Antigens for a Universal Vaccine in Epithelial Ovarian Cancer

    NARCIS (Netherlands)

    Vermeij, R.; Daemen, T.; de Bock, G.H.; de Graeff, P.; Leffers, N.; Lambeck, A.; Ten Hoor, K.A.; Hollema, H.; van der Zee, A.G.J.; Nijman, H.W.

    2010-01-01

    The prognosis of epithelial ovarian cancer (EOC), the primary cause of death from gynaecological malignancies, has only modestly improved over the last decades. Immunotherapeutic treatment using a cocktail of antigens has been proposed as a "universal" vaccine strategy. We determined the expression

  2. Developing antigen-specific therapies in multiple sclerosis: a tale of Tantalus or Ulysses?

    Science.gov (United States)

    van Noort JM

    1999-10-01

    Autoreactive T-cell responses directed to myelin proteins in the central nervous system are widely believed to be crucial in the pathology of multiple sclerosis (MS). However, effective ways of selectively targeting these T-cells in order to alter the clinical course of MS in a predictable manner has yet to be demonstrated. This review discusses two recent developments of crucial importance to the rational development of antigen-specific therapy in MS. The very idea of antigen-specific therapy in MS has long faced the challenge of determinant spreading, i.e., the development of novel autoimmune responses as the consequence of tissue damage. This phenomenon has led many to expect that in ongoing MS, many different pathogenic specificities would accumulate. Obviously, this would render antigen-specific therapy very difficult. Recent data now suggest that determinant spreading is most likely to be a transient phenomenon limited only to the first stages of tissue damage. A second development has changed our perspective on the specificity of individual T-cells and, thus, on the suitability of various ways to implement antigen-specific therapy. Evidence is rapidly accumulating that T-cell receptors are much more cross-reactive than previously assumed. This notion poses unexpected challenges to therapeutic approaches in MS that are based on selective targeting of autoreactive TCR. Vaccination with TCR peptides, administration of anti-TCR antibodies and development of therapeutically altered peptide ligands all depend on a significant level of predictability of pathogenic TCR. With such predictability now turning out to be much lower than was previously hoped, selective TCR-directed strategies for intervention may therefore turn out to be much less effective than anticipated. In the development of antigen-specific therapies, the use of whole protein tolerogens now seems to be the most promising route. Oral, intranasal or iv. administration of antigen remain viable options

  3. Transmission of hepatitis-B virus through salivary blood group antigens in saliva

    International Nuclear Information System (INIS)

    Meo, S.A.; Abdo, A.A.; Baksh, N.D.; Sanie, F.M.

    2010-01-01

    To determine an association between transmission of hepatitis B virus and secretor and non-secretor status of salivary blood group antigens. Study Design: Cross-sectional, analytical study. Place and Duration of Study: The Department of Physiology and Division of Hepatology, College of Medicine, King Khalid University Hospital, King Saud University, Riyadh, Kingdom of Saudi Arabia, from 2007 to 2009. Methodology: Eighty eight known patients, who were positive for Hepatitis B Surface Antigen [HBsAg] were recruited. Saliva was collected for investigating the secretor and non-secretor status by using blood typing kit number Kemtec Educational Science USA. Hepatitis B Surface antigen test was performed on Enzyme Linked Immunosorbent Assay technique. Polymerase chain reaction [PCR] on saliva was also carried out in High Performance Thermal Cycler-Palm- Cycler [Corbett Life Science, Sydney, Australia] and enzymatic amplification of extracted viral DNA was performed using primers covering the promoter of the core region of HBV. Results: Out of the 88 subjects, 61 belong to blood group O, 20 to A and 7 subjects to blood group B. Fifty subjects were secretors [salivary blood group antigens positive] and 38 subjects were non-secretors [salivary blood group antigens negative]. Among core gene positive 25 (69.4%) were secretors and 11 (30.6%) were non-secretors. However, in core gene negative 25 (48.1%) were secretors and 27 (51.9%) were non-secretors. Conclusion: The result shows an association [p=0.047] between secretor and non-secretors status of the salivary blood group antigens with core gene positive and core gene negative. (author)

  4. Detection of proliferating cell nuclear antigens and interleukin-2 beta receptor molecules on mitogen- and antigen-stimulated lymphocytes.

    Science.gov (United States)

    Hesketh, J; Dobbelaere, D; Griffin, J F; Buchan, G

    1993-01-01

    The expression of interleukin-2 receptors (IL-2R) and proliferating cell nuclear antigens (PCNA) were compared for their usefulness as markers of lymphocyte activation. Heterologous polyclonal (anti-bovine IL-2R) and monoclonal (anti-human PCNA) antibodies were used to detect the expression of these molecules on activated deer lymphocytes. Both molecules were co-expressed on blast cells which had been activated with mitogen [concanavalin A (Con A)]. There was detectable up-regulation of IL-2R expression in response to antigen [Mycobacterium bovis-derived purified protein derivative (PPD)] stimulation while PCNA expression mimicked lymphocyte transformation (LT) reactivity. PCNA expression was found to more accurately reflect both antigen- and mitogen-activated lymphocyte activation, as estimated by LT activity. The expression of PCNA was used to identify antigen reactive cells from animals exposed to M. bovis. A very low percentage (1.1 +/- 0.4%) of peripheral blood lymphocytes from non-infected animals could be stimulated to express PCNA by in vitro culture with antigen (PPD). Within the infected group both diseased and healthy, 'in-contact', animals expressed significantly higher levels of PCNA upon antigen stimulation. PMID:8104884

  5. Modeling antigen-antibody nanoparticle bioconjugates and their polymorphs

    Science.gov (United States)

    Desgranges, Caroline; Delhommelle, Jerome

    2018-03-01

    The integration of nanomaterials with biomolecules has recently led to the development of new ways of designing biosensors, and through their assembly, to new hybrid structures for novel and exciting applications. In this work, we develop a coarse-grained model for nanoparticles grafted with antibody molecules and their binding with antigens. In particular, we isolate two possible states for antigen-antibody pairs during the binding process, termed as recognition and anchoring states. Using molecular simulation, we calculate the thermodynamic and structural features of three possible crystal structures or polymorphs, the body-centered cubic, simple cubic, and face-centered cubic phases, and of the melt. This leads us to determine the domain of stability of the three solid phases. In particular, the role played by the switching process between anchoring and recognition states during melting is identified, shedding light on the complex microscopic mechanisms in these systems.

  6. Studies on antigenic cross-reactivity of Trichuris ovis with host mucosal antigens in goat

    OpenAIRE

    Gautam Patra; Seikh Sahanawaz Alam; Sonjoy Kumar Borthakur; Hridayesh Prasad

    2015-01-01

    Objective: To ascertain whether immunodominant antigens of Trichuris ovis might share and cross react with host molecule. Methods: Two crude protein preparations from anterior and posterior parts of Trichuris ovis were characterized along with host mucosal antigen by double immunodiffusion, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting technique. Conventional scanning electron microscopy was performed as per standard procedure. Results: Sharp...

  7. Proliferation induced by Plasmodium falciparum antigen and interleukin-2 production by lymphocytes isolated from malaria-immune individuals

    DEFF Research Database (Denmark)

    Theander, T G; Bygbjerg, I C; Jepsen, S

    1986-01-01

    Affinity-purified Plasmodium falciparum soluble antigens (SPAg) isolated from in vitro cultures of the parasite were shown to be relatively free of nonspecific polyclonal activators. To determine the presence of lymphocytes with specificity against SPAg in the peripheral blood of malaria-immune i......Affinity-purified Plasmodium falciparum soluble antigens (SPAg) isolated from in vitro cultures of the parasite were shown to be relatively free of nonspecific polyclonal activators. To determine the presence of lymphocytes with specificity against SPAg in the peripheral blood of malaria...

  8. Standardization and validation of a cytometric bead assay to assess antibodies to multiple Plasmodium falciparum recombinant antigens

    Directory of Open Access Journals (Sweden)

    Ondigo Bartholomew N

    2012-12-01

    Full Text Available Abstract Background Multiplex cytometric bead assay (CBA have a number of advantages over ELISA for antibody testing, but little information is available on standardization and validation of antibody CBA to multiple Plasmodium falciparum antigens. The present study was set to determine optimal parameters for multiplex testing of antibodies to P. falciparum antigens, and to compare results of multiplex CBA to ELISA. Methods Antibodies to ten recombinant P. falciparum antigens were measured by CBA and ELISA in samples from 30 individuals from a malaria endemic area of Kenya and compared to known positive and negative control plasma samples. Optimal antigen amounts, monoplex vs multiplex testing, plasma dilution, optimal buffer, number of beads required were assessed for CBA testing, and results from CBA vs. ELISA testing were compared. Results Optimal amounts for CBA antibody testing differed according to antigen. Results for monoplex CBA testing correlated strongly with multiplex testing for all antigens (r = 0.88-0.99, P values from Conclusion With optimization, CBA may be the preferred method of testing for antibodies to P. falciparum antigens, as CBA can test for antibodies to multiple recombinant antigens from a single plasma sample and produces a greater range of values in positive samples and lower background readings for blank samples than ELISA.

  9. Detection of HBs antigens and antibodies by immunoenzymology and radioimmunology. Comparison of the results

    International Nuclear Information System (INIS)

    Fauchet, R.; Genetet, B.; Phengsavath

    1981-01-01

    The respective sensitivity threshold of immunoenzymology and radioimmunology in HBs antigen and antibody detection were compared and the optical density (o.d.) and counts per minute (cpm) values were correlated with the concentration in ng/ml of a given sample. The sensitivity comparison between RIA and EIA appears in favour of the former technique for both HBs antigen and HBs antibody detection. However the procedure tried here to detect HBs antibodies by immunoenzymology is simple, requires only the choice of neutralising antigenic pool and sensitivity threshold and could usefully be generalized for teams not authorized to handle radioelements. Quantification of the HBs antigen content in ng/ml is a difficult operation, needing great care in the dilution of the sera tested. The reproducibility is not perfect. The semi-quantitative RIA determination expressed in R.U. (radioimmunological units) seems adequate as a mean to follow the progress of an HBs antibody in both liver disease and vaccination [fr

  10. Antigen specific T-cell responses against tumor antigens are controlled by regulatory T cells in patients with prostate cancer.

    Science.gov (United States)

    Hadaschik, Boris; Su, Yun; Huter, Eva; Ge, Yingzi; Hohenfellner, Markus; Beckhove, Philipp

    2012-04-01

    Immunotherapy is a promising approach in an effort to control castration resistant prostate cancer. We characterized tumor antigen reactive T cells in patients with prostate cancer and analyzed the suppression of antitumor responses by regulatory T cells. Peripheral blood samples were collected from 57 patients with histologically confirmed prostate cancer, 8 patients with benign prostatic hyperplasia and 16 healthy donors. Peripheral blood mononuclear cells were isolated and antigen specific interferon-γ secretion of isolated T cells was analyzed by enzyme-linked immunospot assay. T cells were functionally characterized and T-cell responses before and after regulatory T-cell depletion were compared. As test tumor antigens, a panel of 11 long synthetic peptides derived from a total of 8 tumor antigens was used, including prostate specific antigen and prostatic acid phosphatase. In patients with prostate cancer we noted a 74.5% effector T-cell response rate compared with only 25% in patients with benign prostatic hyperplasia and 31% in healthy donors. In most patients 2 or 3 tumor antigens were recognized. Comparing various disease stages there was a clear increase in the immune response against prostate specific antigens from intermediate to high risk tumors and castration resistant disease. Regulatory T-cell depletion led to a significant boost in effector T-cell responses against prostate specific antigen and prostatic acid phosphatase. Tumor specific effector T cells were detected in most patients with prostate cancer, especially those with castration resistant prostate cancer. Since effector T-cell responses against prostate specific antigens strongly increased after regulatory T-cell depletion, our results indicate that immunotherapy efficacy could be enhanced by decreasing regulatory T cells. Copyright © 2012 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  11. Do ABO blood group antigens hamper the therapeutic efficacy of mesenchymal stromal cells?

    Science.gov (United States)

    Moll, Guido; Hult, Annika; von Bahr, Lena; Alm, Jessica J; Heldring, Nina; Hamad, Osama A; Stenbeck-Funke, Lillemor; Larsson, Stella; Teramura, Yuji; Roelofs, Helene; Nilsson, Bo; Fibbe, Willem E; Olsson, Martin L; Le Blanc, Katarina

    2014-01-01

    Investigation into predictors for treatment outcome is essential to improve the clinical efficacy of therapeutic multipotent mesenchymal stromal cells (MSCs). We therefore studied the possible harmful impact of immunogenic ABO blood groups antigens - genetically governed antigenic determinants - at all given steps of MSC-therapy, from cell isolation and preparation for clinical use, to final recipient outcome. We found that clinical MSCs do not inherently express or upregulate ABO blood group antigens after inflammatory challenge or in vitro differentiation. Although antigen adsorption from standard culture supplements was minimal, MSCs adsorbed small quantities of ABO antigen from fresh human AB plasma (ABP), dependent on antigen concentration and adsorption time. Compared to cells washed in non-immunogenic human serum albumin (HSA), MSCs washed with ABP elicited stronger blood responses after exposure to blood from healthy O donors in vitro, containing high titers of ABO antibodies. Clinical evaluation of hematopoietic stem cell transplant (HSCT) recipients found only very low titers of anti-A/B agglutination in these strongly immunocompromised patients at the time of MSC treatment. Patient analysis revealed a trend for lower clinical response in blood group O recipients treated with ABP-exposed MSC products, but not with HSA-exposed products. We conclude, that clinical grade MSCs are ABO-neutral, but the ABP used for washing and infusion of MSCs can contaminate the cells with immunogenic ABO substance and should therefore be substituted by non-immunogenic HSA, particularly when cells are given to immunocompentent individuals.

  12. Generation and characterization of antigenic variants induced by exposure of tumor cells to UV radiation in vitro

    International Nuclear Information System (INIS)

    Hostetler, L.L.W.

    1988-01-01

    Antigenic changes present in nonantigenic tumor cells exposed to UV radiation (UV) in vitro were investigated by addressing the following questions: (1) Are antigenic variants (AV) produced that are rejected in normal but not immunosuppressed mice? (2) Does generation of AV depend upon intrinsic properties of the cells exposed or result from the action of UV? (3) Is antigenic modification induced by UV due to increased histocompatibility antigen expression? (4) Do AV crossreact immunologically with parental tumor or with other AV? and (5) Is the UV-associated common antigen expressed on UV-induced tumors present on UV-irradiated tumor cells? AV were generated at different frequencies following in vitro UV irradiation of a spontaneous murine fibrosarcoma, a murine melanoma, and two melanoma clones. Immunological experiments demonstrated that the AV and parental cells shared a determinant that was susceptible to immune recognition, but incapable of inducing immunity. In contrast, the AV were noncrossreactive, suggesting that variant-specific antigens were also expressed. Finally, the AV were recognized by UV-induced suppressor cells, indicating that the UV-associated common antigen expressed by UV-induced tumors was also present

  13. Purification and refolding of anti-T-antigen single chain antibodies (scFvs) expressed in Escherichia coli as inclusion bodies.

    Science.gov (United States)

    Yuasa, Noriyuki; Koyama, Tsubasa; Fujita-Yamaguchi, Yoko

    2014-02-01

    T-antigen (Galβ1-3GalNAcα-1-Ser/Thr) is an oncofetal antigen that is commonly expressed as a carbohydrate determinant in many adenocarcinomas. Since it is associated with tumor progression and metastasis, production of recombinant antibodies specific for T-antigen could lead to the development of cancer diagnostics and therapeutics. Previously, we isolated and characterized 11 anti-T-antigen phage clones from a phage library displaying human single-chain antibodies (scFvs) and purified one scFv protein, 1G11. More recently, we purified and characterized 1E8 scFv protein using a Drosophila S2 expression system. In the current study, four anti-T-antigen scFv genes belonging to Groups 1-4 were purified from inclusion bodies expressed in Escherichia coli cells. Inclusion bodies isolated from E. coli cells were denatured in 3.5 M Gdn-HCl. Solubilized His-tagged scFv proteins were purified using Ni(2+)-Sepharose column chromatography in the presence of 3.5 M Gdn-HCl. Purified scFv proteins were refolded according to a previously published method of step-wise dialysis. Two anti-T-antigen scFv proteins, 1E6 and 1E8 that belong to Groups 1 and 2, respectively, were produced in sufficient amounts, thus allowing further characterization of their binding activity with T-antigen. Specificity and affinity constants determined using enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR), respectively, provided evidence that both 1E8 and 1E6 scFv proteins are T-antigen specific and suggested that 1E8 scFv protein has a higher affinity for T-antigen than 1E6 scFv protein.

  14. Characterization of a human antigen specific helper factor

    International Nuclear Information System (INIS)

    Richardson, B.

    1986-01-01

    While antigen (Ag) specific helper factors have been characterized in mice, similar molecules have not been identified in humans. To characterize human antigen specific helper molecules, an IL-2 dependent tetanus toxoid (T.T.) reactive T cell line was fused with a 6-thioguanine resistant CEM line, and hybrids selected in medium containing hypoxanthine and azaserine. Hybrids were screened by culturing the cells with 35 S-Met then reacting the supernatants with T.T. or hepatitis vaccine immobilized on nitrocellulose. One hybrid, TT6BA-O, was identified which secreted a Met-containing molecule which bound T.T. but not hepatitis vaccine. Supernatants from TT6BA-O, but not the parent CEM line, when added to autologous peripheral blood mononuclear cells (PBMC's) stimulated secretion of T.T. specific antibodies (Abs). Specificity controls demonstrated that TT6BA-O supernatant did not induce antibodies to diphtheria toxoid, hepatitis vaccine or pneumococcal polysaccharide, and total immunoglobulin (lg) synthesis was minimally increased. In contrast, pokeweed mitogen stimulated significant lg synthesis as well as Ab's to pneumococcal polysaccharide and T.T. TT6BA-O supernatant induced anti-T.T.Ab's in autologous PBMC's but not PBMC's from 3 unrelated donors, suggesting that the activity of the helper factor is restricted, possibly by the MHC. The molecular weight of the helper factor was estimated at 100,000-150,000 by Sephacryl S-300 chromatography. Finally, the helper factor could be demonstrated to bind and elute from sephorose-immobilized T.T. and anti-DR antisera, but not anti-lg antisera or the T40/25 monoclonal antibody, which binds a nonpolymorphic determinant on the human T cell receptor. These results demonstrate that human Ag specific helper factors exist, bind antigen and bear class II MHC determinants

  15. The relationship between oral Candida carriage and the secretor status of blood group antigens in saliva.

    Science.gov (United States)

    Shin, Eun-Seop; Chung, Sung-Chang; Kim, Young-Ku; Lee, Sung-Woo; Kho, Hong-Seop

    2003-07-01

    The aim of the study was to investigate the relationship between oral Candida carriage and the secretor status of blood group antigens. Unstimulated whole saliva and oral rinse samples were obtained from 180 healthy subjects. These samples were plated on Sabouraud's dextrose agar media to determine oral Candida carriage. Sodium dodecylsulfate polyacrylamide gel electrophoresis and immunoblotting were performed on whole saliva samples to determine the secretor status of blood group antigens. The oral Candida carriage rate was found to be 45.0%. The sensitivity of the concentrated rinse culture proved to be superior. Oral Candida carriage was not significantly related to the blood group or secretor status of ABH or Lewis antigens. No significant relationship was found between oral Candida carriage and salivary flow rate. However, smoking affected oral Candida carriage. Oral Candida carriage in healthy individuals is not significantly related to blood group or secretor status.

  16. Biosynthetic incorporation of [75Se]selenomethionine: a new method for labelling lymphocyte membrane antigens

    International Nuclear Information System (INIS)

    Dosseto, M.; Rohner, C.; Pierres, M.; Goridis, C.

    1981-01-01

    A novel approach for radiolabelling lymphocyte membrane antigens is described. This technique is based on the use of the γ-emitting amino acid analogue [ 75 Se]selenomethionine. Human HLA-A, B, C and DR heavy and light chains and mouse Ia antigens were efficiently labelled by this technique and were precipitated with monoclonal antibodies. Approximately the same radioactivity was incorporated into the HLA-A, B, C chains whether [ 75 Se]selenomethionine, [ 35 S]methionine or [ 3 H]leucine were used as precursors. Easily detectable as a γ-emitter, [ 75 Se]selenomethionine thus constitutes a useful biosynthetic label of lymphocyte surface antigens. The same method was used to label immunoglobulins produced by hybridomas and to determine the nature of the secreted light chains. (Auth.)

  17. Antigen Loss Variants: Catching Hold of Escaping Foes.

    Science.gov (United States)

    Vyas, Maulik; Müller, Rolf; Pogge von Strandmann, Elke

    2017-01-01

    Since mid-1990s, the field of cancer immunotherapy has seen steady growth and selected immunotherapies are now a routine and preferred therapeutic option of certain malignancies. Both active and passive cancer immunotherapies exploit the fact that tumor cells express specific antigens on the cell surface, thereby mounting an immune response specifically against malignant cells. It is well established that cancer cells typically lose surface antigens following natural or therapy-induced selective pressure and these antigen-loss variants are often the population that causes therapy-resistant relapse. CD19 and CD20 antigen loss in acute lymphocytic leukemia and chronic lymphocytic leukemia, respectively, and lineage switching in leukemia associated with mixed lineage leukemia (MLL) gene rearrangements are well-documented evidences in this regard. Although increasing number of novel immunotherapies are being developed, majority of these do not address the control of antigen loss variants. Here, we review the occurrence of antigen loss variants in leukemia and discuss the therapeutic strategies to tackle the same. We also present an approach of dual-targeting immunoligand effectively retargeting NK cells against antigen loss variants in MLL-associated leukemia. Novel immunotherapies simultaneously targeting more than one tumor antigen certainly hold promise to completely eradicate tumor and prevent therapy-resistant relapses.

  18. Conservation of myeloid surface antigens on primate granulocytes.

    Science.gov (United States)

    Letvin, N L; Todd, R F; Palley, L S; Schlossman, S F; Griffin, J D

    1983-02-01

    Monoclonal antibodies reactive with myeloid cell surface antigens were used to study evolutionary changes in granulocyte surface antigens from primate species. Certain of these granulocyte membrane antigens are conserved in phylogenetically distant species, indicating the potential functional importance of these structures. The degree of conservation of these antigens reflects the phylogenetic relationship between primate species. Furthermore, species of the same genus show similar patterns of binding to this panel of anti-human myeloid antibodies. This finding of conserved granulocyte surface antigens suggests that non-human primates may provide a model system for exploring uses of monoclonal antibodies in the treatment of human myeloid disorders.

  19. Antigen Cross-Presentation of Immune Complexes

    Science.gov (United States)

    Platzer, Barbara; Stout, Madeleine; Fiebiger, Edda

    2014-01-01

    The ability of dendritic cells (DCs) to cross-present tumor antigens has long been a focus of interest to physicians, as well as basic scientists, that aim to establish efficient cell-based cancer immune therapy. A prerequisite for exploiting this pathway for therapeutic purposes is a better understanding of the mechanisms that underlie the induction of tumor-specific cytotoxic T-lymphocyte (CTL) responses when initiated by DCs via cross-presentation. The ability of humans DC to perform cross-presentation is of utmost interest, as this cell type is a main target for cell-based immunotherapy in humans. The outcome of a cross-presentation event is guided by the nature of the antigen, the form of antigen uptake, and the subpopulation of DCs that performs presentation. Generally, CD8α+ DCs are considered to be the most potent cross-presenting DCs. This paradigm, however, only applies to soluble antigens. During adaptive immune responses, immune complexes form when antibodies interact with their specific epitopes on soluble antigens. Immunoglobulin G (IgG) immune complexes target Fc-gamma receptors on DCs to shuttle exogenous antigens efficiently into the cross-presentation pathway. This receptor-mediated cross-presentation pathway is a well-described route for the induction of strong CD8+ T cell responses. IgG-mediated cross-presentation is intriguing because it permits the CD8− DCs, which are commonly considered to be weak cross-presenters, to efficiently cross-present. Engaging multiple DC subtypes for cross-presentation might be a superior strategy to boost CTL responses in vivo. We here summarize our current understanding of how DCs use IgG-complexed antigens for the efficient induction of CTL responses. Because of its importance for human cell therapy, we also review the recent advances in the characterization of cross-presentation properties of human DC subsets. PMID:24744762

  20. Rapid profiling of the antigen regions recognized by serum antibodies using massively parallel sequencing of antigen-specific libraries.

    KAUST Repository

    Domina, Maria; Lanza Cariccio, Veronica; Benfatto, Salvatore; D'Aliberti, Deborah; Venza, Mario; Borgogni, Erica; Castellino, Flora; Biondo, Carmelo; D'Andrea, Daniel; Grassi, Luigi; Tramontano, Anna; Teti, Giuseppe; Felici, Franco; Beninati, Concetta

    2014-01-01

    There is a need for techniques capable of identifying the antigenic epitopes targeted by polyclonal antibody responses during deliberate or natural immunization. Although successful, traditional phage library screening is laborious and can map only some of the epitopes. To accelerate and improve epitope identification, we have employed massive sequencing of phage-displayed antigen-specific libraries using the Illumina MiSeq platform. This enabled us to precisely identify the regions of a model antigen, the meningococcal NadA virulence factor, targeted by serum antibodies in vaccinated individuals and to rank hundreds of antigenic fragments according to their immunoreactivity. We found that next generation sequencing can significantly empower the analysis of antigen-specific libraries by allowing simultaneous processing of dozens of library/serum combinations in less than two days, including the time required for antibody-mediated library selection. Moreover, compared with traditional plaque picking, the new technology (named Phage-based Representation OF Immuno-Ligand Epitope Repertoire or PROFILER) provides superior resolution in epitope identification. PROFILER seems ideally suited to streamline and guide rational antigen design, adjuvant selection, and quality control of newly produced vaccines. Furthermore, this method is also susceptible to find important applications in other fields covered by traditional quantitative serology.

  1. Rapid profiling of the antigen regions recognized by serum antibodies using massively parallel sequencing of antigen-specific libraries.

    Directory of Open Access Journals (Sweden)

    Maria Domina

    Full Text Available There is a need for techniques capable of identifying the antigenic epitopes targeted by polyclonal antibody responses during deliberate or natural immunization. Although successful, traditional phage library screening is laborious and can map only some of the epitopes. To accelerate and improve epitope identification, we have employed massive sequencing of phage-displayed antigen-specific libraries using the Illumina MiSeq platform. This enabled us to precisely identify the regions of a model antigen, the meningococcal NadA virulence factor, targeted by serum antibodies in vaccinated individuals and to rank hundreds of antigenic fragments according to their immunoreactivity. We found that next generation sequencing can significantly empower the analysis of antigen-specific libraries by allowing simultaneous processing of dozens of library/serum combinations in less than two days, including the time required for antibody-mediated library selection. Moreover, compared with traditional plaque picking, the new technology (named Phage-based Representation OF Immuno-Ligand Epitope Repertoire or PROFILER provides superior resolution in epitope identification. PROFILER seems ideally suited to streamline and guide rational antigen design, adjuvant selection, and quality control of newly produced vaccines. Furthermore, this method is also susceptible to find important applications in other fields covered by traditional quantitative serology.

  2. Rapid profiling of the antigen regions recognized by serum antibodies using massively parallel sequencing of antigen-specific libraries.

    KAUST Repository

    Domina, Maria

    2014-12-04

    There is a need for techniques capable of identifying the antigenic epitopes targeted by polyclonal antibody responses during deliberate or natural immunization. Although successful, traditional phage library screening is laborious and can map only some of the epitopes. To accelerate and improve epitope identification, we have employed massive sequencing of phage-displayed antigen-specific libraries using the Illumina MiSeq platform. This enabled us to precisely identify the regions of a model antigen, the meningococcal NadA virulence factor, targeted by serum antibodies in vaccinated individuals and to rank hundreds of antigenic fragments according to their immunoreactivity. We found that next generation sequencing can significantly empower the analysis of antigen-specific libraries by allowing simultaneous processing of dozens of library/serum combinations in less than two days, including the time required for antibody-mediated library selection. Moreover, compared with traditional plaque picking, the new technology (named Phage-based Representation OF Immuno-Ligand Epitope Repertoire or PROFILER) provides superior resolution in epitope identification. PROFILER seems ideally suited to streamline and guide rational antigen design, adjuvant selection, and quality control of newly produced vaccines. Furthermore, this method is also susceptible to find important applications in other fields covered by traditional quantitative serology.

  3. Comparative analysis of rabbit hemorrhagic disease virus (RHDV) and new RHDV2 virus antigenicity, using specific virus-like particles

    OpenAIRE

    Bárcena, Juan; Guerra, Beatriz; Angulo, Iván; González, Julia; Valcárcel, Félix; Mata, Carlos P.; Castón, José R.; Blanco, Esther; Alejo, Alí

    2015-01-01

    International audience; In 2010 a new Lagovirus related to rabbit haemorrhagic disease virus (RHDV) emerged in France and has since rapidly spread throughout domestic and wild rabbit populations of several European countries. The new virus, termed RHDV2, exhibits distinctive genetic, antigenic and pathogenic features. Notably, RHDV2 kills rabbits previously vaccinated with RHDV vaccines. Here we report for the first time the generation and characterization of RHDV2-specific virus-like particl...

  4. Immunoglobulin M antibodies against CRA and FRA recombinant antigens of Trypanosoma cruzi in chronic chagasic patients.

    Science.gov (United States)

    Vasconcelos, Romero H T; Azevedo, Elisa A N; Cavalcanti, Maria G A M; Silva, Edimilson D; Ferreira, Antonio G P; Morais, Clarice N L; Gomes, Yara M

    2011-05-01

    Previous works of our research group have demonstrated aspects of the humoral immune response of chronic Chagas disease using the cytoplasmatic repetitive antigen (CRA) and the flagellar repetitive antigen (FRA) of Trypanosoma cruzi. The aim of this work was to analyze the presence of specific immunoglobulin M (IgM) antibodies in chronic chagasic patients using these recombinant antigens of T. cruzi. The positivity of IgM in chronic chagasic patients against CRA and FRA antigens was determined by indirect enzyme-linked immunosorbent assay. We reported no statistical significant differences between the levels of IgM for both recombinant antigens and the different chronic clinical forms of Chagas disease. However, a small proportion of chronic chagasic patients analyzed in this study was positive for this antibody isotype. The findings of this study indicate that the IgM antibodies cannot be used to elucidate the differences in the profile of humoral immune response among chronic chagasic patients with different clinical forms using the CRA and FRA recombinant antigens of T. cruzi. Copyright © 2011 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

  5. Plasma membrane associated, virus-specific polypeptides required for the formation of target antigen complexes recognized by virus-specific cytotoxic T lymphocytes

    International Nuclear Information System (INIS)

    Domber, E.A.

    1986-01-01

    These studies were undertaken to define some of the poxvirus-specific target antigens which are synthesized in infected cells and recognized by vaccinia virus-specific CTLs (VV-CTLs). Since vaccinia virus infected, unmanipulated target cells express numerous virus-specific antigens on the plasma membrane, attempts were made to manipulate expression of the poxvirus genome after infection so that one or a few defined virus-specified antigens were expressed on the surface of infected cells. In vitro [ 51 Cr]-release assays determined that viral DNA synthesis and expression of late viral proteins were not necessary to form a target cell which was fully competent for lysis by VV-CTLs. Under the conditions employed in these experiments, 90-120 minutes of viral protein synthesis were necessary to produce a competent cell for lysis by VV-CTLs. In order to further inhibit the expression of early viral proteins in infected cells, partially UV-inactivated vaccinia virus was employed to infect target cells. It was determined that L-cells infected with virus preparations which had been UV-irradiated for 90 seconds were fully competent for lysis by VV-CTLs. Cells infected with 90 second UV-irr virus expressed 3 predominant, plasma membrane associated antigens of 36-37K, 27-28K, and 19-17K. These 3 viral antigens represent the predominant membrane-associated viral antigens available for interaction with class I, major histocompatibility antigens and hence are potential target antigens for VV-CTLs

  6. Proteolysis of the heavy chain of major histocompatibility complex class I antigens by complement component C1s

    DEFF Research Database (Denmark)

    Eriksson, H; Nissen, Mogens Holst

    1990-01-01

    weights of the fragments are in agreement with the cleavage located in the area between the disulphide loops of the alpha 2-and alpha 3-domains of the heavy chain. In addition human C1s complement is able to cleave H-2 antigens from mouse in a similar fashion but not rat MHC class I antigen or mouse MHC...... class II antigen (I-Ad). Mouse MHC class I antigen-specific determinants could also be detected in supernatant from mouse spleen cells incubated with C1r and C1s. These results indicate the presence in the body fluids of a non-membrane-bound soluble form of the alpha 1-and alpha 2-domains which...

  7. Identification of immunogenic Salmonella enterica serotype Typhi antigens expressed in chronic biliary carriers of S. Typhi in Kathmandu, Nepal.

    Directory of Open Access Journals (Sweden)

    Richelle C Charles

    Full Text Available Salmonella enterica serotype Typhi can colonize and persist in the biliary tract of infected individuals, resulting in a state of asymptomatic chronic carriage. Chronic carriers may act as persistent reservoirs of infection within a community and may introduce infection to susceptible individuals and new communities. Little is known about the interaction between the host and pathogen in the biliary tract of chronic carriers, and there is currently no reliable diagnostic assay to identify asymptomatic S. Typhi carriage.To study host-pathogen interactions in the biliary tract during S. Typhi carriage, we applied an immunoscreening technique called in vivo-induced antigen technology (IVIAT, to identify potential biomarkers unique to carriers. IVIAT identifies humorally immunogenic bacterial antigens expressed uniquely in the in vivo environment, and we hypothesized that S. Typhi surviving in the biliary tract of humans may express a distinct antigenic profile. Thirteen S. Typhi antigens that were immunoreactive in carriers, but not in healthy individuals from a typhoid endemic area, were identified. The identified antigens included a number of putative membrane proteins, lipoproteins, and hemolysin-related proteins. YncE (STY1479, an uncharacterized protein with an ATP-binding motif, gave prominent responses in our screen. The response to YncE in patients whose biliary tract contained S. Typhi was compared to responses in patients whose biliary tract did not contain S. Typhi, patients with acute typhoid fever, and healthy controls residing in a typhoid endemic area. Seven of 10 (70% chronic carriers, 0 of 8 bile culture-negative controls (0%, 0 of 8 healthy Bangladeshis (0%, and 1 of 8 (12.5% Bangladeshis with acute typhoid fever had detectable anti-YncE IgG in blood. IgA responses were also present.Further evaluation of YncE and other antigens identified by IVIAT could lead to the development of improved diagnostic assays to identify asymptomatic

  8. Understanding original antigenic sin in influenza with a dynamical system.

    Science.gov (United States)

    Pan, Keyao

    2011-01-01

    Original antigenic sin is the phenomenon in which prior exposure to an antigen leads to a subsequent suboptimal immune response to a related antigen. Immune memory normally allows for an improved and rapid response to antigens previously seen and is the mechanism by which vaccination works. I here develop a dynamical system model of the mechanism of original antigenic sin in influenza, clarifying and explaining the detailed spin-glass treatment of original antigenic sin. The dynamical system describes the viral load, the quantities of healthy and infected epithelial cells, the concentrations of naïve and memory antibodies, and the affinities of naïve and memory antibodies. I give explicit correspondences between the microscopic variables of the spin-glass model and those of the present dynamical system model. The dynamical system model reproduces the phenomenon of original antigenic sin and describes how a competition between different types of B cells compromises the overall effect of immune response. I illustrate the competition between the naïve and the memory antibodies as a function of the antigenic distance between the initial and subsequent antigens. The suboptimal immune response caused by original antigenic sin is observed when the host is exposed to an antigen which has intermediate antigenic distance to a second antigen previously recognized by the host's immune system.

  9. Distinct Phosphorylation Clusters Determine the Signaling Outcome of Free Fatty Acid Receptor 4/G Protein-Coupled Receptor 120

    DEFF Research Database (Denmark)

    Prihandoko, Rudi; Alvarez-Curto, Elisa; Hudson, Brian D

    2016-01-01

    of these phosphoacceptor sites to alanine completely prevented phosphorylation of mFFA4 but did not limit receptor coupling to extracellular signal regulated protein kinase 1 and 2 (ERK1/2) activation. Rather, an inhibitor of Gq/11proteins completely prevented receptor signaling to ERK1/2. By contrast, the recruitment...... activation. These unique observations define differential effects on signaling mediated by phosphorylation at distinct locations. This hallmark feature supports the possibility that the signaling outcome of mFFA4 activation can be determined by the pattern of phosphorylation (phosphorylation barcode...

  10. Prevalence of human immunodeficiency virus type 1 p24 antigen in U.S. blood donors--an assessment of the efficacy of testing in donor screening. The HIV-Antigen Study Group.

    Science.gov (United States)

    Alter, H J; Epstein, J S; Swenson, S G; VanRaden, M J; Ward, J W; Kaslow, R A; Menitove, J E; Klein, H G; Sandler, S G; Sayers, M H

    1990-11-08

    We performed a multicenter study in 1989 to determine whether screening whole-blood donors for human immunodeficiency virus type 1 (HIV-1) p24 antigen would improve transfusion safety by identifying carriers of the virus who are seronegative for HIV-1 antibody. More than 500,000 donations were tested at 13 U.S. blood centers with test kits from two manufacturers. Units found repeatedly reactive were retested in a central laboratory; if the results were positive, they were confirmed by a neutralization assay. A subgroup of units was also tested for HIV-1 by the polymerase chain reaction. Selected donors confirmed or not confirmed as having p24 antigen were contacted for follow-up interviews to identify risk factors and undergo retesting for HIV-1 markers. Positive tests for p24 antigen were confirmed by neutralization in five donors (0.001 percent of all donations tested), all of whom were also positive for HIV-1 antibody and HIV-1 by polymerase chain reaction. Three of the antigen-positive donors had other markers of infectious disease that would have resulted in the exclusion of their blood; two had risk factors for HIV-1 that should have led to self-exclusion. Of 220 blood units with repeatedly reactive p24 antigen whose presence could not be confirmed by neutralization (0.04 percent of the donations studied), none were positive for HIV-1 antibody, HIV-1 by polymerase chain reaction (120 units tested), or virus culture (76 units tested)--attesting to the specificity of confirmatory neutralization. The finding that no donation studied was positive for p24 antigen and negative for HIV-1 antibody suggests that screening donors for p24 antigen with tests of the current level of sensitivity would not add substantially to the safety of the U.S. blood supply.

  11. Protein antigenic structures recognized by T cells: potential applications to vaccine design.

    Science.gov (United States)

    Berzofsky, J A; Cease, K B; Cornette, J L; Spouge, J L; Margalit, H; Berkower, I J; Good, M F; Miller, L H; DeLisi, C

    1987-08-01

    In summary, our results using the model protein antigen myoglobin indicated, in concordance with others, that helper T lymphocytes recognize a limited number of immunodominant antigenic sites of any given protein. Such immunodominant sites are the focus of a polyclonal response of a number of different T cells specific for distinct but overlapping epitopes. Therefore, the immunodominance does not depend on the fine specificity of any given clone of T cells, but rather on other factors, either intrinsic or extrinsic to the structure of the antigen. A major extrinsic factor is the MHC of the responding individual, probably due to a requirement for the immunodominant peptides to bind to the MHC of presenting cells in that individual. In looking for intrinsic factors, we noted that both immunodominant sites of myoglobin were amphipathic helices, i.e., helices having hydrophilic and hydrophobic residues on opposite sides. Studies with synthetic peptides indicated that residues on the hydrophilic side were necessary for T-cell recognition. However, unfolding of the native protein was shown to be the apparent goal of processing of antigen, presumably to expose something not already exposed on the native molecule, such as the hydrophobic sides of these helices. We propose that such exposure is necessary to interact with something on the presenting cell, such as MHC or membrane, where we have demonstrated the presence of antigenic peptides by blocking of presentation of biotinylated peptide with avidin. The membrane may serve as a short-term memory of peptides from antigens encountered by the presenting cell, for dynamic sampling by MHC molecules to be available for presentation to T cells. These ideas, together with the knowledge that T-cell recognition required only short peptides and therefore had to be based only on primary or secondary structure, not tertiary folding of the native protein, led us to propose that T-cell immunodominant epitopes may tend to be amphipathic

  12. Significance of Serology by Multi-Antigen ELISA for Tissue Helminthiases in Korea

    Science.gov (United States)

    2017-01-01

    It is clinically important to differentiate tissue-invading helminthiasis. The purpose of this study was to assess the specific immunoglobulin G (IgG) antibody positive rates for clonorchiasis, paragonimiasis, cysticercosis, and sparganosis 4 helminthiases from 1996 to 2006 using multi-antigen enzyme-linked immunosorbent assay (ELISA) in Korea. Results of 6,017 samples, which were referred to our institute for serodiagnosis, were analyzed. The subjects with positive serum IgG antibodies were 1,502 (25.0%) for any of the 4 helminthiases. The overall positive numbers for clonorchiasis, paragonimiasis, cysticercosis, and sparganosis were 728 (12.1%), 166 (2.8%), 729 (12.1%), and 263 (4.4%), respectively. The positive serologic reaction to multi-antigens was determined in 309 (20.6%) of the 1,502 total seropositive subjects. Those with multi-antigen positivity were regarded as positive for the antigen of strongest reaction but cross-reaction to others with weak positive reaction. Annual seropositive rates for those 4 tissue helminthiases ranged from 12.1% to 35.7%. The highest rate was observed in age from 60 to 69 years old and prevalence of men (27.4%; 1,030/3,763) was significantly higher than of women (19.1%; 332/1,741). Hospital records of 165 ELISA positive patients were reviewed to confirm correlation with their clinical diagnosis. Paragonimiasis was highly correlated as 81.8% (9/11), cysticercosis 29.9% (20/67), clonorchiasis 29.0% (20/69), and sparganosis 11.1% (2/18). In conclusion, the multi-antigen ELISA using 4 helminth antigens is useful to differentiate suspected tissue-invading helminthiases, especially ELISA diagnosis of paragonimiasis is reliable. The seropositivity is still high among suspected patients in Korea. PMID:28581268

  13. B lymphocytes as natural antigen-presenting cells (APC) of their own Ig receptor determinants

    International Nuclear Information System (INIS)

    Yurin, V.L.; Rudensky, A.Yu.; Rabinovich, O.R.; Kulakova, O.G.; Bobreneva, R.A.

    1986-01-01

    The authors use Igk-lb allotype-specific rat T cell proliferation(Pr) in vitro as a model of natural Ig determinants B cell presentation in Ig-specific T-B cell interactions. As shown before Igk-lb-specific responsiveness of AUG(RT-l/sup c/, Igk-la) and WAG (RT-l, Igk-la) rats is controlled by dominant Ir gene, linked to RT-l/sup c/. Only IgG(Igk-lb)-pulsed splenic APC of AUG(responder) but not WAG(non-responder) origin induce specific F 1 (WAGxAUG) T cell Pr. The same restriction was observed if purified B cells from Igk-l congeneic AUG-lb and WAG-lb rats were used as APC. B cell presentation was found to be sensitive to high irradiation dose(2000 rad). Anti-RT-l monoclonal antibody inhibition studies suggested RT-lB(I-A) molecule as a main restricting element of Igk-lb T cell recognition. B cell and splenic APC presentation of Igk-lb allotype was not inhibited by poly- and monoclonal anti-Igk-lb antibodies. Allelic exclusion of Igk-lb presentation by B cells from heterozygous F 1 (WAG-lbx AUG) rats was demonstrated by panning with antiallotypic reagents. Important, that irradiated anti-Igk-lb T cells induce specific Pr of normal Igk-lb-positive B cells. The data demonstrate MHC-restricted B cell presentation of their own receptor determinants, distinct from serologically-defined epitopes. T cell recognition of these determinants induce specific Pr of Ig-recognizing T cells and Ig-presenting B lymphocytes

  14. Antigen antibody interactions

    CERN Document Server

    DeLisi, Charles

    1976-01-01

    1. 1 Organization of the Immune System One of the most important survival mechanisms of vertebrates is their ability to recognize and respond to the onslaught of pathogenic microbes to which they are conti- ously exposed. The collection of host cells and molecules involved in this recognition­ 12 response function constitutes its immune system. In man, it comprises about 10 cells 20 (lymphocytes) and 10 molecules (immunoglobulins). Its ontogenic development is c- strained by the requirement that it be capable of responding to an almost limitless variety of molecular configurations on foreign substances, while simultaneously remaining inert to those on self components. It has thus evolved to discriminate, with exquisite precision, between molecular patterns. The foreign substances which induce a response, called antigens, are typically large molecules such as proteins and polysaccharides. The portions of these with which immunoglobulins interact are called epitopes or determinants. A typical protein epitope m...

  15. A single point in protein trafficking by Plasmodium falciparum determines the expression of major antigens on the surface of infected erythrocytes targeted by human antibodies.

    Science.gov (United States)

    Chan, Jo-Anne; Howell, Katherine B; Langer, Christine; Maier, Alexander G; Hasang, Wina; Rogerson, Stephen J; Petter, Michaela; Chesson, Joanne; Stanisic, Danielle I; Duffy, Michael F; Cooke, Brian M; Siba, Peter M; Mueller, Ivo; Bull, Peter C; Marsh, Kevin; Fowkes, Freya J I; Beeson, James G

    2016-11-01

    Antibodies to blood-stage antigens of Plasmodium falciparum play a pivotal role in human immunity to malaria. During parasite development, multiple proteins are trafficked from the intracellular parasite to the surface of P. falciparum-infected erythrocytes (IEs). However, the relative importance of different proteins as targets of acquired antibodies, and key pathways involved in trafficking major antigens remain to be clearly defined. We quantified antibodies to surface antigens among children, adults, and pregnant women from different malaria-exposed regions. We quantified the importance of antigens as antibody targets using genetically engineered P. falciparum with modified surface antigen expression. Genetic deletion of the trafficking protein skeleton-binding protein-1 (SBP1), which is involved in trafficking the surface antigen PfEMP1, led to a dramatic reduction in antibody recognition of IEs and the ability of human antibodies to promote opsonic phagocytosis of IEs, a key mechanism of parasite clearance. The great majority of antibody epitopes on the IE surface were SBP1-dependent. This was demonstrated using parasite isolates with different genetic or phenotypic backgrounds, and among antibodies from children, adults, and pregnant women in different populations. Comparisons of antibody reactivity to parasite isolates with SBP1 deletion or inhibited PfEMP1 expression suggest that PfEMP1 is the dominant target of acquired human antibodies, and that other P. falciparum IE surface proteins are minor targets. These results establish SBP1 as part of a critical pathway for the trafficking of major surface antigens targeted by human immunity, and have key implications for vaccine development, and quantifying immunity in populations.

  16. Doxorubicin-anti-carcinoembryonic antigen immunoconjugate activity in vitro.

    Science.gov (United States)

    Richardson, V J; Ford, C H; Tsaltas, G; Gallant, M E

    1989-04-01

    An in vitro model consisting of a series of 11 human cancer cell lines with varying density of expression of membrane carcinoembryonic antigen (CEA) has been used to evaluate conjugates of doxorubicin (Adriamycin) covalently linked by a carbodiimide method to goat polyclonal antibodies and mouse monoclonal antibodies to CEA. Conjugates were produced which retained both antigen binding and drug cytotoxicity. IC50 values were determined for free drug, free drug mixed with unconjugated antibodies and for the immunoconjugates. Cell lines that were very sensitive to free drug (IC50 less than 100 ng/ml) were also found to be highly sensitive to conjugated drug and similarly cell lines resistant to drug (IC50 greater than 1,000 ng/ml) were also resistant to conjugated drug. Although there was no correlation between CEA expression and conjugates efficacy, competitive inhibition studies using autologous antibody to block conjugate binding to cells indicated immunoconjugates specificity for the CEA target.

  17. Prevalence of Hepatitis-B Surface Antigen among Blood Donors in ...

    African Journals Online (AJOL)

    Information is scarce on the prevalence of Hepatitis-B Virus (HBV) infection among blood donors in Taraba State. Hepatitis-B surface antigen (HBsAg) ELISA [Gudans Industrial Hong 2 Kou, China] was used to determine the prevalence of HBsAg among 804 blood donors aged between 11 and 65 years in Federal Medical ...

  18. Assessment of a serum tumour marker for carcinoma of the pancreas: the carbohydrate antigen C.A. 19-9

    International Nuclear Information System (INIS)

    Vincent, D.; Venot, J.; Catanzano, G.; Clement, M.N.; Piquet, M.F.; Veyriras, E.; Beck, C.

    1985-01-01

    A radio-immunological assay with monoclonal antibodies was used to measure the C 19-9 antigen in 51 patients to determine its diagnostic value in cancer of the pancreas. The results show that the C 19-9 antigen is a good marker for carcinoma of the pancreas and that it can be commonly used [fr

  19. Phenotypic H-Antigen Typing by Mass Spectrometry Combined with Genetic Typing of H Antigens, O Antigens, and Toxins by Whole-Genome Sequencing Enhances Identification of Escherichia coli Isolates.

    Science.gov (United States)

    Cheng, Keding; Chui, Huixia; Domish, Larissa; Sloan, Angela; Hernandez, Drexler; McCorrister, Stuart; Robinson, Alyssia; Walker, Matthew; Peterson, Lorea A M; Majcher, Miles; Ratnam, Sam; Haldane, David J M; Bekal, Sadjia; Wylie, John; Chui, Linda; Tyler, Shaun; Xu, Bianli; Reimer, Aleisha; Nadon, Celine; Knox, J David; Wang, Gehua

    2016-08-01

    Mass spectrometry-based phenotypic H-antigen typing (MS-H) combined with whole-genome-sequencing-based genetic identification of H antigens, O antigens, and toxins (WGS-HOT) was used to type 60 clinical Escherichia coli isolates, 43 of which were previously identified as nonmotile, H type undetermined, or O rough by serotyping or having shown discordant MS-H and serotyping results. Whole-genome sequencing confirmed that MS-H was able to provide more accurate data regarding H antigen expression than serotyping. Further, enhanced and more confident O antigen identification resulted from gene cluster based typing in combination with conventional typing based on the gene pair comprising wzx and wzy and that comprising wzm and wzt The O antigen was identified in 94.6% of the isolates when the two genetic O typing approaches (gene pair and gene cluster) were used in conjunction, in comparison to 78.6% when the gene pair database was used alone. In addition, 98.2% of the isolates showed the existence of genes for various toxins and/or virulence factors, among which verotoxins (Shiga toxin 1 and/or Shiga toxin 2) were 100% concordant with conventional PCR based testing results. With more applications of mass spectrometry and whole-genome sequencing in clinical microbiology laboratories, this combined phenotypic and genetic typing platform (MS-H plus WGS-HOT) should be ideal for pathogenic E. coli typing. Copyright © 2016 Cheng et al.

  20. Reduction of T-Helper Cell Responses to Recall Antigen Mediated by Codelivery with Peptidoglycan via the Intestinal Nanomineral-Antigen Pathway.

    Science.gov (United States)

    Hewitt, Rachel E; Robertson, Jack; Haas, Carolin T; Pele, Laetitia C; Powell, Jonathan J

    2017-01-01

    Naturally occurring intestinal nanomineral particles constituently form in the mammalian gut and trap luminal protein and microbial components. These cargo loaded nanominerals are actively scavenged by M cells of intestinal immune follicles, such as Peyer's patches and are passed to antigen-presenting cells. Using peripheral blood mononuclear cell populations as an in vitro model of nanomineral uptake and antigen presentation, we show that monocytes avidly phagocytose nanomineral particles bearing antigen and peptidoglycan (PGN), and that the presence of PGN within particles downregulates their cell surface MHC class II and upregulates programmed death receptor ligand 1. Nanomineral delivery of antigen suppresses antigen-specific CD4 + T cell responses, an effect that is enhanced in the presence of PGN. Blocking the interleukin-10 receptor restores CD4 + T cell responses to antigen codelivered with PGN in nanomineral form. Using human intestinal specimens, we have shown that the in vivo nanomineral pathway operates in an interleukin-10 rich environment. Consequently, the delivery of a dual antigen-PGN cargo by endogenous nanomineral in vivo is likely to be important in the establishment of intestinal tolerance, while their synthetic mimetics present a potential delivery system for therapeutic applications targeting the modulation of Peyer's patch T cell responses.

  1. Antigenic cross-reactivity and immunogenicity of Bothrops venoms from snakes of the Amazon region.

    Science.gov (United States)

    Furtado, Maria de Fátima D; Cardoso, Silvia Travaglia; Soares, Oscar Espellet; Pereira, Aparecida Pietro; Fernandes, Daniel Silva; Tambourgi, Denise Vilarinho; Sant'Anna, Osvaldo Augusto

    2010-04-01

    Snakebites are still a critical public health problem in developing countries or isolated areas. In Brazil, the North Region has a high distribution coefficient worsened by the significant number of eventually unreported cases, due to difficulties in access to health services, to the natural geographic barriers and the vast territory. In the Rio Negro area, the species Bothrops atrox, Bothrops brazili, Lachesis muta muta and Bothriopsis taeniata are thought to be the major species responsible for snakebites. The aim of this study was to qualitatively and quantitatively determine the antigenic cross-reactivity and expression of toxins and the immunogenicity of Bothrops venom species of the Amazon and to evaluate the general efficacy of the therapeutic sera. The in vivo assays demonstrated that the defibrinating activity of B. taeniata venom was absent but that the lethal and hemorrhagic properties were more intense than in the B. atrox venom. The results evidence venom variability among the two B. atrox populations from two distinct Amazonian regions, which may reveal a subjacent speciation process. The results point to new aspects that may guide the improvement of anti-Bothropic therapeutic serum. Copyright 2010 Elsevier Ltd. All rights reserved.

  2. Hepatitis B surface antigen incorporated in dissolvable microneedle array patch is antigenic and thermostable.

    Science.gov (United States)

    Poirier, Danielle; Renaud, Frédéric; Dewar, Vincent; Strodiot, Laurent; Wauters, Florence; Janimak, Jim; Shimada, Toshio; Nomura, Tatsuya; Kabata, Koki; Kuruma, Koji; Kusano, Takayuki; Sakai, Masaki; Nagasaki, Hideo; Oyamada, Takayoshi

    2017-11-01

    Alternatives to syringe-based administration are considered for vaccines. Intradermal vaccination with dissolvable microneedle arrays (MNA) appears promising in this respect, as an easy-to-use and painless method. In this work, we have developed an MNA patch (MNAP) made of hydroxyethyl starch (HES) and chondroitin sulphate (CS). In swines, hepatitis B surface antigen (HBsAg) formulated with the saponin QS-21 as adjuvant, both incorporated in HES-based MNAP, demonstrated the same level of immunogenicity as a commercially available aluminum-adjuvanted HBsAg vaccine, after two immunizations 28 days apart. MNAP application was associated with transient skin reactions (erythema, lump, scab), particularly evident when the antigen was delivered with the adjuvant. The thermostability of the adjuvanted antigen when incorporated in the HES-based matrix was also assessed by storing MNAP at 37, 45 or 50 °C for up to 6 months. We could demonstrate that antigenicity was retained at 37 and 45 °C and only a 10% loss was observed after 6 months at 50 °C. Our results are supportive of MNAP as an attractive alternative to classical syringe-based vaccination. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  3. Added value of antigen ELISA in the diagnosis of neurocysticercosis in resource poor settings.

    Directory of Open Access Journals (Sweden)

    Sarah Gabriël

    Full Text Available BACKGROUND: Neurocysticercosis (NCC is the most common cause of acquired epilepsy in Taenia solium endemic areas, primarily situated in low-income countries. Diagnosis is largely based upon the "Del Brutto diagnostic criteria" using the definitive/probable/no NCC diagnosis approach. Neuroimaging and specific T. solium cysticercosis antibody detection results are at the mainstay of this diagnosis, while antigen detection in serum has never been included. This study aimed at evaluating the addition of antigen detection as a major diagnostic criterion, especially in areas where neuroimaging is absent. METHODS: The B158/B60 monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA for the detection of circulating cysticercus antigen was carried out retrospectively on serum samples collected during a hospital-based study from 83 people with epilepsy (PWE in an endemic area. RESULTS: The addition of antigen results as a major criterion allowed the correct diagnosis of definitive NCC in 10 out of 17 patients as opposed to 0/17 without antigen results in the absence of neuroimaging. A sensitivity of 100% and a specificity of 84% were determined for the diagnosis of active NCC using antigen ELISA. While the use of a higher cutoff improves the specificity of the test to 96%, it decreases its sensitivity to 83%. CONCLUSIONS: In areas where neuroimaging is absent, NCC diagnosis according to the existing criteria is problematic. Taking into account its limitations for diagnosis of inactive NCC, antigen detection can be of added value for diagnosing NCC in PWE by supporting diagnostic and treatment decisions. Therefore, we recommend a revision of the "Del Brutto diagnostic criteria" for use in resource poor areas and suggest the inclusion of serum antigen detection as a major criterion.

  4. Evidence against suppressor cell involvement in naturally acquired tolerance of a minor histocompatibility antigen

    International Nuclear Information System (INIS)

    Johnson, L.L.

    1991-01-01

    The hypothesis was investigated that suppressor cells may be responsible for maintenance of immunologic tolerance of a minor H3 antigen in mice that express the antigen naturally. Lymphoid cell populations from B6.C-H-24c (HW54) mice, a congenic-resistant strain histoincompatible with H-24b-expressing C57BL/6 (B6) mice only with respect to the H-24 locus, were examined in cell-transfer experiments to see if they contained naturally arising H-24c-specific suppressor cells. The H-24 antigen was chosen for these studies because, unlike most other minor and major histocompatibility (H) antigens, it is not detectable on mature lymphoid cells by any of several functional criteria. Thus transfer of HW54 lymphoid cells to B6 hosts could be done without the complication of inducing hyporesponsiveness de novo in the host, as occurs with other minor H antigens that are expressed on lymphocytes. B6 hosts were given HW54 skin grafts along with HW54 lymphoid cells to assess their tolerance of the H-24c-encoded antigen. The hosts were either (1) normal, nonimmune B6 mice; (2) B6 mice rendered immunodeficient by thymectomy and irradiation (TxB) and repopulated with H-24c-immune B6 lymphocytes; or (3) TxB B6 hosts repopulated with nonimmune B6 lymphocytes. In each case it was found that the additionally infused HW54 lymphoid cells did not suppress the ability of these hosts to reject HW54 skin grafts. In other words, HW54 lymphoid cells appear not to possess suppressive activity specific for the H-24c antigen that might maintain antigen-specific natural tolerance. Additional experiments were performed to determine whether HW54 lymphoid cells can inhibit the ability of sublethally irradiated B6 mice to regain the capacity to reject HW54 skin

  5. Common antigenic determinants of haemoglobin as basis of immunological cross-reactivity between chironomid species (Diptera, Chironomidae): studies with human and animal sera.

    Science.gov (United States)

    Baur, X; Dewair, M; Haegele, K; Prelicz, H; Scholl, A; Tichy, H

    1983-01-01

    Chironomids, of which approximately 10,000 species exist, are reported to cause severe immediate type allergic diseases in man. In the present study, immunological cross-reactivity between 14 chironomid species from different continents was proven by RAST inhibition, double immunodiffusion and a new allergoprint technique, based upon PAGE separation of insect crude extracts. Using isolated chironomid haemoglobins and sera of sensitized persons, as well as rabbit antibodies against larval crude extract or against the haemoglobin fraction of Chironomus thummi, it could be proven that cross-reactivity derives at least predominantly from haemoglobin components with common antigenic determinants in the different species. Images Fig. 2 Fig. 7 Fig. 8 Fig. 9 PMID:6197219

  6. Virosomes for antigen and DNA delivery

    NARCIS (Netherlands)

    Daemen, T; de Mare, A; Bungener, L; de Jonge, J; Huckriede, A; Wilschut, J

    2005-01-01

    Specific targeting and delivery as well as the display of antigens on the surface of professional antigen-presenting cells (APCs) are key issues in the design and development of new-generation vaccines aimed at the induction of both humoral and cell-mediated immunity. Prophylactic vaccination

  7. Distinct functional programming of human fetal and adult monocytes.

    Science.gov (United States)

    Krow-Lucal, Elisabeth R; Kim, Charles C; Burt, Trevor D; McCune, Joseph M

    2014-03-20

    Preterm birth affects 1 out of 9 infants in the United States and is the leading cause of long-term neurologic handicap and infant mortality, accounting for 35% of all infant deaths in 2008. Although cytokines including interferon-γ (IFN-γ), interleukin-10 (IL-10), IL-6, and IL-1 are produced in response to in utero infection and are strongly associated with preterm labor, little is known about how human fetal immune cells respond to these cytokines. We demonstrate that fetal and adult CD14(+)CD16(-) classical monocytes are distinct in terms of basal transcriptional profiles and in phosphorylation of signal transducers and activators of transcription (STATs) in response to cytokines. Fetal monocytes phosphorylate canonical and noncanonical STATs and respond more strongly to IFN-γ, IL-6, and IL-4 than adult monocytes. We demonstrate a higher ratio of SOCS3 to IL-6 receptor in adult monocytes than in fetal monocytes, potentially explaining differences in STAT phosphorylation. Additionally, IFN-γ signaling results in upregulation of antigen presentation and costimulatory machinery in adult, but not fetal, monocytes. These findings represent the first evidence that primary human fetal and adult monocytes are functionally distinct, potentially explaining how these cells respond differentially to cytokines implicated in development, in utero infections, and the pathogenesis of preterm labor.

  8. Observations of pretreatment prostate-specific antigen doubling time in 107 patients referred for definitive radiotherapy

    International Nuclear Information System (INIS)

    Lee, W. Robert; Hanks, Gerald E.; Corn, Benjamin W.; Schultheiss, Timothy E.

    1995-01-01

    Purpose: To determine pretreatment prostate-specific antigen doubling times (PSADT) in patients referred for definitive radiotherapy. Methods and Materials: One hundred and seven patients with histologically proven nonmetastatic prostate cancer and an elevated prostate-specific antigen (PSA) who were referred for radiation therapy had three serum PSA values obtained prior to the start of definitive therapy. Prostate-specific antigen doubling times were calculated by linear regression. Results: Prostate-specific antigen values increased during the period of observation in 78 patients (73%). Forty-three patients (40%) had calculated PSADT of less than 2 years and of those patients with pretreatment serum PSA values of greater than 10 ng/mL more than 50% has calculated PSADT of less than 2 years. Conclusions: A significant minority of patients referred for radiotherapy have calculated PSADT of less than 2 years. The significance of this relatively fast growth rate is as yet undetermined, but suggests that patients referred for radiotherapy may have aggressive disease prior to treatment

  9. Characterizing complex polysera produced by antigen-specific immunization through the use of affinity-selected mimotopes.

    Directory of Open Access Journals (Sweden)

    Galina Denisova

    Full Text Available BACKGROUND: Antigen-based (as opposed to whole organism vaccines are actively being pursued for numerous indications. Even though different formulations may produce similar levels of total antigen-specific antibody, the composition of the antibody response can be quite distinct resulting in different levels of therapeutic activity. METHODOLOGY/PRINCIPAL FINDINGS: Using plasmid-based immunization against the proto-oncogene HER-2 as a model, we have demonstrated that affinity-selected epitope mimetics (mimotopes can provide a defined signature of a polyclonal antibody response. Further, using novel computer algorithms that we have developed, these mimotopes can be used to predict epitope targets. CONCLUSIONS/SIGNIFICANCE: By combining our novel strategy with existing methods of epitope prediction based on physical properties of an individual protein, we believe that this method offers a robust method for characterizing the breadth of epitope-specificity within a specific polyserum. This strategy is useful as a tool for monitoring immunity following vaccination and can also be used to define relevant epitopes for the creation of novel vaccines.

  10. Purification of a polyclonal antibody against CD147 for ELISA using antigen-immunoaffinity chromatography

    Science.gov (United States)

    Liu, Shuangshuang; Li, Shasha; Zhang, Yang; Wang, Ye; Zhu, Yumeng; Wang, Bin; Chen, Zhi-Nan

    2017-01-01

    The immunoglobulin superfamily member CD147 is a widely expressed glycoprotein that occurs in both a membrane-spanning and soluble form. Sandwich ELISA is a powerful tool for analyzing soluble antigens. The aim of the present study was to obtain a highly specific polyclonal antibody against human CD147 that can be used for sandwich ELISA analysis. Expression of recombinant CD147 by a eukaryotic expression system was used to immunize rabbits to obtain antiserum. A highly specific polyclonal antibody that was able to detect soluble CD147 in sandwich ELISA was obtained by antigen-immunoaffinity chromatography purification. The purity of rabbit anti-CD147 polyclonal antibodies was ~99%, and ELISA analysis was able to determine the titer of the rabbit anti-CD147 polyclonal antibodies at 1:512,000. The lowest concentration of the standard CD147 antigen that the sandwich ELISA was able to detect was 31.25 pg/ml. The sandwich ELISA system was composed of anti-hepatoma HAb18 monoclonal antibodies and purified rabbit anti-CD147 polyclonal antibodies. The present study demonstrated that antigen-immunoaffinity chromatography may be a good technique for the purification of polyclonal antibodies, which may be used to detect antigen in sandwich ELISAs. PMID:28487989

  11. Proteomic Profiling of Serological Responses to Aspergillus fumigatus Antigens in Patients with Invasive Aspergillosis.

    Science.gov (United States)

    Teutschbein, Janka; Simon, Svenja; Lother, Jasmin; Springer, Jan; Hortschansky, Peter; Morton, C Oliver; Löffler, Jürgen; Einsele, Hermann; Conneally, Eibhlin; Rogers, Thomas R; Guthke, Reinhard; Brakhage, Axel A; Kniemeyer, Olaf

    2016-05-06

    Aspergillus fumigatus is the species that most commonly causes the opportunistic infection invasive aspergillosis (IA) in patients being treated for hematological malignancies. Little is known about the A. fumigatus proteins that trigger the production of Aspergillus-specific IgG antibodies during the course of IA. To characterize the serological response to A. fumigatus protein antigens, mycelial proteins were separated by 2-D gel electrophoresis. The gels were immunoblotted with sera from patients with probable and proven IA and control patients without IA. We identified 49 different fungal proteins, which gave a positive IgG antibody signal. Most of these antigens play a role in primary metabolism and stress responses. Overall, our analysis identified 18 novel protein antigens from A. fumigatus. To determine whether these antigens can be used as diagnostic or prognostic markers or exhibit a protective activity, we employed supervised machine learning with decision trees. We identified two candidates for further analysis, the protein antigens CpcB and Shm2. Heterologously produced Shm2 induced a strongly proinflammatory response in human peripheral blood mononuclear cells after in vitro stimulation. In contrast, CpcB did not activate the immune response of PBMCs. These findings could serve as the basis for the development of an immunotherapy of IA.

  12. Lack of radioimmunodetection and complications associated with monoclonal anticarcinoembryonic antigen antibody cross-reactivity with an antigen on circulating cells

    International Nuclear Information System (INIS)

    Dillman, R.O.; Beauregard, J.C.; Sobol, R.E.; Royston, I.; Bartholomew, R.M.; Hagan, P.S.; Halpern, S.E.

    1984-01-01

    Characterization of several high-affinity murine monoclonal anticarcinoembryonic antigen (CEA) antibodies suggested good specificity except for cross-reactivity with an antigen on granulocytes and erythrocytes which was different from the previously described normal cross-reacting antigen of granulocytes. In vivo studies in athymic mice using an indium conjugate of an anti-CEA monoclonal antibody (MoAb) revealed excellent specific uptake in colorectal carcinoma xenografts. Studies were conducted in humans to determine the limitations produced by the cross-reactivity with granulocytes and erythrocytes. Patients with metastatic colorectal cancer received 3 to 6 mg of anti-CEA MoAb over 10 min or 2 hr. In five of six trials, the MoAb infusion was associated with a 40 to 90% decrease in circulating granulocytes and systemic toxicity including fever, rigors, and emesis. One patient had no change in cell count and had no toxicity. Radionuclide scans with 111 In-anti-CEA MoAb showed marked uptake in the spleen when cells were eliminated, and in the liver, especially when pretreatment CEA levels were high. Metastatic tumor sites failed to concentrate the isotope. This study emphasizes the potential limitations for radioimmunodetection and/or radioimmunotherapy imposed by reactivity with circulating cells, and suggests that certain toxic reactions associated with MoAb infusions are related to destruction of circulating cells rather than allergic reactions to mouse protein. It also emphasizes how variables such as dose and binding affinity of antibody, radioisotope used, and assessment at different observation points can obscure lack of antibody specificity

  13. Lack of radioimmunodetection and complications associated with monoclonal anticarcinoembryonic antigen antibody cross-reactivity with an antigen on circulating cells

    Energy Technology Data Exchange (ETDEWEB)

    Dillman, R.O.; Beauregard, J.C.; Sobol, R.E.; Royston, I.; Bartholomew, R.M.; Hagan, P.S.; Halpern, S.E.

    1984-05-01

    Characterization of several high-affinity murine monoclonal anticarcinoembryonic antigen (CEA) antibodies suggested good specificity except for cross-reactivity with an antigen on granulocytes and erythrocytes which was different from the previously described normal cross-reacting antigen of granulocytes. In vivo studies in athymic mice using an indium conjugate of an anti-CEA monoclonal antibody (MoAb) revealed excellent specific uptake in colorectal carcinoma xenografts. Studies were conducted in humans to determine the limitations produced by the cross-reactivity with granulocytes and erythrocytes. Patients with metastatic colorectal cancer received 3 to 6 mg of anti-CEA MoAb over 10 min or 2 hr. In five of six trials, the MoAb infusion was associated with a 40 to 90% decrease in circulating granulocytes and systemic toxicity including fever, rigors, and emesis. One patient had no change in cell count and had no toxicity. Radionuclide scans with /sup 111/In-anti-CEA MoAb showed marked uptake in the spleen when cells were eliminated, and in the liver, especially when pretreatment CEA levels were high. Metastatic tumor sites failed to concentrate the isotope. This study emphasizes the potential limitations for radioimmunodetection and/or radioimmunotherapy imposed by reactivity with circulating cells, and suggests that certain toxic reactions associated with MoAb infusions are related to destruction of circulating cells rather than allergic reactions to mouse protein. It also emphasizes how variables such as dose and binding affinity of antibody, radioisotope used, and assessment at different observation points can obscure lack of antibody specificity.

  14. Functional, Antigen-Specific Stem Cell Memory (TSCM CD4+ T Cells Are Induced by Human Mycobacterium tuberculosis Infection

    Directory of Open Access Journals (Sweden)

    Cheleka A. M. Mpande

    2018-03-01

    Full Text Available BackgroundMaintenance of long-lasting immunity is thought to depend on stem cell memory T cells (TSCM, which have superior self-renewing capacity, longevity and proliferative potential compared with central memory (TCM or effector (TEFF T cells. Our knowledge of TSCM derives primarily from studies of virus-specific CD8+ TSCM. We aimed to determine if infection with Mycobacterium tuberculosis (M. tb, the etiological agent of tuberculosis, generates antigen-specific CD4+ TSCM and to characterize their functional ontology.MethodsWe studied T cell responses to natural M. tb infection in a longitudinal adolescent cohort of recent QuantiFERON-TB Gold (QFT converters and three cross-sectional QFT+ adult cohorts; and to bacillus Calmette–Guerin (BCG vaccination in infants. M. tb and/or BCG-specific CD4 T cells were detected by flow cytometry using major histocompatibility complex class II tetramers bearing Ag85, CFP-10, or ESAT-6 peptides, or by intracellular cytokine staining. Transcriptomic analyses of M. tb-specific tetramer+ CD4+ TSCM (CD45RA+ CCR7+ CD27+ were performed by microfluidic qRT-PCR, and functional and phenotypic characteristics were confirmed by measuring expression of chemokine receptors, cytotoxic molecules and cytokines using flow cytometry.ResultsM. tb-specific TSCM were not detected in QFT-negative persons. After QFT conversion frequencies of TSCM increased to measurable levels and remained detectable thereafter, suggesting that primary M. tb infection induces TSCM cells. Gene expression (GE profiling of tetramer+ TSCM showed that these cells were distinct from bulk CD4+ naïve T cells (TN and shared features of bulk TSCM and M. tb-specific tetramer+ TCM and TEFF cells. These TSCM were predominantly CD95+ and CXCR3+, markers typical of CD8+ TSCM. Tetramer+ TSCM expressed significantly higher protein levels of CCR5, CCR6, CXCR3, granzyme A, granzyme K, and granulysin than bulk TN and TSCM cells. M. tb-specific TSCM were also

  15. Genetic diversity and antigenicity variation of Babesia bovis merozoite surface antigen-1 (MSA-1) in Thailand.

    Science.gov (United States)

    Tattiyapong, Muncharee; Sivakumar, Thillaiampalam; Takemae, Hitoshi; Simking, Pacharathon; Jittapalapong, Sathaporn; Igarashi, Ikuo; Yokoyama, Naoaki

    2016-07-01

    Babesia bovis, an intraerythrocytic protozoan parasite, causes severe clinical disease in cattle worldwide. The genetic diversity of parasite antigens often results in different immune profiles in infected animals, hindering efforts to develop immune control methodologies against the B. bovis infection. In this study, we analyzed the genetic diversity of the merozoite surface antigen-1 (msa-1) gene using 162 B. bovis-positive blood DNA samples sourced from cattle populations reared in different geographical regions of Thailand. The identity scores shared among 93 msa-1 gene sequences isolated by PCR amplification were 43.5-100%, and the similarity values among the translated amino acid sequences were 42.8-100%. Of 23 total clades detected in our phylogenetic analysis, Thai msa-1 gene sequences occurred in 18 clades; seven among them were composed of sequences exclusively from Thailand. To investigate differential antigenicity of isolated MSA-1 proteins, we expressed and purified eight recombinant MSA-1 (rMSA-1) proteins, including an rMSA-1 from B. bovis Texas (T2Bo) strain and seven rMSA-1 proteins based on the Thai msa-1 sequences. When these antigens were analyzed in a western blot assay, anti-T2Bo cattle serum strongly reacted with the rMSA-1 from T2Bo, as well as with three other rMSA-1 proteins that shared 54.9-68.4% sequence similarity with T2Bo MSA-1. In contrast, no or weak reactivity was observed for the remaining rMSA-1 proteins, which shared low sequence similarity (35.0-39.7%) with T2Bo MSA-1. While demonstrating the high genetic diversity of the B. bovis msa-1 gene in Thailand, the present findings suggest that the genetic diversity results in antigenicity variations among the MSA-1 antigens of B. bovis in Thailand. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Standardization and validation of a cytometric bead assay to assess antibodies to multiple Plasmodium falciparum recombinant antigens.

    Science.gov (United States)

    Ondigo, Bartholomew N; Park, Gregory S; Gose, Severin O; Ho, Benjamin M; Ochola, Lyticia A; Ayodo, George O; Ofulla, Ayub V; John, Chandy C

    2012-12-21

    Multiplex cytometric bead assay (CBA) have a number of advantages over ELISA for antibody testing, but little information is available on standardization and validation of antibody CBA to multiple Plasmodium falciparum antigens. The present study was set to determine optimal parameters for multiplex testing of antibodies to P. falciparum antigens, and to compare results of multiplex CBA to ELISA. Antibodies to ten recombinant P. falciparum antigens were measured by CBA and ELISA in samples from 30 individuals from a malaria endemic area of Kenya and compared to known positive and negative control plasma samples. Optimal antigen amounts, monoplex vs multiplex testing, plasma dilution, optimal buffer, number of beads required were assessed for CBA testing, and results from CBA vs. ELISA testing were compared. Optimal amounts for CBA antibody testing differed according to antigen. Results for monoplex CBA testing correlated strongly with multiplex testing for all antigens (r = 0.88-0.99, P values from <0.0001 - 0.004), and antibodies to variants of the same antigen were accurately distinguished within a multiplex reaction. Plasma dilutions of 1:100 or 1:200 were optimal for all antigens for CBA testing. Plasma diluted in a buffer containing 0.05% sodium azide, 0.5% polyvinylalcohol, and 0.8% polyvinylpyrrolidone had the lowest background activity. CBA median fluorescence intensity (MFI) values with 1,000 antigen-conjugated beads/well did not differ significantly from MFI with 5,000 beads/well. CBA and ELISA results correlated well for all antigens except apical membrane antigen-1 (AMA-1). CBA testing produced a greater range of values in samples from malaria endemic areas and less background reactivity for blank samples than ELISA. With optimization, CBA may be the preferred method of testing for antibodies to P. falciparum antigens, as CBA can test for antibodies to multiple recombinant antigens from a single plasma sample and produces a greater range of values in

  17. Tumor markers cancer antigen 15.3, carcinoembryonic antigen, and tissue polypeptide antigen for monitoring metastatic breast cancer during first-line chemotherapy and follow-up

    DEFF Research Database (Denmark)

    Sölétormos, G; Nielsen, D; Schiøler, V

    1996-01-01

    progressive disease, the median positive lead time was 35 days during therapy and 76 days during follow-up. Tumor marker assessment may document that a therapy is effective and ought to be continued in spite of adverse toxic effects, and that a treatment is ineffective and should be stopped to prevent......We investigated whether model systems integrating stochastic variation into criteria for marker assessment could be used for monitoring metastatic breast cancer. A total of 3989 serum samples was obtained from 204 patients receiving first-line chemotherapy and from 112 of these patients during...... follow-up. Each sample was analyzed for cancer antigen 15.3, carcinoembryonic antigen, and tissue polypeptide antigen. The efficiency for identifying progression and nonprogression was 94% during therapy and 85% during follow-up, with no false-positive marker results for progressive disease. At clinical...

  18. Radioimmunoassay for the detection of Australia-SH antigen

    Energy Technology Data Exchange (ETDEWEB)

    Gerhardt, H [Giessen Univ. (Germany, F.R.). Zentrum fuer Innere Medizin

    1974-06-01

    Among infectious diseases, hepatitis presents a great problem in all countries with a high medical standard. The number of Australia antigen-positive cases rises from year to year, due to the increase in drug-fixer hepatitis and blood transfusions. Highly sensitive and at the same time practicable methods are therefore required for the identification of Australia antigen carriers and their elimination as blood donors. The most sensitive of all currently used tests for the detection of Australia antigen is the 'solid phase' radioimmunoassay since it permits an objective and quantitative measurement of the antigen.

  19. 21 CFR 660.40 - Hepatitis B Surface Antigen.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Hepatitis B Surface Antigen. 660.40 Section 660.40...) BIOLOGICS ADDITIONAL STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Hepatitis B Surface Antigen § 660.40 Hepatitis B Surface Antigen. (a) Proper name and definition. The proper name of this product...

  20. Accurate radioimmunoassay of human growth hormone with separation on polyacrylamide gel electrophoresis of free antigen; antigen-anti-body complex and damaged labelled antigen: a study is performed on this last one for the purpose of obtaining long lasting labelled products

    International Nuclear Information System (INIS)

    Bartolini, P.; Assis, L.M.; Schwarz, I.; Macchione, M.; Pieroni, R.R.

    1977-01-01

    The intent of this work was the obtainement of a radioimmunoassay method, accurate and precise enough, to furnish a suitable way for determining Human Growth Hormone (HGH) in extracts or in physiological fluids, useful more for specific research purposes than for routinized clinical assays, and where the labelled products could be used as long as possible. Only one technique was found that could give an answer to these requirements, though under some aspects more laborious than others: Polyacrylamide Gel Electrophoresis (PAGE). This was used introducing some modifications to the original method of Davis, and it was possible, using tubes 11 cm long, to separate on the same gel, the free, the antibody bound, and the damaged labelled antigen. The method, being able to detect separately and independently these three components and to give a better control on the analytically dangerous 'damaged', furnished accurate and reproducible curves. An example of determination is the one on KABI-Crescormon, that compares the results obatined with the present technique to those presented by another laboratory. Thanks to this method the labelled antigen could be used up to one month of time. After this period a re-purification on Sephadex was introduced so that the same labelled product was profitable for two more months. Parallel to this work a study has been performed on the various components originating in this so called process of 'damaging', and a particular importance has also been given to a more precise knowledge of the amount of antigen, in terms of mass, present in an assay. (orig.) [de

  1. IDENTIFICATION OF ACROSOME AS THE MAIN ANTIGEN OF THE SPERM CELLS PROVOKING AUTOANTIBODIES IN VASECTOMIZED IRANIAN MEN

    Directory of Open Access Journals (Sweden)

    M R Nowroozi

    2008-12-01

    Full Text Available "nVasectomy is one of the extensively used methods of contraception in family planning programs. Antisperm antibodies (ASA develop after vasectomy which can result in auto-immune male infertility. The precise sperm antigens involved in the autoimmune response are still poorly defined, therefore we determined the circulating ASA and identified relevant sperm antigens based on localization of binding sites of ASA to sperm cell antigens, using a rapid, inexpensive and clinically relevant assay in vasectomized men. Results showed that 2.5% of men had ASA at the time of vasectomy, whereas 53.5% of the study population subsequently developed ASA. The numbers of men with circulating ASA increased significantly for the first three months after vasectomy. These antibodies were distinguishable into three groups based on their bindings to different sites of sperm cell antigens including against acrosome and tail in 67.56% and 10.8%, respectively; 21.6% of subjects had antibody to the other parts of the sperm cell antigens. The results of this study are discussed in terms of an autoimmune response against sperm antigens and development of ASA.

  2. Radioimmunological determination of hemoglobin Asub(1c)

    International Nuclear Information System (INIS)

    Javid, J.; Cerami, A.; Koenig, R.J.; Walsh, P.K.P.

    1979-01-01

    The antibodies fighting human haemaglobin A sub(1c) to diagnose diabetes is obtained by immunisation of cats, goats or sheep. The acquisition of antigen antibody complexes in a blood sample is done by radioimmunological determination using antigen labelled with I 125 (several examples). (DG) [de

  3. Radioimmunological test for the cancero-embryonal antigen in evaluation of stomach neoplasm treatment efficiency

    International Nuclear Information System (INIS)

    Klimenkov, A.A.; Tkacheva, G.A.; Gladikov, Yu.V.; Blokhina, N.G.; Ozherel'ev, A.V.

    1979-01-01

    The results of a dynamic determination of the level of the cancero-embryonic antigen are analysed in 30 patients with stomach neoplasm of the 1-3 stages subjected to a radical operation and 22 patients with stage 4 given polychemotherapy. It is shown that information on the nature of the change in the level of the cancero-embryonal antigen in the blood serves as an important criterion for evaluation of the completeness of the tumour mass removal, detection of the disease relapse and comparison of the efficiency of various combinations of antitumor drugs

  4. Prevalence of Rh, Duffy, Kell, Kidd & MNSs blood group antigens in the Indian blood donor population.

    Science.gov (United States)

    Makroo, R N; Bhatia, Aakanksha; Gupta, Richa; Phillip, Jessy

    2013-03-01

    Little data are available regarding the frequencies of the blood group antigens other than ABO and RhD in the Indian population. Knowledge of the antigen frequencies is important to assess risk of antibody formation and to guide the probability of finding antigen-negative donor blood, which is especially useful when blood is required for a patient who has multiple red cell alloantibodies. This study was carried out to determine the frequencies of the D, C, c, E, e, K, k, Fy(a), Fy(b), Jk(a), Jk(b), M, N, S and s antigens in over 3,000 blood donors. Samples from randomly selected blood donors from Delhi and nearby areas (both voluntary and replacement) were collected for extended antigen typing during the period January 2009 to January 2010. Antigens were typed via automated testing on the Galileo instrument using commercial antisera. A total of 3073 blood samples from donors were phenotyped. The prevalence of these antigens was found to be as follows in %: D: 93.6, C: 87, c: 58, E: 20, e: 98, K: 3.5, k: 99.97, F(a) : 87.4, Fy(b) : 57.6, Jk(a) : 81.5, Jk(b) : 67.4, M: 88.7, N: 65.4, S: 54.8 and s: 88.7. This study found the prevalence of the typed antigens among Indian blood donors to be statistically different to those in the Caucasian, Black and Chinese populations, but more similar to Caucasians than to the other racial groups.

  5. Percentage of free serum prostate-specific antigen: a new tool in the early diagnosis of prostatic cancer.

    Science.gov (United States)

    Toubert, M E; Guillet, J; Chiron, M; Meria, P; Role, C; Schlageter, M H; Francois, H; Borschneck, C; Nivelon, F; Desgrandchamps, F; Rastel, D; Cussenot, O; Teillac, P; Le Duc, A; Najean, Y

    1996-11-01

    Prostate-specific antigen (PSA) is a protease able to bind to serum antiproteases as alpha 1 antichymotrypsin (ACT). Free PSA (FPSA) corresponds to the fraction of total PSA (TPSA) which is unbound to ACT. Specific detection of the FPSA seems to be a valuable tool in the distinction between prostatic cancer (PCa) and benign prostatic hyperplasia (BPH). Our aim was to evaluate retrospectively the FPSA/TPSA ratio in comparison to TPSA or FPSA determination, using two new immunoradiometric assays (PSA-RIACT and FPSA-RIACT, CIS bio international, Gif Sur Yvette, France) in the early diagnosis of PCa. 256 men, with TPSA levels between 0.7 and 44.7 ng/ml (median age = 69 years), including 164 sera obtained from patients with BPH and 92 sera from patients with untreated PCa were assayed. All diagnoses were histologically confirmed and patients tested before any adjuvant treatment. The evaluation of the median FPSA/TPSA ratio in the two groups showed significantly different values (BPH group: 24.2%, PCa group: 12.1%, P 10 ng/ml).

  6. A competitive-inhibiton radioimmunoassay for influenza virus envelope antigens

    International Nuclear Information System (INIS)

    Russ, G.; Styk, B.; Vareckova, E.; Polakova, K.

    1976-01-01

    A double-antibody competitive-inhibition radioimmunoassay for influenza virus envelope antigens is described. A viral antigen preparation from influenza A virus recombinant MRC11 [antigenically identical to A/Port Chalmers/1/73 (H3N2)] consisting of haemagglutinin and neuraminidase was labelled with radioiodine. Rabbit antisera were allowed to react with the labelled antigen and the resultant antigen-antibody complexes were precipitated with the appropriate antiglobulin. The competitive-inhibition radioimmunoassay very sensitively elucidated differences even among closely related influenza virus strains. Attempts have been made to eliminate neuraminidase from radioimmunoprecipitation to obtain a competitive-inhibition radioimmunoassay system for haemagglutinin alone. (author)

  7. Profiling Humoral Immune Responses to Clostridium difficile-Specific Antigens by Protein Microarray Analysis.

    Science.gov (United States)

    Negm, Ola H; Hamed, Mohamed R; Dilnot, Elizabeth M; Shone, Clifford C; Marszalowska, Izabela; Lynch, Mark; Loscher, Christine E; Edwards, Laura J; Tighe, Patrick J; Wilcox, Mark H; Monaghan, Tanya M

    2015-09-01

    Clostridium difficile is an anaerobic, Gram-positive, and spore-forming bacterium that is the leading worldwide infective cause of hospital-acquired and antibiotic-associated diarrhea. Several studies have reported associations between humoral immunity and the clinical course of C. difficile infection (CDI). Host humoral immune responses are determined using conventional enzyme-linked immunosorbent assay (ELISA) techniques. Herein, we report the first use of a novel protein microarray assay to determine systemic IgG antibody responses against a panel of highly purified C. difficile-specific antigens, including native toxins A and B (TcdA and TcdB, respectively), recombinant fragments of toxins A and B (TxA4 and TxB4, respectively), ribotype-specific surface layer proteins (SLPs; 001, 002, 027), and control proteins (tetanus toxoid and Candida albicans). Microarrays were probed with sera from a total of 327 individuals with CDI, cystic fibrosis without diarrhea, and healthy controls. For all antigens, precision profiles demonstrated ELISA in the quantification of antitoxin A and antitoxin B IgG. These results indicate that microarray is a suitable assay for defining humoral immune responses to C. difficile protein antigens and may have potential advantages in throughput, convenience, and cost. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  8. Bayesian nonparametric clustering in phylogenetics: modeling antigenic evolution in influenza.

    Science.gov (United States)

    Cybis, Gabriela B; Sinsheimer, Janet S; Bedford, Trevor; Rambaut, Andrew; Lemey, Philippe; Suchard, Marc A

    2018-01-30

    Influenza is responsible for up to 500,000 deaths every year, and antigenic variability represents much of its epidemiological burden. To visualize antigenic differences across many viral strains, antigenic cartography methods use multidimensional scaling on binding assay data to map influenza antigenicity onto a low-dimensional space. Analysis of such assay data ideally leads to natural clustering of influenza strains of similar antigenicity that correlate with sequence evolution. To understand the dynamics of these antigenic groups, we present a framework that jointly models genetic and antigenic evolution by combining multidimensional scaling of binding assay data, Bayesian phylogenetic machinery and nonparametric clustering methods. We propose a phylogenetic Chinese restaurant process that extends the current process to incorporate the phylogenetic dependency structure between strains in the modeling of antigenic clusters. With this method, we are able to use the genetic information to better understand the evolution of antigenicity throughout epidemics, as shown in applications of this model to H1N1 influenza. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  9. Expression of Hepatitis C Virus Core and E2 antigenic recombinant proteins and their use for development of diagnostic assays.

    Science.gov (United States)

    Ali, Amjad; Nisar, Muhammad; Idrees, Muhammad; Rafique, Shazia; Iqbal, Muhammad

    2015-05-01

    Early diagnosis of HCV infection is based on detection of antibodies against HCV proteins using recombinant viral antigens. The present study was designed to select, clone and express the antigenic regions of Core and E2 genes from local HCV-3a genotype and to utilize the antigenic recombinant proteins (Core & E2) to develop highly sensitive, specific and economical diagnostic assays for detection of HCV infection. The antigenic sites were determined within Core and E2 genes and were then cloned in pET-28a expression vector. The right orientation of the desired inserted fragments of Core and E2 were confirmed via sequencing prior to expression and were then transformed in BL21 (DE3) pLysS strains of E. coli and induced with 0.5mM Isopropyl-b-D-thiogalactopyranoside (IPTG) for the production of antigenic recombinant proteins. The produced truncated antigens were then purified by Nickel affinity chromatography and were confirmed by western blotting, immunoblotting and enzyme-linked immunosorbent assay (ELISA). The expressed Core and E2 recombinant antigens were used to develop immunoblotting assay for the detection of anti-HCV antibodies in sera. With immunoblotting, a total of 93-HCV infected sera and 35-HCV negative individuals were tested for the presence of anti-HCV antibodies to the Core and E2 antigens. Recombinant antigen showed 100% reactivity against HCV infected sera, with no cross reactivity against HCV-negative sera. The immunoblot assay mixture of recombinant antigens (Core+E2) showed a strong reaction intensity in the test area (TA) as compared to the individual truncated Core and E2 recombinant antigens. In the in-house ELISA assay, mixed Core and E2 recombinant antigens showed 100% reactivity against a standardized panel of 150-HCV-positive sera and non reactivity against a standardized panel of 150 HCV-negative sera while also being non reactive to sera positive for other viral infections. The antigenic recombinant antigens also were tested for the

  10. Antibody-antigen-adjuvant conjugates enable co-delivery of antigen and adjuvant to dendritic cells in cis but only have partial targeting specificity

    NARCIS (Netherlands)

    Kreutz, M.; Giquel, B.; Hu, Q.; Abuknesha, R.; Uematsu, S.; Akira, S.; Nestle, F.O.; Diebold, S.S.

    2012-01-01

    Antibody-antigen conjugates, which promote antigen-presentation by dendritic cells (DC) by means of targeted delivery of antigen to particular DC subsets, represent a powerful vaccination approach. To ensure immunity rather than tolerance induction the co-administration of a suitable adjuvant is

  11. Expression of variant surface antigens by Plasmodium falciparum parasites in the peripheral blood of clinically immune pregnant women indicates ongoing placental infection

    DEFF Research Database (Denmark)

    Ofori, Michael F; Staalsoe, Trine; Bam, Victoria

    2003-01-01

    Placenta-sequestered Plasmodium falciparum parasites that cause pregnancy-associated malaria (PAM) in otherwise clinically immune women express distinct variant surface antigens (VSA(PAM)) not expressed by parasites in nonpregnant individuals. We report here that parasites from the peripheral blood...... of clinically immune pregnant women also express VSA(PAM), making them a convenient source of VSA(PAM) expressors for PAM vaccine research....

  12. Construction, expression, purification and biotin labeling of a single recombinant multi-epitope antigen for double-antigen sandwich ELISA to detect hepatitis C virus antibody.

    Science.gov (United States)

    He, Jing; Xiu, Bingshui; Wang, Guohua; Chen, Kun; Feng, Xiaoyan; Song, Xiaoguo; Zhu, Cuixia; Yang, Xiqin; Bai, Guanzhong; Ling, Shigan; Zhang, Heqiu

    2011-08-01

    Based on B cell epitope predictions, a recombinant antigen with multiple epitopes from four Hepatitis C Virus fragments (C, NS3, NS4 and NS5) were engineered. The recombinant gene was then highly expressed in E. coli. The non-modified and C-terminal-modified recombinant proteins were used for coating and biotin labeling, respectively, to establish the double-antigen sandwich ELISA. Ten positive reference samples confirmed by the CHIRON RIBA HCV 3.0 SIA kit were detected positive, Forty one plasma samples were positive among samples from 441 volunteers, which indicated that the recombinant antigen could readily react well with plasma HCV antibody. As critical reagents of double-antigen sandwich ELISA, the recombinant multi-epitope antigen and the C-terminal-modified and biotin-conjugated antigen show good antigenicity. In this study, we provide a simple approach to produce multiple epitopes within one recombinant protein in order to avoid the costly expression of less-effective pools of multiple proteins, which is the conventional strategy of diagnostic antigen production for HCV antibody detection.

  13. Bat Caliciviruses and Human Noroviruses Are Antigenically Similar and Have Overlapping Histo-Blood Group Antigen Binding Profiles.

    Science.gov (United States)

    Kocher, Jacob F; Lindesmith, Lisa C; Debbink, Kari; Beall, Anne; Mallory, Michael L; Yount, Boyd L; Graham, Rachel L; Huynh, Jeremy; Gates, J Edward; Donaldson, Eric F; Baric, Ralph S

    2018-05-22

    Emerging zoonotic viral diseases remain a challenge to global public health. Recent surveillance studies have implicated bats as potential reservoirs for a number of viral pathogens, including coronaviruses and Ebola viruses. Caliciviridae represent a major viral family contributing to emerging diseases in both human and animal populations and have been recently identified in bats. In this study, we blended metagenomics, phylogenetics, homology modeling, and in vitro assays to characterize two novel bat calicivirus (BtCalV) capsid sequences, corresponding to strain BtCalV/A10/USA/2009, identified in Perimyotis subflavus near Little Orleans, MD, and bat norovirus. We observed that bat norovirus formed virus-like particles and had epitopes and receptor-binding patterns similar to those of human noroviruses. To determine whether these observations stretch across multiple bat caliciviruses, we characterized a novel bat calicivirus, BtCalV/A10/USA/2009. Phylogenetic analysis revealed that BtCalV/A10/USA/2009 likely represents a novel Caliciviridae genus and is most closely related to "recoviruses." Homology modeling revealed that the capsid sequences of BtCalV/A10/USA/2009 and bat norovirus resembled human norovirus capsid sequences and retained host ligand binding within the receptor-binding domains similar to that seen with human noroviruses. Both caliciviruses bound histo-blood group antigens in patterns that overlapped those seen with human and animal noroviruses. Taken together, our results indicate the potential for bat caliciviruses to bind histo-blood group antigens and overcome a significant barrier to cross-species transmission. Additionally, we have shown that bat norovirus maintains antigenic epitopes similar to those seen with human noroviruses, providing further evidence of evolutionary descent. Our results reiterate the importance of surveillance of wild-animal populations, especially of bats, for novel viral pathogens. IMPORTANCE Caliciviruses are

  14. The effect of HLA mismatches, shared cross-reactive antigen groups, and shared HLA-DR antigens on the outcome after pediatric liver transplantation

    NARCIS (Netherlands)

    Sieders, E; Hepkema, BG; Peeters, PMJG; Ten Vergert, EM; De Jong, KP; Porte, RJ; Bijleveld, CMA; van den Berg, AP; Lems, SPM; Gouw, ASH; Slooff, MJH

    2005-01-01

    The aim of this study was to analyze the effect of human leukocyte antigen (HLA) class I and HLA-DR mismatching, sharing cross-reactive antigen groups (CREGs), and sharing HLA-DR antigens on the outcome after pediatric liver transplantation. Outcome parameters were graft survival, acute rejection,

  15. Cerium oxide-deposited mesoporous silica nanoparticles for the determination of carcinoembryonic antigen in serum using inductively coupled plasma-mass spectrometry

    International Nuclear Information System (INIS)

    Choi, H.W.; Lee, K.H.; Hur, N.H.; Lim, H.B.

    2014-01-01

    Highlights: • Sandwich-type immunoassay using ICP-MS and nanoparticles to determine biomarkers. • CeO 2 -deposited mesoporous silica nanoparticles were synthesized as a probe. • Ratiometric measurement significantly improved the calibration linearity. • Excellent detection limit was achieved by signal amplification. - Abstract: CeO 2 -deposited mesoporous silica nanoparticles were synthesized as a probe to determine carcinoembryonic antigen (CEA) in serum by inductively coupled plasma-mass spectrometry (ICP-MS). The prepared mesoporous nanoparticles were modified and tagged to the target for sandwich-type immunoassay. Fe 3 O 4 magnetic nanoparticles (MNPs) were also synthesized and immobilized with antibody to extract the target biomarker. The calibration curve of the synthesized CeO 2 -deposited silica nanoparticles, which was plotted by the signal ratio of 140 Ce/ 57 Fe measured by ICP-MS vs. the concentration of CEA, showed excellent linearity and sensitivity owing to the signal amplification and low spectral interference. Under optimal conditions, the sandwich-type analytical method was applied to determine CEA in serum spiked in the range of 0.001–5 ng mL −1 and showed a limit of detection of 0.36 ng mL −1 . Since the deposited CeO 2 in the mesoporous silica layer can be substituted by other metal compounds, various kinds of metal-deposited nanoparticles can be prepared as probe materials for multiplex detection in bioanalysis

  16. Original antigenic sin: A comprehensive review.

    Science.gov (United States)

    Vatti, Anup; Monsalve, Diana M; Pacheco, Yovana; Chang, Christopher; Anaya, Juan-Manuel; Gershwin, M Eric

    2017-09-01

    The concept of "original antigenic sin" was first proposed by Thomas Francis, Jr. in 1960. This phenomenon has the potential to rewrite what we understand about how the immune system responds to infections and its mechanistic implications on how vaccines should be designed. Antigenic sin has been demonstrated to occur in several infectious diseases in both animals and humans, including human influenza infection and dengue fever. The basis of "original antigenic sin" requires immunological memory, and our immune system ability to autocorrect. In the context of viral infections, it is expected that if we are exposed to a native strain of a pathogen, we should be able to mount a secondary immune response on subsequent exposure to the same pathogen. "Original antigenic sin" will not contradict this well-established immunological process, as long as the subsequent infectious antigen is identical to the original one. But "original antigenic sin" implies that when the epitope varies slightly, then the immune system relies on memory of the earlier infection, rather than mount another primary or secondary response to the new epitope which would allow faster and stronger responses. The result is that the immunological response may be inadequate against the new strain, because the immune system does not adapt and instead relies on its memory to mount a response. In the case of vaccines, if we only immunize to a single strain or epitope, and if that strain/epitope changes over time, then the immune system is unable to mount an accurate secondary response. In addition, depending of the first viral exposure the secondary immune response can result in an antibody-dependent enhancement of the disease or at the opposite, it could induce anergy. Both of them triggering loss of pathogen control and inducing aberrant clinical consequences. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Antigen injection (image)

    Science.gov (United States)

    Leprosy is caused by the organism Mycobacterium leprae . The leprosy test involves injection of an antigen just under ... if your body has a current or recent leprosy infection. The injection site is labeled and examined ...

  18. Strategy for eliciting antigen-specific CD8+ T cell-mediated immune response against a cryptic CTL epitope of merkel cell polyomavirus large T antigen

    Directory of Open Access Journals (Sweden)

    Gomez Bianca P

    2012-10-01

    Full Text Available Abstract Background Merkel cell carcinoma (MCC is a relatively new addition to the expanding category of oncovirus-induced cancers. Although still comparably rare, the number of cases has risen dramatically in recent years. Further complicating this trend is that MCC is an extremely aggressive neoplasm with poor patient prognosis and limited treatment options for advanced disease. The causative agent of MCC has been identified as the merkel cell polyomavirus (MCPyV. The MCPyV-encoded large T (LT antigen is an oncoprotein that is theorized to be essential for virus-mediated tumorigenesis and is therefore, an excellent MCC antigen for the generation of antitumor immune responses. As a foreign antigen, the LT oncoprotein avoids the obstacle of immune tolerance, which normally impedes the development of antitumor immunity. Ergo, it is an excellent target for anti-MCC immunotherapy. Since tumor-specific CD8+ T cells lead to better prognosis for MCC and numerous other cancers, we have generated a DNA vaccine that is capable of eliciting LT-specific CD8+ T cells. The DNA vaccine (pcDNA3-CRT/LT encodes the LT antigen linked to a damage-associated molecular pattern, calreticulin (CRT, as it has been demonstrated that the linkage of CRT to antigens promotes the induction of antigen-specific CD8+ T cells. Results The present study shows that DNA vaccine-induced generation of LT-specific CD8+ T cells is augmented by linking CRT to the LT antigen. This is relevant since the therapeutic effects of the pcDNA3-CRT/LT DNA vaccine is mediated by LT-specific CD8+ T cells. Mice vaccinated with the DNA vaccine produced demonstrably more LT-specific CD8+ T cells. The DNA vaccine was also able to confer LT-specific CD8+ T cell-mediated protective and therapeutic effects to prolong the survival of mice with LT-expressing tumors. In the interest of determining the LT epitope which most MCC-specific CD8+ T cells recognize, we identified the amino acid sequence of the

  19. The MASP family of Trypanosoma cruzi: changes in gene expression and antigenic profile during the acute phase of experimental infection.

    Directory of Open Access Journals (Sweden)

    Sara Lopes dos Santos

    Full Text Available Trypanosoma cruzi is the etiological agent of Chagas disease, a debilitating illness that affects millions of people in the Americas. A major finding of the T. cruzi genome project was the discovery of a novel multigene family composed of approximately 1,300 genes that encode mucin-associated surface proteins (MASPs. The high level of polymorphism of the MASP family associated with its localization at the surface of infective forms of the parasite suggests that MASP participates in host-parasite interactions. We speculate that the large repertoire of MASP sequences may contribute to the ability of T. cruzi to infect several host cell types and/or participate in host immune evasion mechanisms.By sequencing seven cDNA libraries, we analyzed the MASP expression profile in trypomastigotes derived from distinct host cells and after sequential passages in acutely infected mice. Additionally, to investigate the MASP antigenic profile, we performed B-cell epitope prediction on MASP proteins and designed a MASP-specific peptide array with 110 putative epitopes, which was screened with sera from acutely infected mice.We observed differential expression of a few MASP genes between trypomastigotes derived from epithelial and myoblast cell lines. The more pronounced MASP expression changes were observed between bloodstream and tissue-culture trypomastigotes and between bloodstream forms from sequential passages in acutely infected mice. Moreover, we demonstrated that different MASP members were expressed during the acute T. cruzi infection and constitute parasite antigens that are recognized by IgG and IgM antibodies. We also found that distinct MASP peptides could trigger different antibody responses and that the antibody level against a given peptide may vary after sequential passages in mice. We speculate that changes in the large repertoire of MASP antigenic peptides during an infection may contribute to the evasion of host immune responses during the

  20. A small antigenic determinant of the Chikungunya virus E2 protein is sufficient to induce neutralizing antibodies which are partially protective in mice.

    Directory of Open Access Journals (Sweden)

    Christopher Weber

    2015-04-01

    Full Text Available The mosquito-borne Chikungunya virus (CHIKV causes high fever and severe joint pain in humans. It is expected to spread in the future to Europe and has recently reached the USA due to globalization, climate change and vector switch. Despite this, little is known about the virus life cycle and, so far, there is no specific treatment or vaccination against Chikungunya infections. We aimed here to identify small antigenic determinants of the CHIKV E2 protein able to induce neutralizing immune responses.E2 enables attachment of the virus to target cells and a humoral immune response against E2 should protect from CHIKV infections. Seven recombinant proteins derived from E2 and consisting of linear and/or structural antigens were created, and were expressed in and purified from E. coli. BALB/c mice were vaccinated with these recombinant proteins and the mouse sera were screened for neutralizing antibodies. Whereas a linear N-terminally exposed peptide (L and surface-exposed parts of the E2 domain A (sA alone did not induce neutralizing antibodies, a construct containing domain B and a part of the β-ribbon (called B+ was sufficient to induce neutralizing antibodies. Furthermore, domain sA fused to B+ (sAB+ induced the highest amount of neutralizing antibodies. Therefore, the construct sAB+ was used to generate a recombinant modified vaccinia virus Ankara (MVA, MVA-CHIKV-sAB+. Mice were vaccinated with MVA-CHIKV-sAB+ and/or the recombinant protein sAB+ and were subsequently challenged with wild-type CHIKV. Whereas four vaccinations with MVA-CHIKV-sAB+ were not sufficient to protect mice from a CHIKV infection, protein vaccination with sAB+ markedly reduced the viral titers of vaccinated mice.The recombinant protein sAB+ contains important structural antigens for a neutralizing antibody response in mice and its formulation with appropriate adjuvants might lead to a future CHIKV vaccine.

  1. The prognostic significance of preoperative serum cancer antigen 15-3 levels in endometrial carcinomas

    Science.gov (United States)

    Tas, Emre E.; Yavuz, Ayse F.

    2017-01-01

    Objectives: To determine the associations between serum cancer antigen 15-3 levels and prognostic factors in patients with endometrial carcinomas. Additionally, we investigated the clinical utility of serum cancer antigen 15-3 levels in the selection of low-risk patients with endometrioid type, tumor size <2 cm, myometrial invasion ≤50%, and histological grade 1-2. Methods: Ninety-six patients, who were surgically staged at Ankara Yildirim Beyazit University, Ankara, Turkey, between 2007 and 2016, were retrospectively analyzed. Demographic, clinical, and surgical characteristics were retrieved from the patients’ hospital records. A p<0.05 was considered significant. Results: Fifteen patients had advanced (≥Stage II) disease, 14 patients had Type 2 histology, 20 patients had Grade 3 tumors, 23 patients had lymphovascular space invasion, and 10 patients had positive lymph node involvement. Serum cancer antigen 15-3 levels were significantly higher in patients with advanced (≥Stage II) disease, Type 2 histology, Grade 3 tumors, lymp°hovascular space invasion, and positive lymph node involvement (p<0.05). Serum cancer antigen 15-3 levels were also significantly correlated with tumor size (p=0.006). Serum cancer antigen 15-3 levels were significantly lower (95% confidence interval: 0.57−0.79; p=0.03) in low-risk patients compared to other endometrial carcinoma patients. A cutoff of 25.0 IU/mL was used to identify high-risk patients with a specificity of 100%. Conclusion: Serum cancer antigen 15-3 levels significantly correlated with prognostic factors and were a useful diagnostic tool for endometrial carcinomas. PMID:29114696

  2. The prognostic significance of preoperative serum cancer antigen 15-3 levels in endometrial carcinomas

    Directory of Open Access Journals (Sweden)

    Emre E. Tas

    2017-11-01

    Full Text Available Objectives: To determine the associations between serum cancer antigen 15-3 levels and prognostic factors in patients with endometrial carcinomas. Additionally, we investigated the clinical utility of serum cancer antigen 15-3 levels in the selection of low-risk patients with endometrioid type, tumor size less than 2 cm, myometrial invasion ≤50%, and histological grade 1-2. Methods: Ninety-six patients, who were surgically staged at Ankara Yildirim Beyazit University, Ankara, Turkey, between 2007 and 2016, were retrospectively analyzed. Demographic, clinical, and surgical characteristics were retrieved from the patients’ hospital records. A p less than 0.05 was considered significant. Results: Fifteen patients had advanced (≥Stage II disease, 14 patients had Type 2 histology, 20 patients had Grade 3 tumors, 23 patients had lymphovascular space invasion, and 10 patients had positive lymph node involvement. Serum cancer antigen 15-3 levels were significantly higher in patients with advanced (≥Stage II disease, Type 2 histology, Grade 3 tumors, lymphovascular space invasion, and positive lymph node involvement (p less than 0.05. Serum cancer antigen 15-3 levels were also significantly correlated with tumor size (p=0.006. Serum cancer antigen 15-3 levels were significantly lower (95% confidence interval: 0.57−0.79; p=0.03 in low-risk patients compared to other endometrial carcinoma patients. A cutoff of 25.0 IU/mL was used to identify high-risk patients with a specificity of 100%. Conclusion: Serum cancer antigen 15-3 levels significantly correlated with prognostic factors and were a useful diagnostic tool for endometrial carcinomas.

  3. Reduction of T-Helper Cell Responses to Recall Antigen Mediated by Codelivery with Peptidoglycan via the Intestinal Nanomineral–Antigen Pathway

    Science.gov (United States)

    Hewitt, Rachel E.; Robertson, Jack; Haas, Carolin T.; Pele, Laetitia C.; Powell, Jonathan J.

    2017-01-01

    Naturally occurring intestinal nanomineral particles constituently form in the mammalian gut and trap luminal protein and microbial components. These cargo loaded nanominerals are actively scavenged by M cells of intestinal immune follicles, such as Peyer’s patches and are passed to antigen-presenting cells. Using peripheral blood mononuclear cell populations as an in vitro model of nanomineral uptake and antigen presentation, we show that monocytes avidly phagocytose nanomineral particles bearing antigen and peptidoglycan (PGN), and that the presence of PGN within particles downregulates their cell surface MHC class II and upregulates programmed death receptor ligand 1. Nanomineral delivery of antigen suppresses antigen-specific CD4+ T cell responses, an effect that is enhanced in the presence of PGN. Blocking the interleukin-10 receptor restores CD4+ T cell responses to antigen codelivered with PGN in nanomineral form. Using human intestinal specimens, we have shown that the in vivo nanomineral pathway operates in an interleukin-10 rich environment. Consequently, the delivery of a dual antigen–PGN cargo by endogenous nanomineral in vivo is likely to be important in the establishment of intestinal tolerance, while their synthetic mimetics present a potential delivery system for therapeutic applications targeting the modulation of Peyer’s patch T cell responses. PMID:28367148

  4. [The isolation and evaluation of Aspergillus fumigatus antigens].

    Science.gov (United States)

    Lirio, V de S; de Assis, C M; Cano, M I; Lacaz, C da S

    1992-01-01

    Antigens from three strains of Aspergillus fumigatus (354, 356, and JIG) and an antiserum against the mixing of these antigens have been produced, and evaluated immunochemically. The antigens were obtained through a modified Coleman & Kaufman technique (culture filtrate concentrated by acetone). Analysis by the immunodiffusion test (ID) against homologous serum has yielded 100% sensitivity (with the studied sera). Concerning heterologous sera we found reactivity with a serum of a patient of candidiasis and another with histoplasmosis. The same result was obtained with a reference antigen in immunodiffusion, showing similar standards of response. Titration of the antiserum by ID and counterimmunoelectrophoresis showed a title of 1:32, and by complement fixation (micro-technique) a title of 1:128. Using immunoelectrophoresis (IEF), the produced antiserum yielded 8 lines of precipitation (5 in the anodic pole and 3 in the cathodic one). In SDS-PAGE at 12.5% the antigen has presented a rather complex electrophoretic profile (26 proteic subunits with a molecular weight ranging from 18 a > 100 kDa). Immunogenicity of the antigen was observed in all fractions of SDS-PAGE when the immunoblotting against the antiserum was carried out.

  5. Structure-based non-canonical amino acid design to covalently crosslink an antibody–antigen complex

    Science.gov (United States)

    Xu, Jianqing; Tack, Drew; Hughes, Randall A.; Ellington, Andrew D.; Gray, Jeffrey J.

    2014-01-01

    Engineering antibodies to utilize non-canonical amino acids (NCAA) should greatly expand the utility of an already important biological reagent. In particular, introducing crosslinking reagents into antibody complementarity determining regions (CDRs) should provide a means to covalently crosslink residues at the antibody–antigen interface. Unfortunately, finding the optimum position for crosslinking two proteins is often a matter of iterative guessing, even when the interface is known in atomic detail. Computer-aided antibody design can potentially greatly restrict the number of variants that must be explored in order to identify successful crosslinking sites. We have therefore used Rosetta to guide the introduction of an oxidizable crosslinking NCAA, l-3,4-dihydroxyphenylalanine (l-DOPA), into the CDRs of the anti-protective antigen scFv antibody M18, and have measured crosslinking to its cognate antigen, domain 4 of the anthrax protective antigen. Computed crosslinking distance, solvent accessibility, and interface energetics were three factors considered that could impact the efficiency of l-DOPA-mediated crosslinking. In the end, 10 variants were synthesized, and crosslinking efficiencies were generally 10% or higher, with the best variant crosslinking to 52% of the available antigen. The results suggest that computational analysis can be used in a pipeline for engineering crosslinking antibodies. The rules learned from l-DOPA crosslinking of antibodies may also be generalizable to the formation of other crosslinked interfaces and complexes. PMID:23680795

  6. Minor Antigen Disparities Impede Induction of Long Lasting Chimerism and Tolerance through Bone Marrow Transplantation with Costimulation Blockade

    Directory of Open Access Journals (Sweden)

    Sinda Bigenzahn

    2016-01-01

    Full Text Available Mixed chimerism and tolerance can be successfully induced in rodents through allogeneic bone marrow transplantation (BMT with costimulation blockade (CB, but varying success rates have been reported with distinct models and protocols. We therefore investigated the impact of minor antigen disparities on the induction of mixed chimerism and tolerance. C57BL/6 (H2b mice received nonmyeloablative total body irradiation (3 Gy, costimulation blockade (anti-CD40L mAb and CTLA4Ig, and 2×107 bone marrow cells (BMC from either of three donor strains: Balb/c (H2d (MHC plus multiple minor histocompatibility antigen (mHAg mismatched, B10.D2 (H2d or B10.A (H2a (both MHC mismatched, but mHAg matched. Macrochimerism was followed over time by flow cytometry and tolerance was tested by skin grafting. 20 of 21 recipients of B10.D2 BMC but only 13 of 18 of Balb/c BMC and 13 of 20 of B10.A BMC developed stable long-term multilineage chimerism (p<0.05 for each donor strain versus B10.D2. Significantly superior donor skin graft survival was observed in successfully established long-term chimeras after mHAg matched BMT compared to mHAg mismatched BMT (p<0.05. Both minor and major antigen disparities pose a substantial barrier for the induction of chimerism while the maintenance of tolerance after nonmyeloablative BMT and costimulation blockade is negatively influenced by minor antigen disparities.

  7. Heritability of antibody isotype and subclass responses to Plasmodium falciparum antigens.

    Directory of Open Access Journals (Sweden)

    Nancy O Duah

    2009-10-01

    Full Text Available It is important to understand the extent to which genetic factors regulate acquired immunity to common infections. A classical twin study design is useful to estimate the heritable component of variation in measurable immune parameters.This study assessed the relative heritability of different plasma antibody isotypes and subclasses (IgG1, IgG2, IgG3, IgG4, IgM, IgA and IgE naturally acquired to P. falciparum blood stage antigens AMA1, MSP1-19, MSP2 (two allelic types and MSP3 (two allelic types. Separate analyses were performed on plasma from 213 pairs of Gambian adult twins, 199 child twin pairs sampled in a dry season when there was little malaria transmission, and another set of 107 child twin pairs sampled at the end of the annual wet season when malaria was common. There were significantly positive heritability (h(2 estimates for 48% (20/42 of the specific antibody assays (for the seven isotypes and subclasses to the six antigens tested among the adults, 48% (20/42 among the children in the dry season and 31% (13/42 among the children in the wet season. In children, there were significant heritability estimates for IgG4 reactivity against each of the antigens, and this subclass had higher heritability than the other subclasses and isotypes. In adults, 75% (15/20 of the significantly heritable antigen-specific isotype responses were attributable to non-HLA class II genetic variation, whereas none showed a significant HLA contribution.Genome-wide approaches are now warranted to map the major genetic determinants of variable antibody isotype and subclass responses to malaria, alongside evaluation of their impact on infection and disease. Although plasma levels of IgG4 to malaria antigens are generally low, the exceptionally high heritability of levels of this subclass in children deserves particular investigation.

  8. Review of Mycobacteriumavium subsp. paratuberculosis antigen candidates with diagnostic potential

    DEFF Research Database (Denmark)

    Mikkelsen, Heidi; Aagaard, Claus; Nielsen, Søren Saxmose

    2011-01-01

    antigens, heat shock antigens and hypothetical antigens. Strategies for evaluation of novel antigen candidates are discussed critically. Relatively few of the described antigens were evaluated for their use in CMI based diagnostic assays and so far, no obvious candidate has been identified...... to development of antibodies and shedding of detectable amounts of MAP. At present, available diagnostic assays are limited by the lack of MAP specific antigens included in these assays resulting in poor specificity. The objective of this review is to provide a systematic overview of diagnostic MAP antigen...... faeces; however, these diagnostic tools are often not applicable until years after infection. Detection of MAP specific cell-mediated immune (CMI) responses can serve as an alternative and be implemented in a diagnostic tool. CMI responses can be measured at an early stage of infection, prior...

  9. The fate of heterologous antigen (131I-HSA) in the organs of chickens exposed to total-body X-irradiation before a secondary antigenic stimulus

    International Nuclear Information System (INIS)

    Prohazka, Z.; Hampl, J.; Krejci, J.

    1975-01-01

    A study was made on the effect of ionizing radiation on the rate of elimination of 131 I-labelled human serum albumin from the blood and its organ deposition in chickens exposed to 1200 R (LD 50 ) at various intervals before secondary antigen injection. In unirradiated control chickens, the elimination of antigen after its secondary injection followed the typical three-phase pattern, characterized by an early onset and a rapid progress of the third phase. The elimination curve from irradiated birds paralleled rather closely that from the controls during the first and second phases while the phase of immune elimination was hardly perceptible. No major differences were found between the individual irrradiated groups. The irradiated birds also showed less formation of antibodies and antigen-antibody complexes and a lower antigen content of the organs than the unirradiated controls. From the results it appears that the specific antigen uptake from the blood of chickens during the first and second phases of elimination of a secondary dose of antigen is radioresistant; the temporal relation between X-irradiation and secondary antigen injection does not play a substantial role in impairment of the secondary antibody response to soluble antigens in chickens

  10. Patterns of predicted T-cell epitopes associated with antigenic drift in influenza H3N2 hemagglutinin.

    Directory of Open Access Journals (Sweden)

    E Jane Homan

    Full Text Available Antigenic drift allowing escape from neutralizing antibodies is an important feature of transmission and survival of influenza viruses in host populations. Antigenic drift has been studied in particular detail for influenza A H3N2 and well defined antigenic clusters of this virus documented. We examine how host immunogenetics contributes to determination of the antibody spectrum, and hence the immune pressure bringing about antigenic drift. Using uTOPE™ bioinformatics analysis of predicted MHC binding, based on amino acid physical property principal components, we examined the binding affinity of all 9-mer and 15-mer peptides within the hemagglutinin 1 (HA1 of 447 H3N2 virus isolates to 35 MHC-I and 14 MHC-II alleles. We provide a comprehensive map of predicted MHC-I and MHC-II binding affinity for a broad array of HLA alleles for the H3N2 influenza HA1 protein. Each HLA allele exhibited a characteristic predicted binding pattern. Cluster analysis for each HLA allele shows that patterns based on predicted MHC binding mirror those described based on antibody binding. A single amino acid mutation or position displacement can result in a marked difference in MHC binding and hence potential T-helper function. We assessed the impact of individual amino acid changes in HA1 sequences between 10 virus isolates from 1968-2002, representative of antigenic clusters, to understand the changes in MHC binding over time. Gain and loss of predicted high affinity MHC-II binding sites with cluster transitions were documented. Predicted high affinity MHC-II binding sites were adjacent to antibody binding sites. We conclude that host MHC diversity may have a major determinant role in the antigenic drift of influenza A H3N2.

  11. 21 CFR 660.1 - Antibody to Hepatitis B Surface Antigen.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 7 2010-04-01 2010-04-01 false Antibody to Hepatitis B Surface Antigen. 660.1... Hepatitis B Surface Antigen § 660.1 Antibody to Hepatitis B Surface Antigen. (a) Proper name and definition. The proper name of this product shall be Antibody to Hepatitis B Surface Antigen. The product is...

  12. Identification of candidate vaccine antigens of bovine hemoparasites Theileria parva and Babesia bovis by use of helper T cell clones.

    Science.gov (United States)

    Brown, W C; Zhao, S; Logan, K S; Grab, D J; Rice-Ficht, A C

    1995-03-01

    Current vaccines for bovine hemoparasites utilize live attenuated organisms or virulent organisms administered concurrently with antiparasitic drugs. Although such vaccines can be effective, for most hemoparasites the mechanisms of acquired resistance to challenge infection with heterologous parasite isolates have not been clearly defined. Selection of potentially protective antigens has traditionally made use of antibodies to identify immunodominant proteins. However, numerous studies have indicated that induction of high antibody titers neither predicts the ability of an antigen to confer protective immunity nor correlates with protection. Because successful parasites have evolved antibody evasion tactics, alternative strategies to identify protective immunogens should be used. Through the elaboration of cytokines, T helper 1-(Th1)-like T cells and macrophages mediate protective immunity against many intracellular parasites, and therefore most likely play an important role in protective immunity against bovine hemoparasites. CD4+ T cell clones specific for soluble or membrane antigens of either Theileria parva schizonts or Babesia bovis merozoites were therefore employed to identify parasite antigens that elicit strong Th cell responses in vitro. Soluble cytosolic parasite antigen was fractionated by gel filtration, anion exchange chromatography or hydroxylapatite chromatography, or a combination thereof, and fractions were tested for the ability to induce proliferation of Th cell clones. This procedure enabled the identification of stimulatory fractions containing T. parva proteins of approximately 10 and 24 kDa. Antisera raised against the purified 24 kDa band reacted with a native schizont protein of approximately 30 kDa. Babesia bovis-specific Th cell clones tested against fractionated soluble Babesia bovis merozoite antigen revealed the presence of at least five distinct antigenic epitopes. Proteins separated by gel filtration revealed four patterns of

  13. Co-delivery of antigen and IL-12 by Venezuelan equine encephalitis virus replicon particles enhances antigen-specific immune responses and anti-tumor effects

    Science.gov (United States)

    Osada, Takuya; Berglund, Peter; Morse, Michael A.; Hubby, Bolyn; Lewis, Whitney; Niedzwiecki, Donna; Hobeika, Amy; Burnett, Bruce; Devi, Gayathri R.; Clay, Timothy M.; Smith, Jonathan; Lyerly, H. Kim

    2013-01-01

    We recently demonstrated that Venezuelan equine encephalitis (VEE) virus-based replicon particles (VRP) encoding tumor antigens could break tolerance in the immunomodulatory environment of advanced cancer. We hypothesized that local injection of VRP expressing Interleukin-12 (IL-12) at the site of injections of VRP-based cancer vaccines would enhance the tumor-antigen-specific T cell and antibody responses and anti-tumor efficacy. Mice were immunized with VRP encoding the human tumor-associated antigen, carcinoembryonic antigen (CEA) (VRP-CEA(6D)) and VRP-IL-12 was also administered at the same site or at a distant location. CEA-specific T cell and antibody responses were measured. To determine antitumor activity, mice were implanted with MC38-CEA-2 cells and immunized with VRP-CEA with and without VRP-IL-12 and tumor growth and mouse survival were measured. VRP-IL-12 greatly enhanced CEA-specific T cell and antibody responses when combined with VRP-CEA(6D) vaccination. VRP IL-12 was superior to IL-12 protein at enhancing immune responses. Vaccination with VRP-CEA(6D) plus VRP-IL-12 was superior to VRP-CEA(6D) or VRP-IL-12 alone in inducing anti-tumor activity and prolonging survival in tumor-bearing mice. Importantly, local injection of VRP-IL-12 at the VRP-CEA(6D) injection site provided more potent activation of CEA-specific immune responses than VRP-IL-12 injected at a distant site from the VRP-CEA injections. Together, this study shows that VRP-IL-12 enhances vaccination with VRP-CEA(6D) and was more effective at activating CEA-specific T cell responses when locally expressed at the vaccine site. Clinical trials evaluating the adjuvant effect of VRP-IL-12 at enhancing the immunogenicity of cancer vaccines are warranted. PMID:22488274

  14. Polymer nanoparticles for cross-presentation of exogenous antigens and enhanced cytotoxic T-lymphocyte immune response

    Directory of Open Access Journals (Sweden)

    Song C

    2016-08-01

    Full Text Available Chanyoung Song,* Young-Woock Noh,* Yong Taik Lim SKKU Advanced Institute of Nanotechnology (SAINT, School of Chemical Engineering, Sungkyunkwan University, Suwon, South Korea *These authors contributed equally to this work Abstract: Effective induction of an antigen-specific cytotoxic T lymphocyte (CTL immune response is one of the key goals of cancer immunotherapy. We report the design and fabrication of polyethylenimine (PEI-coated polymer nanoparticles (NPs as efficient antigen-delivery carriers that can induce antigen cross-presentation and a strong CTL response. After synthesis of poly(d,l-lactide-co-glycolide (PLGA NPs containing ovalbumin (OVA by the double-emulsion solvent-evaporation method, cationic-charged PLGA NPs were generated by coating them with PEI. In a methyl tetrazolium salt assay, no discernible cytotoxic effect of PEI-coated PLGA (OVA NPs was observed. The capacity and mechanism of PEI-coated PLGA (OVA NPs for antigen delivery and cross-presentation on dendritic cells (DCs were determined by fluorescence microscopy and flow cytometry. PEI-coated PLGA (OVA NPs were internalized efficiently via phagocytosis or macropinocytosis in DCs and induced efficient cross-presentation of the antigen on MHC class I molecules via both endosome escape and a lysosomal processing mechanism. The DCs treated with PEI-coated PLGA (OVA NPs induced a release of IL-2 cytokine from OVA-specific CD8-OVA1.3 T cells more efficiently than DCs treated with PLGA (OVA NPs. Therefore, the PEI-coated PLGA (OVA NPs can induce antigen cross-presentation and are expected to be used for induction of a strong CTL immune response and for efficient anticancer immunotherapy. Keywords: antigen delivery, dendritic cells, polymer NPs, vaccine, cross-presentation

  15. Thermodynamics of antibody-antigen interaction revealed by mutation analysis of antibody variable regions.

    Science.gov (United States)

    Akiba, Hiroki; Tsumoto, Kouhei

    2015-07-01

    Antibodies (immunoglobulins) bind specific molecules (i.e. antigens) with high affinity and specificity. In order to understand their mechanisms of recognition, interaction analysis based on thermodynamic and kinetic parameters, as well as structure determination is crucial. In this review, we focus on mutational analysis which gives information about the role of each amino acid residue in antibody-antigen interaction. Taking anti-hen egg lysozyme antibodies and several anti-small molecule antibodies, the energetic contribution of hot-spot and non-hot-spot residues is discussed in terms of thermodynamics. Here, thermodynamics of the contribution from aromatic, charged and hydrogen bond-forming amino acids are discussed, and their different characteristics have been elucidated. The information gives fundamental understanding of the antibody-antigen interaction. Furthermore, the consequences of antibody engineering are analysed from thermodynamic viewpoints: humanization to reduce immunogenicity and rational design to improve affinity. Amino acid residues outside hot-spots in the interface play important roles in these cases, and thus thermodynamic and kinetic parameters give much information about the antigen recognition. Thermodynamic analysis of mutant antibodies thus should lead to advanced strategies to design and select antibodies with high affinity. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  16. Oral dendritic cells mediate antigen-specific tolerance by stimulating TH1 and regulatory CD4+ T cells.

    Science.gov (United States)

    Mascarell, Laurent; Lombardi, Vincent; Louise, Anne; Saint-Lu, Nathalie; Chabre, Henri; Moussu, Hélène; Betbeder, Didier; Balazuc, Anne-Marie; Van Overtvelt, Laurence; Moingeon, Philippe

    2008-09-01

    A detailed characterization of oral antigen-presenting cells is critical to improve second-generation sublingual allergy vaccines. To characterize oral dendritic cells (DCs) within lingual and buccal tissues from BALB/c mice with respect to their surface phenotype, distribution, and capacity to polarize CD4(+) T-cell responses. In situ analysis of oral DCs was performed by immunohistology. Purified DCs were tested in vitro for their capacity to capture, process, and present the ovalbumin antigen to naive CD4(+) T cells. In vivo priming of ovalbumin-specific T cells adoptively transferred to BALB/c mice was analyzed by cytofluorometry in cervical lymph nodes after sublingual administration of mucoadhesive ovalbumin. Three subsets of oral DCs with a distinct tissue distribution were identified: (1) a minor subset of CD207(+) Langerhans cells located in the mucosa itself, (2) a major subpopulation of CD11b(+)CD11c(-) and CD11b(+)CD11c(+) myeloid DCs at the mucosal/submucosal interface, and (3) B220(+)120G8(+) plasmacytoid DCs found in submucosal tissues. Purified myeloid and plasmacytoid oral DCs capture and process the antigen efficiently and are programmed to elicit IFN-gamma and/or IL-10 production together with a suppressive function in naive CD4(+) T cells. Targeting the ovalbumin antigen to oral DCs in vivo by using mucoadhesive particles establishes tolerance in the absence of cell depletion through the stimulation of IFN-gamma and IL-10-producing CD4(+) regulatory T cells in cervical lymph nodes. The oral immune system is composed of various subsets of tolerogenic DCs organized in a compartmentalized manner and programmed to induce T(H)1/regulatory T-cell responses.

  17. Pathological Outcome following Radical Prostatectomy in Men with Prostate Specific Antigen Greater than 10 ng/ml and Histologically Favorable Risk Prostate Cancer.

    Science.gov (United States)

    Yu, Jiwoong; Kwon, Young Suk; Kim, Sinae; Han, Christopher Sejong; Farber, Nicholas; Kim, Jongmyung; Byun, Seok Soo; Kim, Wun-Jae; Jeon, Seong Soo; Kim, Isaac Yi

    2016-05-01

    Active surveillance is now the treatment of choice in men with low risk prostate cancer. Although there is no consensus on which patients are eligible for active surveillance, prostate specific antigen above 10 ng/ml is generally excluded. In an attempt to determine the validity of using a prostate specific antigen cutoff of 10 ng/ml to counsel men considering active surveillance we analyzed a multi-institution database to determine the pathological outcome in men with prostate specific antigen greater than 10 ng/ml but histologically favorable risk prostate cancer. We queried a prospectively maintained database of men with histologically favorable risk prostate cancer who underwent radical prostatectomy between 2003 and 2015. The cohort was categorized into 3 groups based on prostate specific antigen level, including low-less than 10 ng/ml, intermediate-10 or greater to less than 20 and high-20 or greater. Associations of prostate specific antigen group with adverse pathological and oncologic outcomes were analyzed. Of 2,125 patients 1,327 were categorized with histologically favorable risk disease. However on multivariate analyses the rates of up staging and upgrading were similar between the intermediate and low prostate specific antigen groups. In contrast compared to the intermediate prostate specific antigen group the high group had higher incidences of up staging (p = 0.02) and upgrading to 4 + 3 or greater disease (p = 0.046). Biochemical recurrence-free survival rates revealed no pairwise intergroup differences except between the low and high groups. Patients with preoperatively elevated prostate specific antigen between 10 and less than 20 ng/ml who otherwise had histologically favorable risk prostate cancer were not at higher risk for adverse pathological outcomes than men with prostate specific antigen less than 10 ng/ml. Copyright © 2016 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  18. Immune responses of B. malayi thioredoxin (TRX) and venom allergen homologue (VAH) chimeric multiple antigen for lymphatic filariasis.

    Science.gov (United States)

    Anugraha, Gandhirajan; Jeyaprita, Parasurama Jawaharlal; Madhumathi, Jayaprakasam; Sheeba, Tamilvanan; Kaliraj, Perumal

    2013-12-01

    Although multiple vaccine strategy for lymphatic filariasis has provided tremendous hope, the choice of antigens used in combination has determined its success in the previous studies. Multiple antigens comprising key vaccine candidates from different life cycle stages would provide a promising strategy if the antigenic combination is chosen by careful screening. In order to analyze one such combination, we have used a chimeric construct carrying the well studied B. malayi antigens thioredoxin (BmTRX) and venom allergen homologue (BmVAH) as a fusion protein (TV) and evaluated its immune responses in mice model. The efficacy of fusion protein vaccine was explored in comparison with the single antigen vaccines and their cocktail. In mice, TV induced significantly high antibody titer of 1,28,000 compared to cocktail vaccine TRX+VAH (50,000) and single antigen vaccine TRX (16,000) or VAH (50,000). Furthermore, TV elicited higher level of cellular proliferative response together with elevated levels of IFN-γ, IL-4 and IL-5 indicating a Th1/Th2 balanced response. The isotype antibody profile showed significantly high level of IgG1 and IgG2b confirming the balanced response elicited by TV. Immunization with TV antigen induced high levels of both humoral and cellular immune responses compared to either cocktail or antigen given alone. The result suggests that TV is highly immunogenic in mice and hence the combination needs to be evaluated for its prophylactic potential.

  19. The actin cytoskeleton modulates the activation of iNKT cells by segregating CD1d nanoclusters on antigen-presenting cells

    Science.gov (United States)

    Torreno-Pina, Juan A.; Manzo, Carlo; Salio, Mariolina; Aichinger, Michael C.; Oddone, Anna; Lakadamyali, Melike; Shepherd, Dawn; Besra, Gurdyal S.; Cerundolo, Vincenzo

    2016-01-01

    Invariant natural killer T (iNKT) cells recognize endogenous and exogenous lipid antigens presented in the context of CD1d molecules. The ability of iNKT cells to recognize endogenous antigens represents a distinct immune recognition strategy, which underscores the constitutive memory phenotype of iNKT cells and their activation during inflammatory conditions. However, the mechanisms regulating such “tonic” activation of iNKT cells remain unclear. Here, we show that the spatiotemporal distribution of CD1d molecules on the surface of antigen-presenting cells (APCs) modulates activation of iNKT cells. By using superresolution microscopy, we show that CD1d molecules form nanoclusters at the cell surface of APCs, and their size and density are constrained by the actin cytoskeleton. Dual-color single-particle tracking revealed that diffusing CD1d nanoclusters are actively arrested by the actin cytoskeleton, preventing their further coalescence. Formation of larger nanoclusters occurs in the absence of interactions between CD1d cytosolic tail and the actin cytoskeleton and correlates with enhanced iNKT cell activation. Importantly and consistently with iNKT cell activation during inflammatory conditions, exposure of APCs to the Toll-like receptor 7/8 agonist R848 increases nanocluster density and iNKT cell activation. Overall, these results define a previously unidentified mechanism that modulates iNKT cell autoreactivity based on the tight control by the APC cytoskeleton of the sizes and densities of endogenous antigen-loaded CD1d nanoclusters. PMID:26798067

  20. The evolutionary dynamics of variant antigen genes in Babesia reveal a history of genomic innovation underlying host-parasite interaction

    KAUST Repository

    Jackson, Andrew P.

    2014-05-05

    Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5? ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct. 2014 The Author(s) 2014.

  1. The evolutionary dynamics of variant antigen genes in Babesia reveal a history of genomic innovation underlying host-parasite interaction

    KAUST Repository

    Jackson, Andrew P.; Otto, Thomas D.; Darby, Alistair; Ramaprasad, Abhinay; Xia, Dong; Echaide, Ignacio Eduardo; Farber, Marisa; Gahlot, Sunayna; Gamble, John; Gupta, Dinesh; Gupta, Yask; Jackson, Louise; Malandrin, Laurence; Malas, Tareq B.; Moussa, Ehab; Nair, Mridul; Reid, Adam J.; Sanders, Mandy; Sharma, Jyotsna; Tracey, Alan; Quail, Mike A.; Weir, William; Wastling, Jonathan M.; Hall, Neil; Willadsen, Peter; Lingelbach, Klaus; Shiels, Brian; Tait, Andy; Berriman, Matt; Allred, David R.; Pain, Arnab

    2014-01-01

    Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5? ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct. 2014 The Author(s) 2014.

  2. The evolutionary dynamics of variant antigen genes in Babesia reveal a history of genomic innovation underlying host–parasite interaction

    Science.gov (United States)

    Jackson, Andrew P.; Otto, Thomas D.; Darby, Alistair; Ramaprasad, Abhinay; Xia, Dong; Echaide, Ignacio Eduardo; Farber, Marisa; Gahlot, Sunayna; Gamble, John; Gupta, Dinesh; Gupta, Yask; Jackson, Louise; Malandrin, Laurence; Malas, Tareq B.; Moussa, Ehab; Nair, Mridul; Reid, Adam J.; Sanders, Mandy; Sharma, Jyotsna; Tracey, Alan; Quail, Mike A.; Weir, William; Wastling, Jonathan M.; Hall, Neil; Willadsen, Peter; Lingelbach, Klaus; Shiels, Brian; Tait, Andy; Berriman, Matt; Allred, David R.; Pain, Arnab

    2014-01-01

    Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5′ ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct. PMID:24799432

  3. Enhancement of the priming efficacy of DNA vaccines encoding dendritic cell-targeted antigens by synergistic toll-like receptor ligands

    Directory of Open Access Journals (Sweden)

    Kornbluth Richard S

    2009-08-01

    Full Text Available Abstract Background Targeting of protein antigens to dendritic cells (DC via the DEC205 receptor enhances presentation of antigen-derived peptides on MHC-I and MHC-II molecules and, in the presence of costimulatory signals, antigen-specific immune responses. The immunogenicity and efficacy of DNA vaccination can also be enhanced by fusing the encoded antigen to single chain antibodies directed against DEC205. To further improve this strategy, we evaluated different toll-like receptor ligands (TLR and CD40 ligands (CD40L as adjuvants for DNA vaccines encoding a DEC205-single-chain antibody fused to the ovalbumin model antigen or HIV-1 Gag and assessed the priming efficacy of DNA in a DNA prime adenoviral vector boost immunization regimen. Results Mice were primed with the adjuvanted DEC-205 targeted DNA vaccines and boosted with adenoviral vectors encoding the same antigens. CD8+ T cell responses were determined after the adenoviral booster immunization, to determine how well the different DNA immunization regimens prime for the adenoviral boost. In the absence of adjuvants, targeting of DNA-encoded ovalbumin to DCs suppressed CD8+ T-cell responses after the adenoviral booster immunization. CD8+ T-cell responses to the DEC205 targeted DNA vaccines increased only slightly by adding either the TLR-9 ligand CpG, the TLR-3 ligand Poly I:C, or CD40 ligand expression plasmids. However, the combination of both TLR-ligands led to a strong enhancement of CD8+ T-cell responses compared to a non-targeted DNA vaccine. This finding was confirmed using HIV Gag as antigen. Conclusion Although DNA prime adenoviral vector boost immunizations belong to the strongest inducers of cytotoxic T cell responses in different animal models and humans, the CD8+ T cell responses can be further improved by targeting the DNA encoded antigen to DEC205 in the presence of synergistic TLR ligands CpG and Poly I:C.

  4. Distinct pattern of lesion distribution in multiple sclerosis is associated with different circulating T-helper and helper-like innate lymphoid cell subsets.

    Science.gov (United States)

    Gross, Catharina C; Schulte-Mecklenbeck, Andreas; Hanning, Uta; Posevitz-Fejfár, Anita; Korsukewitz, Catharina; Schwab, Nicholas; Meuth, Sven G; Wiendl, Heinz; Klotz, Luisa

    2017-06-01

    Distinct lesion topography in relapsing-remitting multiple sclerosis (RRMS) might be due to different antigen presentation and/or trafficking routes of immune cells into the central nervous system (CNS). To investigate whether distinct lesion patterns in multiple sclerosis (MS) might be associated with a predominance of distinct circulating T-helper cell subset as well as their innate counterparts. Flow cytometric analysis of lymphocytes derived from the peripheral blood of patients with exclusively cerebral (n = 20) or predominantly spinal (n = 12) disease manifestation. Patients with exclusively cerebral or preferential spinal lesion manifestation were associated with increased proportions of circulating granulocyte-macrophage colony-stimulating factor (GM-CSF) producing T H 1 cells or interleukin (IL)-17-producing T H 17 cells, respectively. In contrast, proportions of peripheral IL-17/IL-22-producing lymphoid tissue inducer (LTi), the innate counterpart of T H 17 cells, were enhanced in RRMS patients with exclusively cerebral lesion topography. Distinct T-helper and T-helper-like innate lymphoid cell (ILC) subsets are associated with different lesion topography in RRMS.

  5. Antigenic variation and the genetics and epigenetics of the PfEMP1 erythrocyte surface antigens in Plasmodium falciparum malaria

    DEFF Research Database (Denmark)

    Arnot, David E; Jensen, Anja T R

    2011-01-01

    . Sterile immunity is not achieved and chronic parasitization of apparently healthy adults is the norm. In this article, we analyse the best understood malaria "antigenic variation" system, that based on Plasmodium falciparum's PfEMP1-type cytoadhesion antigens, and critically review recent literature...

  6. Cytotoxic T lymphocyte recognition of HLA-A/B antigens introduced into EL4 cells by cell-liposome fusion.

    Science.gov (United States)

    Engelhard, V H; Powers, G A; Moore, L C; Holterman, M J; Correa-Freire, M C

    1984-01-01

    HLA-A2 and -B7 antigens were introduced into EL4 (H-2b) cells by cell-liposome fusion and were used as targets or stimulators for cytotoxic T lymphocytes (CTL) generated in C57B1/6 (H-2b) mice. It was found that such EL4-HLA cells were not recognized by CTL that had been raised against either a human cell line bearing these HLA antigens or the purified HLA-A2 and -B7 antigens reconstituted into liposomes. In addition, EL4-HLA cells were not capable of inducing CTL that could recognize a human cell line bearing HLA-A2 and -B7 antigens. Instead, EL4-HLA cells induced CTL that specifically lysed EL4-HLA cells and not human cells expressing HLA-A2 and -B7. CTL recognition required the presence of HLA antigens on the EL4 cell surface and was inhibited by antibodies against either H-2b or HLA-A/B. Monoclonal antibody binding studies showed that the expected polymorphic determinants of the HLA-A2 and -B7 antigens were still present on EL4-HLA cells. However, the specificity of CTL or their precursors that are capable of recognizing HLA-A2 or -B7 was altered after these antigens became associated with the EL4 surface. Possible explanations for these results are discussed.

  7. Topographic study of the ADP/ATP transport protein. Localization of ADP and atractyloside fixation sites. Identification of the antigenic domains

    International Nuclear Information System (INIS)

    Boulay, Francois

    1983-01-01

    The objectives of this research thesis were: to determine the intramolecular localisation of binding sites of atractyloside and adenine-nucleotides; to determine whether antibodies obtained against the ADP/ATP carrier protein and isolated from beef heart mitochondria possess a reactivity specific to the organ or the species, where antigenic determinants are localized and whether there is conservation of the antigenic structure from one species to the other; to study how to follow and interpret conformational changes of the protein under the effect of ADP and inhibitors (carboxy-atractyloside or bongkrekic acid), and where the SH group unmasked by ADP and bongkrekic acid is localized [fr

  8. Re-purification of labelled ferritin antigen with HPLC

    International Nuclear Information System (INIS)

    Zhang Haoyi; Jin Lichun

    2002-01-01

    Objective: To improve the quality of long-term stored labelled ferritin antigen with HPLC. Methods: The antigen was analyzed and purified with HPLC and again analyzed with RIA afterwards. Results: Ferritin antigen underwent significant polymerization after long-term (aggregation) storage. After re-purification with HPLC, its immuno-activity and labelled specific radioactivity were both significantly improved. Conclusion: Quality of stored ferritin RIA kit could be greatly improved after re-purification with HPLC

  9. Human Tumor Antigens Yesterday, Today, and Tomorrow.

    Science.gov (United States)

    Finn, Olivera J

    2017-05-01

    The question of whether human tumors express antigens that can be recognized by the immune system has been answered with a resounding YES. Most were identified through spontaneous antitumor humoral and cellular immune responses found in cancer patients and include peptides, glycopeptides, phosphopeptides, viral peptides, and peptides resulting from common mutations in oncogenes and tumor-suppressor genes, or common gene fusion events. Many have been extensively tested as candidates for anticancer vaccines. More recently, attention has been focused on the potentially large number of unique tumor antigens, mutated neoantigens, that are the predicted products of the numerous mutations revealed by exome sequencing of primary tumors. Only a few have been confirmed as targets of spontaneous immunity and immunosurveillance, and even fewer have been tested in preclinical and clinical settings. The field has been divided for a long time on the relative importance of shared versus mutated antigens in tumor surveillance and as candidates for vaccines. This question will eventually need to be answered in a head to head comparison in well-designed clinical trials. One advantage that shared antigens have over mutated antigens is their potential to be used in vaccines for primary cancer prevention. Cancer Immunol Res; 5(5); 347-54. ©2017 AACR . ©2017 American Association for Cancer Research.

  10. Identification of protective antigens for vaccination against systemic salmonellosis

    Directory of Open Access Journals (Sweden)

    Dirk eBumann

    2014-08-01

    Full Text Available There is an urgent medical need for improved vaccines with broad serovar coverage and high efficacy against systemic salmonellosis. Subunit vaccines offer excellent safety profiles but require identification of protective antigens, which remains a challenging task. Here, I review crucial properties of Salmonella antigens that might help to narrow down the number of potential candidates from more than 4000 proteins encoded in Salmonella genomes, to a more manageable number of 50-200 most promising antigens. I also discuss complementary approaches for antigen identification and potential limitations of current pre-clinical vaccine testing.

  11. Biological and clinical meaning of myeloid antigen expression in the acute lymphocytic leukemia in children

    International Nuclear Information System (INIS)

    Marsan Suarez, Vianed; Sanchez Segura, Miriam; Socarras Ferrer, Bertha B; Valle Perez, Lazaro O del

    2009-01-01

    In 238 children presenting with acute lymphoid leukemia (ALL) authors studied the possible association between the myeloid antigens expression with determined biologic and clinic features at disease onset. The cellular immunophenotyping was performed by ultraimmunocytochemical method. From the total of diagnosed ALLs, the 21,8% were LLA-Mi+. There was a lymphadenopathies predominance (71,2%), splenomegaly (65,4%) and hepatomegaly (57,7%) in patients with LLA-Mi+ and very significant differences (p =0,003, p = 0,0068, and p = 0,000, respectively. There was also alight predominance of mediastinum adenopathies, CNS infiltration and hemorrahagic manifestations in patients with LLA-Mi+, no statistically significant. Results showed that in our patients the myeloid antigen expression on the lymphoid blasts influenced on appearance of determined presentation of morphologic and clinical features in children

  12. Characterisation of antibody responses in pigs induced by recombinant oncosphere antigens from Taenia solium.

    Science.gov (United States)

    Jayashi, César M; Gonzalez, Armando E; Castillo Neyra, Ricardo; Kyngdon, Craig T; Gauci, Charles G; Lightowlers, Marshall W

    2012-12-14

    Recombinant antigens cloned from the oncosphere life cycle stage of the cestode parasite Taenia solium (T. solium) have been proven to be effective as vaccines for protecting pigs against infections with T. solium. Previous studies have defined three different host protective oncosphere antigens, TSOL18, TSOL16 and TSOL45. In this study, we evaluated the potential for combining the antigens TSOL16 and TSOL18 as a practical vaccine. Firstly, in a laboratory trial, we compared the immunogenicity of the combined antigens (TSOL16/18) versus the immunogenicity of the antigens separately. Secondly, in a field trial, we tested the ability of the TSOL16/18 vaccine to induce detectable antibody responses in animals living under environmental stress and traditionally reared in areas where T. solium cysticercosis is endemic; and finally, we characterised the immune response of the study population. Pigs of 8-16 weeks of age were vaccinated with 200 μg each of TSOL16 and TSOL18, plus 5mg of Quil-A. Specific total IgG, IgG(1) and IgG(2) antibody responses induced by TSOL16 and TSOL18 were determined with ELISA. The immunogenicity of both antigens was retained in the combined TSOL16/18 vaccine. The combined vaccine TSOL16/18 induced detectable specific anti-TSOL18 antibody responses in 100% (113/113) and specific anti-TSOL16 in 99% (112/113) of the vaccinated animals measured at 2 weeks following the booster vaccination. From the two IgG antibody subtypes analysed we found there was stronger response to IgG(2). Copyright © 2012 Elsevier Ltd. All rights reserved.

  13. Dissection of T-cell antigen specificity in human melanoma

    DEFF Research Database (Denmark)

    Andersen, Rikke Sick; Albæk Thrue, Charlotte; Junker, Niels

    2012-01-01

    Tumor-infiltrating lymphocytes (TIL) isolated from melanoma patients and expanded in vitro by interleukin (IL)-2 treatment can elicit therapeutic response after adoptive transfer, but the antigen specificities of the T cells transferred have not been determined. By compiling all known melanoma-as...... from different fragments of resected melanoma lesions. In summary, our findings provide an initial definition of T-cell populations contributing to tumor recognition in TILs although the specificity of many tumor-reactive TILs remains undefined....

  14. Impact of obesity on the predictive accuracy of prostate-specific antigen density and prostate-specific antigen in native Korean men undergoing prostate biopsy.

    Science.gov (United States)

    Kim, Jae Heon; Doo, Seung Whan; Yang, Won Jae; Lee, Kwang Woo; Lee, Chang Ho; Song, Yun Seob; Jeon, Yoon Su; Kim, Min Eui; Kwon, Soon-Sun

    2014-10-01

    To evaluate the impact of obesity on the biopsy detection of prostate cancer. We retrospectively reviewed data of 1182 consecutive Korean patients (≥50 years) with serum prostate-specific antigen levels of 3-10 ng/mL who underwent initial extended 12-cores biopsy from September 2009 to March 2013. Patients who took medications that were likely to influence the prostate-specific antigen level were excluded. Receiver operating characteristic curves were plotted for prostate-specific antigen and prostate-specific antigen density predicting cancer status among non-obese and obese men. A total of 1062 patients (mean age 67.1 years) were enrolled in the analysis. A total of 230 men (21.7%) had a positive biopsy. In the overall study sample, the area under the receiver operator characteristic curve of serum prostate-specific antigen for predicting prostate cancer on biopsy were 0.584 and 0.633 for non-obese and obese men, respectively (P = 0.234). However, the area under the curve for prostate-specific antigen density in predicting cancer status showed a significant difference (non-obese 0.696, obese 0.784; P = 0.017). There seems to be a significant difference in the ability of prostate-specific antigen density to predict biopsy results between non-obese and obese men. Obesity positively influenced the overall ability of prostate-specific antigen density to predict prostate cancer. © 2014 The Japanese Urological Association.

  15. Screening for simian foamy virus infection by using a combined antigen Western blot assay: evidence for a wide distribution among Old World primates and identification of four new divergent viruses

    International Nuclear Information System (INIS)

    Hussain, Althaf I.; Shanmugam, Vedapuri; Bhullar, Vinod B.; Beer, Brigitte E.; Vallet, Dominique; Gautier-Hion, Annie; Wolfe, Nathan D.; Karesh, William B.; Kilbourn, Annelisa M.; Tooze, Zeena; Heneine, Walid; Switzer, William M.

    2003-01-01

    Simian foamy viruses (SFVs) belong to a genetically and antigenically diverse class of retroviruses that naturally infect a wide range of nonhuman primates (NHPs) and can also be transmitted to humans occupationally exposed to NHPs. Current serologic detection of SFV infection requires separate Western blot (WB) testing by using two different SFV antigens [SFV AGM (African green monkey) and SFV CPZ (chimpanzee)]. However, this method is labor intensive and validation is limited to only small numbers of NHPs. To facilitate serologic SFV testing, we developed a WB assay that combines antigens from both SFV AGM and SFV CPZ . The combined-antigen WB (CA-WB) assay was validated with 145 serum samples from 129 NHPs (32 African and Asian species) and 16 humans, all with known SFV infection status determined by PCR. Concordant CA-WB results were obtained for all 145 PCR-positive or -negative primate and human specimens, giving the assay a 100% sensitivity and specificity. In addition, no reactivity was observed in sera from persons positive for human immunodeficiency virus or human T cell lymphotropic virus (HIV/HTLV) (n = 25) or HIV/HTLV-negative U.S. blood donors (n = 100). Using the CA-WB assay, we screened 360 sera from 43 Old World primate species and found an SFV prevalence of about 68% in both African and Asian primates. We also isolated SFV from the blood of four seropositive primates (Allenopithecus nigroviridis, Trachypithecus francoisi, Hylobates pileatus, and H. leucogenys) not previously known to be infected with SFV. Phylogenetic analysis of integrase sequences from these isolates confirmed that all four SFVs represent new, distinct, and highly divergent lineages. These results demonstrate the ability of the CA-WB assay to detect infection in a large number of NHP species, including previously uncharacterized infections with divergent SFVs

  16. Use of Ionizing Radiations to Prepare Radiovaccines and Radio-Antigens

    International Nuclear Information System (INIS)

    Tumanyan, MA; Hruschev, V.G.

    1967-01-01

    The possibility of employing ionizing radiations at certain doses to kill micro-organisms was used to produce vaccines against intestinal infections, and also to obtain from these bacteria antigens capable of being used as chemical vaccines. Typhoid fever and dysentery radiovaccines and radio-antigens were prepared, and the effect of various gamma ray doses on their toxicity and their antigenic and immunogenic properties was tested. The doses used did not change properties of these products as compared with those of vaccines and antigens produced by normal means. The paper also discusses the possibility of using radiation to sterilize fabricated vaccines and antigens, including radiovaccines and radio-antigens, anitoxins, antitoxic serums and nutrient media for the culture of micro-organisms. Data on the irradiation apparatus used for these investigations are reported. (author) [ru

  17. Microradioimmunoassay for antibodies to tumor-associated antigens

    International Nuclear Information System (INIS)

    Huang, J.C.C.; Berczi, I.; Froese, G.; Tsay, H.M.; Sehon, A.H.

    1975-01-01

    A versatile microradioimmunoassay for the detection of antibodies to tumor-associated and other tissue antigens was described. The method involved: the preparation of solid-phase antigen with cultured (already adhered) or noncultured cells (sedimented by centrifugation) fixed to Micro-Test plates with neutral buffered formaldehyde or absolute methanol; the incubation of the antigen with test or control sera; and the incubation of the antigen with radioiodinated antiglobulin antibody. The nonspecific background of radioactivity was reduced to an acceptable level by the fixed cells being precoated in the wells with 0.5 percent bovine serum albumin in phosphate-buffered saline which was also used for the dilution of sera and labeled antiglobulin antibody. Tumor cells in primary cultures gave a high background, as compared to long-term cultures, which was due to the presence of immunoglobulins (most likely tumor-specific antibody). The specific antibody response to a syngeneic mouse tumor was demonstrated by this technique. (auth)

  18. Prostate-Specific Antigen (PSA) Test: MedlinePlus Lab Test Information

    Science.gov (United States)

    ... medlineplus.gov/labtests/prostatespecificantigenpsatest.html Prostate-Specific Antigen (PSA) Test To use the sharing features on this ... enable JavaScript. What is a prostate-specific antigen (PSA) test? A prostate-specific antigen (PSA) test measures ...

  19. Isolation of the new antigen receptor from wobbegong sharks, and use as a scaffold for the display of protein loop libraries.

    Science.gov (United States)

    Nuttall, S D; Krishnan, U V; Hattarki, M; De Gori, R; Irving, R A; Hudson, P J

    2001-08-01

    The new antigen receptor (NAR) from nurse sharks consists of an immunoglobulin variable domain attached to five constant domains, and is hypothesised to function as an antigen-binding antibody-like molecule. To determine whether the NAR is present in other species we have isolated a number of new antigen receptor variable domains from the spotted wobbegong shark (Orectolobus maculatus) and compared their structure to that of the nurse shark protein. To determine whether these wNARs can function as antigen-binding proteins, we have used them as scaffolds for the construction of protein libraries in which the CDR3 loop was randomised, and displayed the resulting recombinant domains on the surface of fd bacteriophages. On selection against several protein antigens, the highest affinity wNAR proteins were generated against the Gingipain K protease from Porphyromonas gingivalis. One wNAR protein bound Gingipain K specifically by ELISA and BIAcore analysis and, when expressed in E. coli and purified by affinity chromatography, eluted from an FPLC column as a single peak consistent with folding into a monomeric protein. Naturally occurring nurse shark and wobbegong NAR variable domains exhibit conserved cysteine residues within the CDR1 and CDR3 loops which potentially form disulphide linkages and enhance protein stability; proteins isolated from the in vitro NAR wobbegong library showed similar selection for such paired cysteine residues. Thus, the New Antigen Receptor represents a protein scaffold with possible stability advantages over conventional antibodies when used in in vitro molecular libraries.

  20. Comparison between an immunochromatographic test with an amplified ELISA for detecting e antigen and anti-e antigen antibodies in chronic Hepatitis B

    International Nuclear Information System (INIS)

    Mainet Gonzalez, Damian; Palenzuela Gardon, Daniel O; Aguilar Rubido; Julio C

    2009-01-01

    The disappearance of the e antigen and the appearance of anti-e antigen antibodies are two biomarkers that indicate favorable prognosis in Hepatitis B. In this study the Advanced QualityTM immunochromatographic test for detecting those biomarkers was compared to the Vidas semi-quantitative ELISA test. Our hypothesis was that it is possible to use these biomarkers measured in a rapid and simple Advanced QualityTM immunochromatographic test for evaluating the therapeutic response in clinical trials with chronic hepatitis B patients. The two methods were done following the manufacturer's instructions. The sera were taken from 69 patients with chronic hepatitis B of the clinical trial of the CIGB 440 therapeutic candidate. The immunochromatographic test and ELISA for detecting e antigen and anti-e antigen antibodies presented from substantial to almost perfect agreement in the evaluation of the sera of chronic Hepatitis B patients in a clinical trial. The immunochromatographic test for detecting e antigen had a low positive average agreement and a high negative average agreement compared to the ELISA. Nevertheless, the immunochromatographic test for detecting anti-e antigen antibodies had a high negative and positive average agreement in comparison to the ELISA. The immunochromagraphic test for the e antigen had a lower positive average agreement compared to the ELISA and some patients infected with Hepatitis B virus could not be detected by the former assay. The immunochromatographic test for anti-e antigen antibodies showed a similar performance to that of ELISA and could therefore be used in clinical trials for chronic Hepatitis B in health institutions without the need of a highly qualified lab technician. (author)

  1. A radioimmunoassay for human antibody specific for microbial antigens

    International Nuclear Information System (INIS)

    Tew, J.G.; Burmeister, J.; Greene, E.J.; Pflaumer, S.K.; Goldstein, J.

    1977-01-01

    A simple and sensitive method for detecting and quantitating antibody specific or microbial antigens is described. Bacterial, fungal, parasitic or viral antigens attached to bromoacetyl cellulose or the intact cells themselves were added to a series of two-fold dilutions of human serum. After a short incubation period, which allowed human antibody to attach to the antigens, the complex was thoroughly washed and carbon-14 labeled anti-human light chain antibody was added to each dilution. The resulting complex was washed, collected on a filter pad, placed in a scintillation vial and radioassayed. The relationship between radioactivity bound and -log 2 of the serum dilution was linear. The endpoint for each assay and a confidence interval was calculated by doing inverse prediction from simple linear regression. Results obtained using this assay indicated the presence of antibody in a pool of normal human sera specific for herpes virus and for both cell surface and intracellular antigens of Streptococcus mutans, Naegleria fowleri and Cryptococcus neoformans. In general the dominant response was against the intracellular antigens rather than cell surface antigens

  2. Cerium oxide-deposited mesoporous silica nanoparticles for the determination of carcinoembryonic antigen in serum using inductively coupled plasma-mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Choi, H.W. [Department of Chemistry, NSBI, Dankook University, 126 Jukjeon-dong, Suji-gu, Yongin-si, Gyeonggi-do 448-701 (Korea, Republic of); Lee, K.H.; Hur, N.H. [Department of Chemistry, Sogang University, Shinsu-dong, Mapo-gu, Seoul (Korea, Republic of); Lim, H.B., E-mail: plasma@dankook.ac.kr [Department of Chemistry, NSBI, Dankook University, 126 Jukjeon-dong, Suji-gu, Yongin-si, Gyeonggi-do 448-701 (Korea, Republic of)

    2014-10-17

    Highlights: • Sandwich-type immunoassay using ICP-MS and nanoparticles to determine biomarkers. • CeO{sub 2}-deposited mesoporous silica nanoparticles were synthesized as a probe. • Ratiometric measurement significantly improved the calibration linearity. • Excellent detection limit was achieved by signal amplification. - Abstract: CeO{sub 2}-deposited mesoporous silica nanoparticles were synthesized as a probe to determine carcinoembryonic antigen (CEA) in serum by inductively coupled plasma-mass spectrometry (ICP-MS). The prepared mesoporous nanoparticles were modified and tagged to the target for sandwich-type immunoassay. Fe{sub 3}O{sub 4} magnetic nanoparticles (MNPs) were also synthesized and immobilized with antibody to extract the target biomarker. The calibration curve of the synthesized CeO{sub 2}-deposited silica nanoparticles, which was plotted by the signal ratio of {sup 140}Ce/{sup 57}Fe measured by ICP-MS vs. the concentration of CEA, showed excellent linearity and sensitivity owing to the signal amplification and low spectral interference. Under optimal conditions, the sandwich-type analytical method was applied to determine CEA in serum spiked in the range of 0.001–5 ng mL{sup −1} and showed a limit of detection of 0.36 ng mL{sup −1}. Since the deposited CeO{sub 2} in the mesoporous silica layer can be substituted by other metal compounds, various kinds of metal-deposited nanoparticles can be prepared as probe materials for multiplex detection in bioanalysis.

  3. Distinct Gut-Derived Bacteria Differentially Affect Three Types of Antigen-Presenting Cells and Impact on NK- and T-Cell Responses

    DEFF Research Database (Denmark)

    Fink, Lisbeth Nielsen; Hansen, Anne Marie Valentin; Frøkiær, Hanne

    Objectives Gut bacteria are assumed essential for development and maintenance of a balanced immune system. Specifically, stimulation of antigen-presenting cells (APCs) by gut bacteria is important for polarisation of the immune response. This experiment was designed to reveal similarities...... and differences between the reaction patterns of three types of human APCs when stimulated with intestinal bacteria. Furthermore, the effect of these APCs on NK-cells and T-cells was examined. Methodology The APCs used in this study were blood monocytes, blood dendritic cells, and dendritic cells differentiated...... from monocytes. Monocyte-derived dendritic cells constitute a commonly used model of dendritic cell function. The APCs were cultured for 18 h with four different gut bacteria: Lactobacillus acidophilus X37, Lactobacillus reuteri DSM 12246, E. coli Nissle 1917 or Bifidobacterium longum Q46. Results...

  4. SV40 large T-p53 complex: evidence for the presence of two immunologically distinct forms of p53

    International Nuclear Information System (INIS)

    Milner, J.; Gamble, J.

    1985-01-01

    The transforming protein of SV40 is the large T antigen. Large T binds a cellular protein, p53, which is potentially oncogenic by virtue of its functional involvement in the control of cell proliferation. This raises the possibility that p53 may mediate, in part, the transforming function of SV40 large T. Two immunologically distinct forms of p53 have been identified in normal cells: the forms are cell-cycle dependent, one being restricted to nondividing cells (p53-Go) and the second to dividing cells (p53-G divided by). The authors have now dissociated and probed the multimeric complex of SV40 large T-p53 for the presence of immunologically distinct forms of p53. Here they present evidence for the presence of p53-Go and p53-G divided by complexed with SV40 large T

  5. Genetic and antigenic characterization of novel pestivirus genotypes: implications for classification

    International Nuclear Information System (INIS)

    Becher, Paul; Avalos Ramirez, Ramiro; Orlich, Michaela; Cedillo Rosales, Sibilina; Koenig, Matthias; Schweizer, Matthias; Stalder, Hanspeter; Schirrmeier, Horst; Thiel, Heinz-Juergen

    2003-01-01

    Currently, the genus Pestivirus comprises the four approved species Bovine viral diarrhea virus 1 (BVDV-1), BVDV-2, Classical swine fever virus (CSFV), and Border disease virus (BDV) and one tentative fifth species represented by a single strain (H138) isolated from a giraffe in Kenya more than 30 years ago. To further address the issue of heterogeneity of pestiviruses we have determined the entire N pro and E2 coding sequences for several new pestivirus isolates. Interestingly, phylogenetic analysis revealed that one pestivirus isolated in the 1990s in Africa is closely related to strain H138. Moreover, several novel pestiviruses isolated from sheep group together with the previously described strain V60 (Reindeer-1) isolated from a reindeer, whereas one ovine pestivirus strain (Gifhorn) significantly differs from all previously described pestiviruses, including BDV. We propose to term these mainly sheep-derived pestiviruses BDV-2 (V60-like isolates) and BDV-3 (Gifhorn); consequently, the 'classical' BDV isolates should be termed BDV-1. As an additional criterion for segregation of pestiviruses, the antigenic relatedness of pestivirus isolates covering all observed major genotypes was studied by cross-neutralization assays. Analysis of the antigenic similarities indicated the presence of seven major antigenic groups corresponding to BVDV-1, BVDV-2, CSFV, BDV-1, BDV-2, BDV-3, and 'giraffe'. Taking into account the host origin, the lack of differences concerning the course of disease, and the results of our genetic and antigenic analyses, we suggest that BDV-1, BDV-2, and BDV-3 should be considered as major genotypes within the species BDV

  6. Flow Cytometric Analysis of T, B, and NK Cells Antigens in Patients with Mycosis Fungoides

    Directory of Open Access Journals (Sweden)

    Serkan Yazıcı

    2015-01-01

    Full Text Available We retrospectively analyzed the clinicopathological correlation and prognostic value of cell surface antigens expressed by peripheral blood mononuclear cells in patients with mycosis fungoides (MF. 121 consecutive MF patients were included in this study. All patients had peripheral blood flow cytometry as part of their first visit. TNMB and histopathological staging of the cases were retrospectively performed in accordance with International Society for Cutaneous Lymphomas/European Organization of Research and Treatment of Cancer (ISCL/EORTC criteria at the time of flow cytometry sampling. To determine prognostic value of cell surface antigens, cases were divided into two groups as stable and progressive disease. 17 flow cytometric analyses of 17 parapsoriasis (PP and 11 analyses of 11 benign erythrodermic patients were included as control groups. Fluorescent labeled monoclonal antibodies were used to detect cell surface antigens: T cells (CD3+, CD4+, CD8+, TCRαβ+, TCRγδ+, CD7+, CD4+CD7+, CD4+CD7−, and CD71+, B cells (HLA-DR+, CD19+, and HLA-DR+CD19+, NKT cells (CD3+CD16+CD56+, and NK cells (CD3−CD16+CD56+. The mean value of all cell surface antigens was not statistically significant between parapsoriasis and MF groups. Along with an increase in cases of MF stage statistically significant difference was found between the mean values of cell surface antigens. Flow cytometric analysis of peripheral blood cell surface antigens in patients with mycosis fungoides may contribute to predicting disease stage and progression.

  7. Generation of monoclonal antibodies against highly conserved antigens.

    Directory of Open Access Journals (Sweden)

    Hongzhe Zhou

    Full Text Available BACKGROUND: Therapeutic antibody development is one of the fastest growing areas of the pharmaceutical industry. Generating high-quality monoclonal antibodies against a given therapeutic target is very crucial for the success of the drug development. However, due to immune tolerance, some proteins that are highly conserved between mice and humans are not very immunogenic in mice, making it difficult to generate antibodies using a conventional approach. METHODOLOGY/PRINCIPAL FINDINGS: In this report, the impaired immune tolerance of NZB/W mice was exploited to generate monoclonal antibodies against highly conserved or self-antigens. Using two highly conserved human antigens (MIF and HMGB1 and one mouse self-antigen (TNF-alpha as examples, we demonstrate here that multiple clones of high affinity, highly specific antibodies with desired biological activities can be generated, using the NZB/W mouse as the immunization host and a T cell-specific tag fused to a recombinant antigen to stimulate the immune system. CONCLUSIONS/SIGNIFICANCE: We developed an efficient and universal method for generating surrogate or therapeutic antibodies against "difficult antigens" to facilitate the development of therapeutic antibodies.

  8. Genome analysis of Excretory/Secretory proteins in Taenia solium reveals their Abundance of Antigenic Regions (AAR).

    Science.gov (United States)

    Gomez, Sandra; Adalid-Peralta, Laura; Palafox-Fonseca, Hector; Cantu-Robles, Vito Adrian; Soberón, Xavier; Sciutto, Edda; Fragoso, Gladis; Bobes, Raúl J; Laclette, Juan P; Yauner, Luis del Pozo; Ochoa-Leyva, Adrián

    2015-05-19

    Excretory/Secretory (ES) proteins play an important role in the host-parasite interactions. Experimental identification of ES proteins is time-consuming and expensive. Alternative bioinformatics approaches are cost-effective and can be used to prioritize the experimental analysis of therapeutic targets for parasitic diseases. Here we predicted and functionally annotated the ES proteins in T. solium genome using an integration of bioinformatics tools. Additionally, we developed a novel measurement to evaluate the potential antigenicity of T. solium secretome using sequence length and number of antigenic regions of ES proteins. This measurement was formalized as the Abundance of Antigenic Regions (AAR) value. AAR value for secretome showed a similar value to that obtained for a set of experimentally determined antigenic proteins and was different to the calculated value for the non-ES proteins of T. solium genome. Furthermore, we calculated the AAR values for known helminth secretomes and they were similar to that obtained for T. solium. The results reveal the utility of AAR value as a novel genomic measurement to evaluate the potential antigenicity of secretomes. This comprehensive analysis of T. solium secretome provides functional information for future experimental studies, including the identification of novel ES proteins of therapeutic, diagnosis and immunological interest.

  9. Kinetics of HBsub(s) antigen in man

    International Nuclear Information System (INIS)

    Drouet, J.; Courouce-Pauty, A.M.; Thevenoux, A.M.; Soulier, J.P.; Chanard, J.; Vallee, G.; Funck-Brentano, J.L.

    1975-01-01

    The metabolism of HBsub(s) antigen had been studied in three human volunteers. One had chronic hepatitis and two were silent carriers. The HBsub(s) antigen had been isolated and purified from the plasma of each of the three subjects and, after iodination, reinjected to the same donor. The parameters of plasma kinetics of 131 I HBsub(s)Ag have been analyzed according to a two compartmental model on the basis of the radioactivity of TCA precipitate (TP) and immunoprecipitate (IP). The fast initial volume of distribution was approximately equal in the three subjects (46.6ml/kg). The metabolic clearance rate (MCR) of IP was the very same in two subjects but is four times higher in one of the silent carrier. The total renewal time (TRT) was about 3.3 days. Assuming that the HBsub(s) antigen extraction was of the order of 65% the plasma HBsub(s) antigen concentration per liter of plasma would be 12 and 53mg/liter for two silent carriers and 61 mg/liter for the patient with chronic hepatitis. The radioactive efflux from the model (calculated as IP.MCR multiplied by HBsub(s) antigen concentration) was identical for the two silent carriers and 50% higher in the patient with chronic hepatitis. The increase possibly reflects an increased synthesis of HBsub(s) antigen in the patient with chronic hepatitis. The cumulative urinary radioactivity when added to the whole body counting demonstrated that radioactivity was excreted solely in the urine. The ratio of organ counting to precordium counting did not vary significantly with time in all subjects [fr

  10. Precision cancer immunotherapy: optimizing dendritic cell-based strategies to induce tumor antigen-specific T-cell responses against individual patient tumors.

    Science.gov (United States)

    Osada, Takuya; Nagaoka, Koji; Takahara, Masashi; Yang, Xiao Yi; Liu, Cong-Xiao; Guo, Hongtao; Roy Choudhury, Kingshuk; Hobeika, Amy; Hartman, Zachary; Morse, Michael A; Lyerly, H Kim

    2015-05-01

    Most dendritic cell (DC)-based vaccines have loaded the DC with defined antigens, but loading with autologos tumor-derived antigens would generate DCs that activate personalized tumor-specific T-cell responses. We hypothesized that DC matured with an optimized combination of reagents and loaded with tumor-derived antigens using a clinically feasible electroporation strategy would induce potent antitumor immunity. We first studied the effects on DC maturation and antigen presentation of the addition of picibanil (OK432) to a combination of zoledronic acid, tumor necrosis factor-α, and prostaglandin E2. Using DC matured with the optimized combination, we tested 2 clinically feasible sources of autologous antigen for electroloading, total tumor mRNA or total tumor lysate, to determine which stimulated more potent antigen-specific T cells in vitro and activated more potent antitumor immunity in vivo. The combination of tumor necrosis factor-α/prostaglandin E2/zoledronic acid/OK432 generated DC with high expression of maturation markers and antigen-specific T-cell stimulatory function in vitro. Mature DC electroloaded with tumor-derived mRNA [mRNA electroporated dendritic cell (EPDC)] induced greater expansion of antigen-specific T cells in vitro than DC electroloaded with tumor lysate (lysate EPDC). In a therapeutic model of MC38-carcinoembryonic antigen colon cancer-bearing mice, vaccination with mRNA EPDC induced the most efficient anti-carcinoembryonic antigen cellular immune response, which significantly suppressed tumor growth. In conclusion, mature DC electroloaded with tumor-derived mRNA are a potent cancer vaccine, especially useful when specific tumor antigens for vaccination have not been identified, allowing autologous tumor, and if unavailable, allogeneic cell lines to be used as an unbiased source of antigen. Our data support clinical testing of this strategy.

  11. Rapid and specific biotin labelling of the erythrocyte surface antigens of both cultured and ex-vivo Plasmodium parasites

    Directory of Open Access Journals (Sweden)

    Thompson Joanne

    2007-05-01

    Full Text Available Abstract Background Sensitive detection of parasite surface antigens expressed on erythrocyte membranes is necessary to further analyse the molecular pathology of malaria. This study describes a modified biotin labelling/osmotic lysis method which rapidly produces membrane extracts enriched for labelled surface antigens and also improves the efficiency of antigen recovery compared with traditional detergent extraction and surface radio-iodination. The method can also be used with ex-vivo parasites. Methods After surface labelling with biotin in the presence of the inhibitor furosemide, detergent extraction and osmotic lysis methods of enriching for the membrane fractions were compared to determine the efficiency of purification and recovery. Biotin-labelled proteins were identified on silver-stained SDS-polyacrylamide gels. Results Detergent extraction and osmotic lysis were compared for their capacity to purify biotin-labelled Plasmodium falciparum and Plasmodium chabaudi erythrocyte surface antigens. The pellet fraction formed after osmotic lysis of P. falciparum-infected erythrocytes is notably enriched in suface antigens, including PfEMP1, when compared to detergent extraction. There is also reduced co-extraction of host proteins such as spectrin and Band 3. Conclusion Biotinylation and osmotic lysis provides an improved method to label and purify parasitised erythrocyte surface antigen extracts from both in vitro and ex vivo Plasmodium parasite preparations.

  12. Nephritogenic antigen determinants in epidermal and renal basement membranes of kindreds with Alport-type familial nephritis.

    OpenAIRE

    Kashtan, C; Fish, A J; Kleppel, M; Yoshioka, K; Michael, A F

    1986-01-01

    We probed epidermal basement membranes (EBM) of acid-urea denatured skin from members of kindreds with Alport-type familial nephritis (FN) for the presence of antigens reactive with Goodpasture sera (GPS) and serum (FNS) from an Alport patient who developed anti-glomerular basement membrane (GBM) nephritis in a renal allograft. By immunoblotting, GPS reacted primarily with the 28,000 molecular weight (mol wt) monomer but also the 24,000 mol wt and 26,000 mol wt monomers of the noncollagenous ...

  13. Harnessing Dendritic Cells for Tumor Antigen Presentation

    Energy Technology Data Exchange (ETDEWEB)

    Nierkens, Stefan [Department of Tumor Immunology, Nijmegen Centre for Molecular Life Sciences, Radboud University Nijmegen Medical Centre, Geert Grooteplein 28, Nijmegen 6525 GA (Netherlands); Janssen, Edith M., E-mail: edith.janssen@cchmc.org [Division of Molecular Immunology, Cincinnati Children' s Hospital Research Foundation, University of Cincinnati College of Medicine, 3333 Burnet Avenue, Cincinnati, OH 45229 (United States)

    2011-04-26

    Dendritic cells (DC) are professional antigen presenting cells that are crucial for the induction of anti-tumor T cell responses. As a consequence, research has focused on the harnessing of DCs for therapeutic interventions. Although current strategies employing ex vivo-generated and tumor-antigen loaded DCs have been proven feasible, there are still many obstacles to overcome in order to improve clinical trial successes and offset the cost and complexity of customized cell therapy. This review focuses on one of these obstacles and a pivotal step for the priming of tumor-specific CD8{sup +} and CD4{sup +} T cells; the in vitro loading of DCs with tumor antigens.

  14. T Lymphocyte-Endothelial Interactions: Emerging Understanding of Trafficking and Antigen-Specific Immunity

    Directory of Open Access Journals (Sweden)

    Christopher Vincent Carman

    2015-11-01

    Full Text Available Antigen-specific immunity requires regulated trafficking of T cells in and out of diverse tissues in order to orchestrate lymphocyte development, immune surveillance, responses and memory. The endothelium serves as a unique barrier, as well as a sentinel, between the blood and the tissues and as such it plays an essential locally tuned role in regulating T cell migration and information exchange. While it is well established that chemoattractants and adhesion molecules are major determinants of T cell trafficking, emerging studies have now enumerated a large number of molecular players as well as a range of discrete cellular remodeling activities (e.g. transmigratory cups and invadosome-like protrusions, IPLs that participate in directed migration and pathfinding by T cells. In addition to providing trafficking cues, intimate cell-cell interaction between lymphocytes and endothelial cells provide instruction to T cells that influence their activation and differentiation states. Perhaps the most intriguing and underappreciated of these ‘sentinel’ roles is the ability of the endothelium to act as a non-hematopoietic ‘semi-professional’ antigen-presenting cell. Close contacts between circulating T cells and antigen-presenting endothelium may play unique non-redundant roles in shaping adaptive immune responses within the periphery. A better understanding of the mechanisms directing T cell trafficking and the antigen-presenting role of the endothelium may not only increase our knowledge of the adaptive immune response but also empower the utility of emerging immunomodulatory therapeutics.

  15. The physico-chemical, antigenic, and functional heterogeneity of human serum amyloid A

    International Nuclear Information System (INIS)

    Martin, M.E.; HUq, A.; Rosenthal, C.J.

    1986-01-01

    In the present study we attempted to develop a rapid method to isolate serum amyloid A isomers (SAA is.) and to determine whether this physicochemical heterogeneity corresponds to an antigenic and functional one. Pure human low molecular SAA (SAAL) was prepared from the serum of 6 patients (pts.) using standard techniques. Preparative isoelectric focusing in agarose/sephadex gels was used to separate SAAL is. Monoclonal antibodies (m. abs.) to SAAL and to AA were prepared by hybridization of P3XU-1 nonsecretory murine myeloma cells with murine spleen cells from Balb/c mice immunized with pooled SAAL and AA respectively. Four distinctly migrating SAAL isomers with PI's of 4.9, 5.8, 6.6, and 7.2 were isolated from 6 pts. while only three isomers were separated from the pt. with myasthenia gravis. Four m. abs. to SAAL, one to AA, six m. abs. to SAAL-2 is. and one to SAAL-1 is. were generated in murine ascitic fluid. Dishes coated with the four human SAA is., human AA, various mammalian and human proteins as well as with serum from 31 pts. with metastatic Ca. and 23 pts. with inflammatory diseases (ID) were reacted with the m. abs. The amount of binding was determined using 125 I labelled goat antimouse serum. The m. abs. to SAA were found specific for human SAA recognizing two different patterns in relationship to the intensity of binding to SAA is. One of them (7A2-43) had a greater affinity for SAA from pts with ID, while the other (5A6-5) reacted stronger with SAA from pts with metastatic Ca

  16. Cross-reactive Legionella antigens and the antibody response during infection

    DEFF Research Database (Denmark)

    Bangsborg, Jette Marie; Shand, G; Pearlman, E

    1991-01-01

    In order to define cross-reactive Legionella antigens suitable for diagnostic purposes, we investigated sonicate antigens from two Legionella species, including two serogroups of L. pneumophila. The antigens were reacted with heterologous and homologous rabbit antisera in Western blot. Sera from ...

  17. The molecular determinants of antibody recognition and antigenic drift in the H3 hemagglutinin of swine influenza A virus

    Science.gov (United States)

    Influenza A virus (IAV) of the H3 subtype is an important pathogen that affects both humans and swine. The main intervention strategy for preventing infection is vaccination to induce neutralizing antibodies against the surface glycoprotein hemagglutinin (HA). However, due to antigenic drift, vaccin...

  18. Investigation of the antigenic evolution of field isolates using the reverse genetics system of infectious bursal disease virus (IBDV).

    Science.gov (United States)

    Durairaj, Vijay; Sellers, Holly S; Linnemann, Erich G; Icard, Alan H; Mundt, Egbert

    2011-10-01

    The antigenic profiles of over 300 infectious bursal disease virus (IBDV) isolates were analyzed using a panel of monoclonal antibodies in a reverse genetics system. In addition, the sequences of a large portion of the neutralizing-antibody-inducing VP2 of IBDV were determined. Phylogenetic analysis of nucleotide and amino acid sequences in combination with the antigenic profiles obtained using the monoclonal antibody panel, revealed a lack of correlation between antigenicity and isolate's placement within the phylogenetic tree. In-depth analysis of amino acid exchanges revealed that changes within a certain region of the VP2 molecule resulted in differences in the antigenicity of the virus. This comprehensive analysis of VP2 sequences indicated a high selective pressure in the field that was likely due to vaccination programs, which increase the rate of evolution of the virus.

  19. Construction and expression of hepatitis B surface antigen escape variants within the "a" determinant by site directed mutagenesis.

    Science.gov (United States)

    Golsaz Shirazi, Forough; Amiri, Mohammad Mehdi; Mohammadi, Hamed; Bayat, Ali Ahmad; Roohi, Azam; Khoshnoodi, Jalal; Zarnani, Amir Hassan; Jeddi-Tehrani, Mahmood; Kardar, Gholam Ali; Shokri, Fazel

    2013-09-01

    The antibody response to hepatitis B surface antigen (HBsAg) controls hepatitis B virus infection. The "a" determinant of HBsAg is the most important target for protective antibody response, diagnosis and immunoprophylaxis. Mutations in this area may induce immune escape mutants and affect the performance of HBsAg assays. To construct clinically relevant recombinant mutant forms of HBsAg and assessment of their reactivity with anti-HBs monoclonal antibodies (MAbs). Wild type (wt) and mutant (mt) HBsAg genes were constructed by site directed mutagenesis and SEOing PCR. The amplified genes were inserted into pCMV6-neo plasmid and transfected in CHO cell line. The expression of wt- and mtHBsAg was assessed by commercial ELISA assays and stable cells were established and cloned by limiting dilution. The recombinant mutants were further characterized using a panel of anti-HBs monoclonal antibodies (MAbs) and the pattern of their reactivity was assessed by ELISA. Ten HBsAg mutants having single mutation within the "a" determinant including P120E, T123N, Q129H, M133L, K141E, P142S, D144A, G145R, N146S and C147S together with a wt form were successfully constructed and expressed in CHO cells. Reactivity of anti-HBs MAbs with mtHBsAgs displayed different patterns. The effect of mutations on antibody binding differed depending on the amino acid involved and its location within the ''a'' determinant. Mutation at amino acids 123 and 145 resulted in either complete loss or significant reduction of binding to all anti-HBs MAbs. Our panel of mtHBsAgs is a valuable tool for assessment of the antibody response to HBV escape mutants and may have substantial implications in HBV immunological diagnostics.

  20. Monoclonal antibodies to antigens on human neutrophils, activated T lymphocytes, and acute leukemia blast cells

    International Nuclear Information System (INIS)

    Miterev, G.Yu.; Burova, G.F.; Puzhitskaya, M.S.; Danilevich, S.V.; Bulycheva, T.I.

    1987-01-01

    The authors describe the production of two mouse hybridomas secreting monoclonal antibodies to antigenic determinants of the surface membranes of human neutrophils, activated T lymphocytes, and acute leukemic blast cells. The degree of lymphocyte stimulation was estimated from incorporation of 3 H-thymidine with parallel microculture. Monoclonal antibodies of supernatants of hybridoma cultures shown here reacted in both immunofluorescence test and cytotoxicity test with surface membrane antigens on the majority of neutrophils and PHA-activated peripheral blood lymphocytes from healthy subjects, but did not give positive reactions with unactivated lymphocytes, adherent monocytes, erythrocytes, and alloantigen-stimulated lymphocytes

  1. Monoclonal antibodies to antigens on human neutrophils, activated T lymphocytes, and acute leukemia blast cells

    Energy Technology Data Exchange (ETDEWEB)

    Miterev, G.Yu.; Burova, G.F.; Puzhitskaya, M.S.; Danilevich, S.V.; Bulycheva, T.I.

    1987-11-01

    The authors describe the production of two mouse hybridomas secreting monoclonal antibodies to antigenic determinants of the surface membranes of human neutrophils, activated T lymphocytes, and acute leukemic blast cells. The degree of lymphocyte stimulation was estimated from incorporation of /sup 3/H-thymidine with parallel microculture. Monoclonal antibodies of supernatants of hybridoma cultures shown here reacted in both immunofluorescence test and cytotoxicity test with surface membrane antigens on the majority of neutrophils and PHA-activated peripheral blood lymphocytes from healthy subjects, but did not give positive reactions with unactivated lymphocytes, adherent monocytes, erythrocytes, and alloantigen-stimulated lymphocytes.

  2. Antigen-specific cytotoxic T cell and antigen-specific proliferating T cell clones can be induced to cytolytic activity by monoclonal antibodies against T3

    NARCIS (Netherlands)

    Spits, H.; Yssel, H.; Leeuwenberg, J.; de Vries, J. E.

    1985-01-01

    T3 is a human differentiation antigen expressed exclusively on mature T cells. In this study it is shown that anti-T3 monoclonal antibodies, in addition to their capacity to induce T cells to proliferate, are able to induce antigen-specific cytotoxic T lymphocyte clones to mediate antigen

  3. Antigen-antibody reactions of UV-irradiated phage DNA

    International Nuclear Information System (INIS)

    Fink, A.

    1976-01-01

    The observation of others could be confirmed that UV-irradiated DNA is a better immunogen than unirradiated DNA. The author's immune sera contained a high amount of antibodies with a specific action against photoproducts in the DNA. The thymine dimer was identified as relevant photoproduct and thus as antigenic determinant. In comparison, the amount of unspecific antibodies reacting with denaturated DNA was low and varied between sera. Thymin-dimer antibodies showed a high specificity without cross-reaction with other pyrimidine dimers such as anti CC and anti CT; they belong to the class of IgG molecules. UV-irradiated dinucleotide dTpT is sufficient to induce the formation of antibodies reacting with the cis-syn thymine dimers in UV-irradiated DNA. Antibody binding is proportional to the UV doses applied to the DNA. When using completely denaturated DNA, there is a linear increase changing into a plateau at higher doses. The extent of antigen-antibody binding is strongly dependent on the degree of denaturation of the DNA. With increasing denaturation, the antibody binding of the DNA increases. The antigen-antibody reaction can thus be used to estimate the degree of denaturation of the DNA. There were no signs of an influence of the degree of denaturation of the DNA on the quantum yield of thymine dimers. The different amounts of antibodies is therefore due to the masking of thymine dimers in native DNA. When irradiating intact phage particles, there was no sign of an influence of the phages' protein covers on the antibody binding capacity of DNA compared with DNA irradiated in vitro. (orig.) [de

  4. Comparison of microglia and infiltrating CD11c+ cells as antigen presenting cells for T cell proliferation and cytokine response

    DEFF Research Database (Denmark)

    Wlodarczyk, Agnieszka; Løbner, Morten; Cédile, Oriane

    2014-01-01

    BACKGROUND: Tissue-resident antigen-presenting cells (APC) exert a major influence on the local immune environment. Microglia are resident myeloid cells in the central nervous system (CNS), deriving from early post-embryonic precursors, distinct from adult hematopoietic lineages. Dendritic cells...... (DC) and macrophages infiltrate the CNS during experimental autoimmune encephalomyelitis (EAE). Microglia are not considered to be as effective APC as DC or macrophages. METHODS: In this work we compared the antigen presenting capacity of CD11c+ and CD11c- microglia subsets with infiltrating CD11c......+ APC, which include DC. The microglial subpopulations (CD11c- CD45dim CD11b+ and CD11c+ CD45dim CD11b+) as well as infiltrating CD11c+ CD45high cells were sorted from CNS of C57BL/6 mice with EAE. Sorted cells were characterised by flow cytometry for surface phenotype and by quantitative real-time PCR...

  5. Immunoradiometric quantitation of tissue plasminogen activator-related antigen in human plasma: crypticity phenomenon and relationship to plasma fibrinolysis

    International Nuclear Information System (INIS)

    Wun, T.C.; Capuano, A.

    1987-01-01

    A two-site immunoradiometric assay for tissue plasminogen activator (tPA) antigen has been developed using immunoaffinity purified antibody. Various treatments enhanced the detection of tPA antigen in the plasma samples. Maximum detection was obtained by acidification of plasma to pH 4.8 to 6.5 or addition of 0.5 mol/L of L-lysine or L-arginine. Acidification or addition of lysine to plasma is also required for maximum immunoadsorption of plasma tPA antigen on anti-tPA-Ig-sepharose. These results indicate that plasma tPA antigen is partially cryptic to antibody in untreated plasma. The plasma tPA antigen isolated by immunoadsorption of either untreated plasma or acidified plasma on anti-tPA-Ig-sepharose consists mainly of a 100-kd plasminogen activator species as determined by fibrin-agar zymography. The 100-kd activity is possibly a tPA:inhibitor complex. A standardized sample preparation method was conveniently adopted by mixing 3 vol of plasma and 1 vol of 2 mol/L of L-lysine for the assay. Reconstitution and recovery studies showed that the method is specific and permits full detection of both free tPA and tPA:inhibitor complex. The validity of the assay is further supported by the finding that the spontaneous plasma fibrinolysis previously demonstrated to be dependent on plasma tPA antigen is correlated with tPA antigen content. Using the standardized assay, we found that tPA antigen concentrations in 16 blood bank plasmas are equivalent to 3.7 to 20 ng of 60 kd tPA/mL. In all the plasma tested, more than half of the antigen is undetected unless the plasma is treated as described above

  6. Immunoradiometric quantitation of tissue plasminogen activator-related antigen in human plasma: crypticity phenomenon and relationship to plasma fibrinolysis

    Energy Technology Data Exchange (ETDEWEB)

    Wun, T.C.; Capuano, A.

    1987-05-01

    A two-site immunoradiometric assay for tissue plasminogen activator (tPA) antigen has been developed using immunoaffinity purified antibody. Various treatments enhanced the detection of tPA antigen in the plasma samples. Maximum detection was obtained by acidification of plasma to pH 4.8 to 6.5 or addition of 0.5 mol/L of L-lysine or L-arginine. Acidification or addition of lysine to plasma is also required for maximum immunoadsorption of plasma tPA antigen on anti-tPA-Ig-sepharose. These results indicate that plasma tPA antigen is partially cryptic to antibody in untreated plasma. The plasma tPA antigen isolated by immunoadsorption of either untreated plasma or acidified plasma on anti-tPA-Ig-sepharose consists mainly of a 100-kd plasminogen activator species as determined by fibrin-agar zymography. The 100-kd activity is possibly a tPA:inhibitor complex. A standardized sample preparation method was conveniently adopted by mixing 3 vol of plasma and 1 vol of 2 mol/L of L-lysine for the assay. Reconstitution and recovery studies showed that the method is specific and permits full detection of both free tPA and tPA:inhibitor complex. The validity of the assay is further supported by the finding that the spontaneous plasma fibrinolysis previously demonstrated to be dependent on plasma tPA antigen is correlated with tPA antigen content. Using the standardized assay, we found that tPA antigen concentrations in 16 blood bank plasmas are equivalent to 3.7 to 20 ng of 60 kd tPA/mL. In all the plasma tested, more than half of the antigen is undetected unless the plasma is treated as described above.

  7. Radioimmunoassay for a human prostate specific antigen

    International Nuclear Information System (INIS)

    Machida, T.; Miki, M.; Ohishi, Y.; Kido, A.; Morikawa, J.; Ogawa, Y.

    1983-01-01

    As a marker for prostatic cancer, a prostate-specific antigen was purified from human prostatic tissues. Double antibody radioimmunoassay utilizing immune reaction was developed on the basis of the purified prostatic antigen (PA). Measurement results have revealed that PA radioimmunoassay is much better than prostatic acid phosphatase (PAP) radioimmunoassay in the diagnosis of prostatic cancer

  8. Antigen detection systems

    Science.gov (United States)

    Infectious agents or their constituent parts (antigens or nucleic acids) can be detected in fresh, frozen, or fixed tissues or other specimens, using a variety of direct or indirect assays. The assays can be modified to yield the greatest sensitivity and specificity but in most cases a particular m...

  9. Humoral and cell-mediated immune response against human retinal antigens in relation to ocular onchocerciasis

    NARCIS (Netherlands)

    van der Lelij, A.; Rothova, A.; Stilma, J. S.; Vetter, J. C.; Hoekzema, R.; Kijlstra, A.

    1990-01-01

    Autoimmune mechanisms are thought to be involved in the pathogenesis of the chorioretinal changes in ocular onchocerciasis. The humoral autoimmune response was determined by measuring serum levels of autoantibodies, directed against human S-antigen and interphotoreceptor retinoid binding protein

  10. Immunization against Rabies with Plant-Derived Antigen

    Science.gov (United States)

    Modelska, Anna; Dietzschold, Bernard; Sleysh, N.; Fu, Zhen Fang; Steplewski, Klaudia; Hooper, D. Craig; Koprowski, Hilary; Yusibov, Vidadi

    1998-03-01

    We previously demonstrated that recombinant plant virus particles containing a chimeric peptide representing two rabies virus epitopes stimulate virus neutralizing antibody synthesis in immunized mice. We show here that mice immunized intraperitoneally or orally (by gastric intubation or by feeding on virus-infected spinach leaves) with engineered plant virus particles containing rabies antigen mount a local and systemic immune response. After the third dose of antigen, given intraperitoneally, 40% of the mice were protected against challenge infection with a lethal dose of rabies virus. Oral administration of the antigen stimulated serum IgG and IgA synthesis and ameliorated the clinical signs caused by intranasal infection with an attenuated rabies virus strain.

  11. The Antigen Presenting Cells Instruct Plasma Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Wei eXu

    2014-01-01

    Full Text Available The professional antigen presenting cells (APCs, including many subsets of dendritic cells and macrophages, not only mediate prompt but nonspecific response against microbes, but also bridge the antigen-specific adaptive immune response through antigen presentation. In the latter, typically activated B cells acquire cognate signals from T helper cells in the germinal center of lymphoid follicles to differentiate into plasma cells, which generate protective antibodies. Recent advances have revealed that many APC subsets provide not only signal 1 (the antigen, but also signal 2 to directly instruct the differentiation process of plasma cells in a T cell-independent manner. Herein, the different signals provided by these APC subsets to direct B cell proliferation, survival, class switching and terminal differentiation are discussed. We furthermore propose that the next generation of vaccines for boosting antibody response could be designed by targeting APCs.

  12. The antigen presenting cells instruct plasma cell differentiation.

    Science.gov (United States)

    Xu, Wei; Banchereau, Jacques

    2014-01-06

    The professional antigen presenting cells (APCs), including many subsets of dendritic cells and macrophages, not only mediate prompt but non-specific response against microbes, but also bridge the antigen-specific adaptive immune response through antigen presentation. In the latter, typically activated B cells acquire cognate signals from T helper cells in the germinal center of lymphoid follicles to differentiate into plasma cells (PCs), which generate protective antibodies. Recent advances have revealed that many APC subsets provide not only "signal 1" (the antigen), but also "signal 2" to directly instruct the differentiation process of PCs in a T-cell-independent manner. Herein, the different signals provided by these APC subsets to direct B cell proliferation, survival, class switching, and terminal differentiation are discussed. We furthermore propose that the next generation of vaccines for boosting antibody response could be designed by targeting APCs.

  13. Mucosal Blood Group Antigen Expression Profiles and HIV Infections: A Study among Female Sex Workers in Kenya.

    Directory of Open Access Journals (Sweden)

    Nadia Musimbi Chanzu

    Full Text Available The ABO blood group antigens are carbohydrate moieties expressed on human red blood cells however; these antigens can also be expressed on some other cells particularly the surface of epithelial cells and may be found in mucosal secretions. In many human populations 80% secrete ABO antigens (termed 'secretors' while 20% do not (termed 'non-secretors'. Furthermore, there are disease conditions that are associated with secretor status.To investigate correlations between secretor status and HIV infection among female sex workers in Nairobi, Kenya.This cross-sectional study recruited 280 female sex workers aged 18-65 years from the Pumwani Majengo cohort, Kenya. Blood typing was determined by serological techniques using monoclonal antibodies to the ABO blood group antigens. Secretor phenotyping was determined using anti-H specific lectins specific to salivary, vaginal and cervical blood group H antigen using the agglutination inhibition technique and correlated to individual HIV sero-status. Participants were additionally screened for Bacterial vaginosis, Neisseria gonorrhoea and Trichomonas vaginalis.Out of the 280 participants, 212 (75.7% were secretors and 68 (24.3% were non-secretors. The incidence of all infections: HIV, Bacterial vaginosis, Neisseria gonorrhoea and Trichomonas vaginalis was higher among secretors compared to non-secretors. However, this difference was only statistically significant for HIV infection incidence rates: HIV infected secretors (83.7% versus HIV un-infected secretors (71.8% (p = 0.029 Based on ABO phenotype stratification, the incidence of HIV infection was higher among blood group A secretors (26/52 = 50%, in comparison to B (12/39 = 33.3%: p = 0.066, AB (3/9 = 33.3%: p = 0.355, and O secretors (36/112 = 32.1%: p = 0.028.This is the first report to document the variable expression of the ABH blood group antigens profiling secretor and non-secretor phenotypes in the female genital tract among a high-risk population

  14. Effectiveness of the combined evaluation of KLK3 genetics and free-to-total prostate specific antigen ratio for prostate cancer diagnosis.

    Science.gov (United States)

    Zambon, Carlo-Federico; Prayer-Galetti, Tommaso; Basso, Daniela; Padoan, Andrea; Rossi, Elisa; Secco, Silvia; Pelloso, Michela; Fogar, Paola; Navaglia, Filippo; Moz, Stefania; Zattoni, Filiberto; Plebani, Mario

    2012-10-01

    Of serum prostate specific antigen variability 40% depends on inherited factors. We ascertained whether the knowledge of KLK3 genetics would enhance prostate specific antigen diagnostic performance in patients with clinical suspicion of prostate cancer. We studied 1,058 men who consecutively underwent prostate biopsy for clinical suspicion of prostate cancer. At histology prostate cancer was present in 401 cases and absent in 657. Serum total prostate specific antigen and the free-to-total prostate specific antigen ratio were determined. Four polymorphisms of the KLK3 gene (rs2569733, rs2739448, rs925013 and rs2735839) and 1 polymorphism of the SRD5A2 gene (rs523349) were studied. The influence of genetics on prostate specific antigen variability was evaluated by multivariate linear regression analysis. The performance of total prostate specific antigen and the free-to-total prostate specific antigen ratio alone or combined with a genetically based patient classification were defined by ROC curve analyses. For prostate cancer diagnosis the free-to-total prostate specific antigen ratio index alone (cutoff 11%) was superior to total prostate specific antigen (cutoff 4 ng/ml) and to free-to-total prostate specific antigen ratio reflex testing (positive predictive value 61%, 43% and 54%, respectively). Prostate specific antigen correlated with KLK3 genetics (rs2735839 polymorphism p = 0.001, and rs2569733, rs2739448 and rs925013 haplotype combination p = 0.003). In patients with different KLK3 genetics 2 optimal free-to-total prostate specific antigen ratio cutoffs (11% and 14.5%) were found. For free-to-total prostate specific antigen ratio values between 11% and 14.5% the prostate cancer probability ranged from 30.0% to 47.4% according to patient genetics. The free-to-total prostate specific antigen ratio is superior to total prostate specific antigen for prostate cancer diagnosis, independent of total prostate specific antigen results. Free-to-total prostate

  15. Reduction-sensitive dextran nanogels aimed for intracellular delivery of antigens

    NARCIS (Netherlands)

    Li, Dandan; Kordalivand, Neda; Fransen, Marieke F.; Ossendorp, Ferry; Raemdonck, Koen; Vermonden, Tina; Hennink, Wim E.; Van Nostrum, Cornelus F.

    2015-01-01

    Targeting of antigens to dendritic cells (DCs) to induce strong cellular immune response can be established by loading in a nano-sized carrier and keeping the antigen associated with the particles until they are internalized by DCs. In the present study, a model antigen (ovalbumin, OVA) is

  16. Dengue NS1 Antigen - for Early Detection of Dengue Virus Infection

    Directory of Open Access Journals (Sweden)

    Amol Hartalkar

    2015-08-01

    Full Text Available Objectives: To evaluate the efficacy of NS1 antigen assay for early diagnosis of dengue virus infection in a tertiary care hospital. Methods: This cross sectional study was carried out in department of Medicine from August to December 2013. Total 100 patients with dengue fever were included. Complete blood count, alanine aminotransferase (ALT, aspartate aminotransferase (AST, Dengue NS1 antigen and IgM and IgG antibodies of dengue virus were done in all cases. Results: Of the 100 sera tested, 75% were positive for dengue virus infection based on dengue NS1 antigen, IgM antibody and IgG antibody. Dengue NS1 antigen and IgM, IgG antibody were able to detect dengue virus infection between day 1 to day 8 in 92% of samples, 86.7% of samples and 82.6% of samples respectively. Sixty nine percent (69% were found positive for dengue NS1 antigen, 65% were IgM positive and 62% were IgG positive. Based on the dengue NS1 antigen and IgM antibody combination, 74% were positive for dengue virus infections. Sensitivity of Dengue NS1 antigen was 92.3% and specificity of 74.28% in comparison to IgM antibody. Detection rate increased to 75%, based on the antigen and IgG antibody combination. Sensitivity of dengue NS1 antigen was 90.3% and specificity of 65.8% in comparison to IgG antibody. Conclusion: Dengue NS1 antigen is a useful, sensitive and specific test for early diagnosis of dengue virus infection and it improves diagnostic efficiency in combination with antibody test. Key words: Dengue fever, NS1 antigen. Introduction: Dengue fever (DF is the most common arboviral illness in humans. Each year, an estimated 50-100 million cases of dengue fever and 500,000 cases of dengue hemorrhagic fever occur worldwide, with 30000 deaths (mainly in children. Globally 2.5-3 billion people in approximately 112 tropical and subtropical countries are at risk of dengue.of samples respectively. Sixty nine percent (69% were found positive for dengue NS1 antigen, 65% were Ig

  17. Mycoplasma fermentans glycolipid-antigen as a pathogen of rheumatoid arthritis

    International Nuclear Information System (INIS)

    Kawahito, Yutaka; Ichinose, Sizuko; Sano, Hajime; Tsubouchi, Yasunori; Kohno, Masataka; Yoshikawa, Toshikazu; Tokunaga, Daisaku; Hojo, Tatsuya; Harasawa, Ryo; Nakano, Teruaki; Matsuda, Kazuhiro

    2008-01-01

    Mycoplasma fermentans has been suspected as one of the causative pathogenic microorganisms of rheumatoid arthritis (RA) however, the pathogenic mechanism is still unclear. We, previously, reported that glycolipid-antigens (GGPL-I and III) are the major antigens of M. fermentans. Monoclonal antibody against the GGPL-III could detect the existence of the GGPL-III antigens in synovial tissues from RA patients. GGPL-III antigens were detected in 38.1% (32/84) of RA patient's tissues, but not in osteoarthritis (OA) and normal synovial tissues. Immunoelectron microscopy revealed that a part of GGPL-III antigens are located at endoplasmic reticulum. GGPL-III significantly induced TNF-α and IL-6 production from peripheral blood mononulear cells, and also proliferation of synovial fibroblasts. Further study is necessary to prove that M. fermentans is a causative microorganism of RA; however, the new mechanisms of disease pathogenesis provides hope for the development of effective and safe immunotherapeutic strategies based on the lipid-antigen, GGPL-III, in the near future

  18. Identification of Low- and High-Impact Hemagglutinin Amino Acid Substitutions That Drive Antigenic Drift of Influenza A(H1N1 Viruses.

    Directory of Open Access Journals (Sweden)

    William T Harvey

    2016-04-01

    Full Text Available Determining phenotype from genetic data is a fundamental challenge. Identification of emerging antigenic variants among circulating influenza viruses is critical to the vaccine virus selection process, with vaccine effectiveness maximized when constituents are antigenically similar to circulating viruses. Hemagglutination inhibition (HI assay data are commonly used to assess influenza antigenicity. Here, sequence and 3-D structural information of hemagglutinin (HA glycoproteins were analyzed together with corresponding HI assay data for former seasonal influenza A(H1N1 virus isolates (1997-2009 and reference viruses. The models developed identify and quantify the impact of eighteen amino acid substitutions on the antigenicity of HA, two of which were responsible for major transitions in antigenic phenotype. We used reverse genetics to demonstrate the causal effect on antigenicity for a subset of these substitutions. Information on the impact of substitutions allowed us to predict antigenic phenotypes of emerging viruses directly from HA gene sequence data and accuracy was doubled by including all substitutions causing antigenic changes over a model incorporating only the substitutions with the largest impact. The ability to quantify the phenotypic impact of specific amino acid substitutions should help refine emerging techniques that predict the evolution of virus populations from one year to the next, leading to stronger theoretical foundations for selection of candidate vaccine viruses. These techniques have great potential to be extended to other antigenically variable pathogens.

  19. Identification of Low- and High-Impact Hemagglutinin Amino Acid Substitutions That Drive Antigenic Drift of Influenza A(H1N1) Viruses

    Science.gov (United States)

    Harvey, William T.; Benton, Donald J.; Gregory, Victoria; Hall, James P. J.; Daniels, Rodney S.; Bedford, Trevor; Haydon, Daniel T.; Hay, Alan J.; McCauley, John W.; Reeve, Richard

    2016-01-01

    Determining phenotype from genetic data is a fundamental challenge. Identification of emerging antigenic variants among circulating influenza viruses is critical to the vaccine virus selection process, with vaccine effectiveness maximized when constituents are antigenically similar to circulating viruses. Hemagglutination inhibition (HI) assay data are commonly used to assess influenza antigenicity. Here, sequence and 3-D structural information of hemagglutinin (HA) glycoproteins were analyzed together with corresponding HI assay data for former seasonal influenza A(H1N1) virus isolates (1997–2009) and reference viruses. The models developed identify and quantify the impact of eighteen amino acid substitutions on the antigenicity of HA, two of which were responsible for major transitions in antigenic phenotype. We used reverse genetics to demonstrate the causal effect on antigenicity for a subset of these substitutions. Information on the impact of substitutions allowed us to predict antigenic phenotypes of emerging viruses directly from HA gene sequence data and accuracy was doubled by including all substitutions causing antigenic changes over a model incorporating only the substitutions with the largest impact. The ability to quantify the phenotypic impact of specific amino acid substitutions should help refine emerging techniques that predict the evolution of virus populations from one year to the next, leading to stronger theoretical foundations for selection of candidate vaccine viruses. These techniques have great potential to be extended to other antigenically variable pathogens. PMID:27057693

  20. Antigen-targeting strategies using single-domain antibody fragments

    NARCIS (Netherlands)

    Duarte, Joao Nuno Silva

    2017-01-01

    Antibodies display high selectivity and affinity and have been the preferred platform for antigen targeting. Despite the development of antigen-delivery systems that enable T cell activation, targeting approaches that enhance antibody responses need improvement. This need specially applies to poorly

  1. The evolutionary dynamics of variant antigen genes in Babesia reveal a history of genomic innovation underlying host-parasite interaction.

    Science.gov (United States)

    Jackson, Andrew P; Otto, Thomas D; Darby, Alistair; Ramaprasad, Abhinay; Xia, Dong; Echaide, Ignacio Eduardo; Farber, Marisa; Gahlot, Sunayna; Gamble, John; Gupta, Dinesh; Gupta, Yask; Jackson, Louise; Malandrin, Laurence; Malas, Tareq B; Moussa, Ehab; Nair, Mridul; Reid, Adam J; Sanders, Mandy; Sharma, Jyotsna; Tracey, Alan; Quail, Mike A; Weir, William; Wastling, Jonathan M; Hall, Neil; Willadsen, Peter; Lingelbach, Klaus; Shiels, Brian; Tait, Andy; Berriman, Matt; Allred, David R; Pain, Arnab

    2014-06-01

    Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5' ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  2. Manipulating the Lewis antigen specificity of the cholesterol-dependent cytolysin lectinolysin

    Directory of Open Access Journals (Sweden)

    Sara eLawrence

    2012-11-01

    Full Text Available The cholesterol-dependent cytolysins (CDCs attack cells by punching large holes in their membranes. Lectinolysin from Streptococcus mitis is unique among CDCs due to the presence of an N-terminal lectin domain that enhances the pore-forming activity of the toxin. We recently determined the crystal structures of the lectin domain in complex with various glycans. These structures revealed the molecular basis for the Lewis antigen specificity of the toxin. Based on this information we have used in silico molecular modelling to design a mutant toxin, which we predicted would increase its specificity for Lewis y, an antigen found on the surface of cancer cells. Surprisingly, we found by surface plasmon resonance binding experiments that the resultant mutant lectin domain exhibited higher specificity for Lewis b antigens instead. We then undertook comparative crystallographic and molecular dynamics simulation studies of the wild-type and mutant lectin domains to understand the molecular basis for the disparity between the theoretical and experimental results. The crystallographic results revealed that the net number of interactions between Lewis y and wild-type versus mutant was unchanged whereas there was a loss of a hydrogen bond between mutant and Lewis b compared to wild-type. In contrast, the molecular dynamics studies revealed that the Lewis b antigen spent more time in the binding pocket of the mutant compared to wild-type and the reverse was true for Lewis y. The results of these simulation studies are consistent with the conclusions drawn from the surface plasmon resonance studies. This work is part of a program to engineer lectinolysin so that it will target and kill specific cells in human diseases.

  3. Monoclonal antibodies to Nocardia asteroides and Nocardia brasiliensis antigens.

    OpenAIRE

    Jiménez, T; Díaz, A M; Zlotnik, H

    1990-01-01

    Nocardia asteroides and Nocardia brasiliensis whole-cell extracts were used as antigens to generate monoclonal antibodies (MAbs). Six stable hybrid cell lines secreting anti-Nocardia spp. MAbs were obtained. These were characterized by enzyme-linked immunosorbent assay, Western blot (immunoblot), and immunofluorescence assay. Although all the MAbs exhibited different degrees of cross-reactivity with N. asteroides and N. brasiliensis antigens as well as with culture-filtrate antigens from Myco...

  4. Use of a solid-phase radioimmunoassay and formalin-fixed whole bacterial antigen in the detection of antigen-specific immunoglobulin in prostatic fluid.

    OpenAIRE

    Shortliffe, L M; Wehner, N; Stamey, T A

    1981-01-01

    The prostatic fluid of two patients with Escherichia coli bacterial prostatitis was analyzed for evidence of a local immune response to bacterial infection. A solid-phase radioimmunoassay was modified to measure the immunoglobulin (Ig)A and IgG antigen-specific antibody responses to infecting bacteria in serum and prostatic fluid from patient. Formalin-fixed whole E. coli were used as antigen. In one patient with acute E. coli prostatic infection, measurements of antigen-specific antibody con...

  5. Prostate-specific antigen patterns in US and European populations : Comparison of six diverse cohorts

    NARCIS (Netherlands)

    Simpkin, Andrew J.; Donovan, Jenny L.; Tilling, Kate; Athene Lane, J.; Martin, Richard M.; Albertsen, Peter C.; Bill-Axelson, Anna; Ballentine Carter, H.; Bosch, J. L H Ruud; Ferrucci, Luigi; Hamdy, Freddie C.; Holmberg, Lars; Jeffrey Metter, E.; Neal, David E.; Parker, Christopher C.; Metcalfe, Chris

    2016-01-01

    Objective: To determine whether there are differences in prostate-specific antigen (PSA) levels at diagnosis or changes in PSA levels between US and European populations of men with and without prostate cancer (PCa). Subjects and Methods: We analysed repeated measures of PSA from six clinically and

  6. One target, different effects: a comparison of distinct therapeutic antibodies against the same targets.

    Science.gov (United States)

    Shim, Hyunbo

    2011-10-31

    To date, more than 30 antibodies have been approved worldwide for therapeutic use. While the monoclonal antibody market is rapidly growing, the clinical use of therapeutic antibodies is mostly limited to treatment of cancers and immunological disorders. Moreover, antibodies against only five targets (TNF-α, HER2, CD20, EGFR, and VEGF) account for more than 80 percent of the worldwide market of therapeutic antibodies. The shortage of novel, clinically proven targets has resulted in the development of many distinct therapeutic antibodies against a small number of proven targets, based on the premise that different antibody molecules against the same target antigen have distinct biological and clinical effects from one another. For example, four antibodies against TNF-α have been approved by the FDA -- infliximab, adalimumab, golimumab, and certolizumab pegol -- with many more in clinical and preclinical development. The situation is similar for HER2, CD20, EGFR, and VEGF, each having one or more approved antibodies and many more under development. This review discusses the different binding characteristics, mechanisms of action, and biological and clinical activities of multiple monoclonal antibodies against TNF-α, HER-2, CD20, and EGFR and provides insights into the development of therapeutic antibodies.

  7. Selection of antigenic markers on a GFP-Cκ fusion scaffold with high sensitivity by eukaryotic ribosome display

    International Nuclear Information System (INIS)

    Yang Yongmin; Barankiewicz, Teresa J.; He Mingyue; Taussig, Michael J.; Chen, Swey-Shen

    2007-01-01

    Ribosome display is a cell-free system permitting gene selection through the physical association of genetic material (mRNA) and its phenotypic (protein) product. While often used to select single-chain antibodies from large libraries by panning against immobilized antigens, we have adapted ribosome display for use in the 'reverse' format in order to select high affinity antigenic determinants against solid-phase antibody. To create an antigenic scaffold, DNA encoding green fluorescent protein (GFP) was fused to a light chain constant domain (Cκ) with stop codon deleted, and with 5' signals (T7 promoter, Kozak) enabling coupled transcription/translation in a eukaryotic cell-free system. Epitopes on either GFP (5') or Cκ (3') were selected by anti-GFP or anti-Cκ antibodies, respectively, coupled to magnetic beads. After selection, mRNA was amplified directly from protein-ribosome-mRNA (PRM) complexes by in situ PCR followed by internal amplification and reassembly PCR. As little as 10 fg of the 1 kb DNA construct, i.e. approximately 7500 molecules, could be recovered following a single round of interaction with solid-phase anti-GFP antibody. This platform is highly specific and sensitive for the antigen-antibody interaction and may permit selection and reshaping of high affinity antigenic variants of scaffold proteins

  8. Molecular characterization of Marek's disease herpesvirus B antigen

    International Nuclear Information System (INIS)

    Isfort, R.J.; Sithole, I.; Kung, H.J.; Velicer, L.F.

    1986-01-01

    The Marek's disease herpesvirus (MDHV) B antigen (MDHV-B) was identified and molecularly characterized as a set of three glycoproteins of 100,000, 60,000, and 49,000 apparent molecular weight (gp100, gp60, and gp49, respectively) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) after immunoprecipitation from [ 35 S]methionine-labeled infected cells by specific rabbit antiserum directed against MDHV-B (RαB), as previously determined by immunodiffusion. Further identification was accomplished by blocking this immunoprecipitation with highly purified MDHV-B. The same set of three polypeptides was also immunoprecipitated from [ 35 S] methionine- and 14 C-labeled infected cells into two other sera shown to have anti-B activity. These data serve to clarify the molecular identification of the polypeptides found in common between MDHV and HVT by linking them to MDHV-B. Collectively, the data presented here and by others support the conclusion that all three glycoproteins now identified as gp100, gp60, and gp49 have MDHV-B determinants. Finally, detection of the same three polypeptides with well-absorbed RαPM, which was directed against purified infected-cell plasma membranes, suggests that at least one component of the B-antigen complex has a plasma membrane location in the infected cell. These preliminary data point to the future membrane biochemistry and membrane immunology experiments needed to understand the MDHV system, and they may explain the high level of immunogenicity of MDHV-B in the infected chicken, as shown by its immunoprecipitation with immune chicken serum

  9. Distinct effects on diversifying selection by two mechanisms of immunity against Streptococcus pneumoniae.

    Directory of Open Access Journals (Sweden)

    Yuan Li

    Full Text Available Antigenic variation to evade host immunity has long been assumed to be a driving force of diversifying selection in pathogens. Colonization by Streptococcus pneumoniae, which is central to the organism's transmission and therefore evolution, is limited by two arms of the immune system: antibody- and T cell- mediated immunity. In particular, the effector activity of CD4(+ T(H17 cell mediated immunity has been shown to act in trans, clearing co-colonizing pneumococci that do not bear the relevant antigen. It is thus unclear whether T(H17 cell immunity allows benefit of antigenic variation and contributes to diversifying selection. Here we show that antigen-specific CD4(+ T(H17 cell immunity almost equally reduces colonization by both an antigen-positive strain and a co-colonized, antigen-negative strain in a mouse model of pneumococcal carriage, thus potentially minimizing the advantage of escape from this type of immunity. Using a proteomic screening approach, we identified a list of candidate human CD4(+ T(H17 cell antigens. Using this list and a previously published list of pneumococcal Antibody antigens, we bioinformatically assessed the signals of diversifying selection among the identified antigens compared to non-antigens. We found that Antibody antigen genes were significantly more likely to be under diversifying selection than the T(H17 cell antigen genes, which were indistinguishable from non-antigens. Within the Antibody antigens, epitopes recognized by human antibodies showed stronger evidence of diversifying selection. Taken together, the data suggest that T(H17 cell-mediated immunity, one form of T cell immunity that is important to limit carriage of antigen-positive pneumococcus, favors little diversifying selection in the targeted antigen. The results could provide new insight into pneumococcal vaccine design.

  10. Hepatitis B virus core antigen determines viral persistence in a C57BL/6 mouse model.

    Science.gov (United States)

    Lin, Yi-Jiun; Huang, Li-Rung; Yang, Hung-Chih; Tzeng, Horng-Tay; Hsu, Ping-Ning; Wu, Hui-Lin; Chen, Pei-Jer; Chen, Ding-Shinn

    2010-05-18

    We recently developed a mouse model of hepatitis B virus (HBV) persistence, in which a single i.v. hydrodynamic injection of HBV DNA to C57BL/6 mice allows HBV replication and induces a partial immune response, so that about 20-30% of the mice carry HBV for more than 6 months. The model was used to identify the viral antigen crucial for HBV persistence. We knocked out individual HBV genes by introducing a premature termination codon to the HBV core, HBeAg, HBx, and polymerase ORFs. The specific-gene-deficient HBV mutants were hydrodynamically injected into mice and the HBV profiles of the mice were monitored. About 90% of the mice that received the HBcAg-mutated HBV plasmid exhibited high levels of hepatitis B surface antigenemia and maintained HBsAg expression for more than 6 months after injection. To map the region of HBcAg essential for viral clearance, we constructed a set of serial HBcAg deletion mutants for hydrodynamic injection. We localized the essential region of HBcAg to the carboxyl terminus, specifically to the 10 terminal amino acids (HBcAg176-185). The majority of mice receiving this HBV mutant DNA did not elicit a proper HBcAg-specific IFN-gamma response and expressed HBV virions for 6 months. These results indicate that the immune response triggered in mice by HBcAg during exposure to HBV is important in determining HBV persistence.

  11. Serum carcinoembryonic antigen tends to decrease in poorly-differentiated colorectal cancer

    Directory of Open Access Journals (Sweden)

    Ester Morina Silalahi

    2015-12-01

    This was a cross-sectional study conducted on 40 CRC subjects from July 2012 until May 2013. Determination of serum CEA and CA 19-9 levels and histopathological (cellular differentiation grades in CRC biopsies was done in all subjects. RESULTS The study involved forty CRC patients, consisting of 22 males and 18 females, with mean age of 51.93 ± 11.63 years, CEA levels of 51.93 ± 84.07 ng/ml and CA 19-9 levels of 33.81 ± 62.39 U/ml. Carcino-embryonic antigen levels tended to decrease with decreasing CRC histopathological grade, while CA 19-9 levels increased in well-differentiated CRC. However, both relationships were statistically not significant (with p=0.314 and p=0.787, respectively. CONCLUSIONS Carcinoembryonic antigen (CEA levels tend to decrease with decreasing histopathological grade of CRC, and CA 19-9 levels tend to increase in well-differentiated CRC.

  12. Association Between PAI-1 Activity Levels and t-PA Antigen with Glycemic Status in Prediabetic Population

    Directory of Open Access Journals (Sweden)

    Andi Fachruddin Benyamin

    2016-11-01

    Full Text Available Aim: to evaluate an association between fibrinolysis defect and glycemic status in prediabetic population by assessing the levels of t-PA antigen and PAI-1 activity. Methods: it was an observational study with cross-sectional approach. There were 72 subjects aged 30-50 years who had met the inclusion criteria. The diagnosis of diabetes mellitus (DM and glycemic index were determined based on the American Diabetes Association (ADA criteria. The PAI-1 and t-PA antigen levels were measured quantitatively using enzyme-linked immunosorbent assay (ELISA. Analysis between the levels of t-PA antigen and PAI-1 activity was performed using ANOVA. Results: the t-PA antigen level was significantly higher in subjects with impaired glucose tolerance (IGT and impaired fasting blood glucose (IFBG as well as subject with impaired fasting blood glucose (IFBG than those with normal glucose tolerance (NGT (p=0.047. The PAI-1 activity was significantly higher in subjects with IGT, IFBG and subjects with IFBG than NGT (p=0.024. There was a significant association between glycemic status in prediabetic subjects and PAI-1 activity (p=0.04. Conclusion: the level of t-PA antigen and PAI-1 activity were significantly higher in prediabetic subjects than those with NGT; and there was a significant association between glycemic status in prediabetic subjects and PAI-1 activity.

  13. Toxoplasma gondii: II. Tachyzoite antigenic characterization of eigth strains

    Directory of Open Access Journals (Sweden)

    Regina Mitsuka

    1998-01-01

    Full Text Available Eight Toxoplasma gondii strains were analyzed using ELISA and Western blot techniques, in order to demonstrate possible immunological differences. The analyzed strains were: LIV IV, LIV V and S 11 isolated from swine, RH and VPS from a human being, AS 28 from a wild mouse, HV III from a dog and CN from a cat. With the ELISA assay the eight strains showed similar reactivity with homologous and heterologous sera. The antigenic suspension, consisting of total cellular extract of tachyzoites, was effective in the indirect ELISA assay, with the positive sera reacting strongly and the negative not reacting with the antigens. The Western blot analysis showed that the T. gondii strains have similar antigenic profiles with a few variations. Three bands were observed in all strains: one of about 33 kDa (p33, another of 54 kDa (p54 and a third one of 66 kDa (p66. The HV III strain, isolated from a dog, did not show three antigens (50, 70 and 75 kDa that were present in the others. However, this difference was not detected by the ELISA assay. Only two antigens (62 kDa of the CN and 67 kDa of the LIV IV were strain-specific antigens.

  14. Unpolarized release of vaccinia virus and HIV antigen by colchicine treatment enhances intranasal HIV antigen expression and mucosal humoral responses.

    Directory of Open Access Journals (Sweden)

    Yan Zhang

    Full Text Available The induction of a strong mucosal immune response is essential to building successful HIV vaccines. Highly attenuated recombinant HIV vaccinia virus can be administered mucosally, but even high doses of immunization have been found unable to induce strong mucosal antibody responses. In order to solve this problem, we studied the interactions of recombinant HIV vaccinia virus Tiantan strain (rVTT-gagpol in mucosal epithelial cells (specifically Caco-2 cell layers and in BALB/c mice. We evaluated the impact of this virus on HIV antigen delivery and specific immune responses. The results demonstrated that rVTT-gagpol was able to infect Caco-2 cell layers and both the nasal and lung epithelia in BALB/c mice. The progeny viruses and expressed p24 were released mainly from apical surfaces. In BALB/c mice, the infection was limited to the respiratory system and was not observed in the blood. This showed that polarized distribution limited antigen delivery into the whole body and thus limited immune response. To see if this could be improved upon, we stimulated unpolarized budding of the virus and HIV antigens by treating both Caco-2 cells and BALB/c mice with colchicine. We found that, in BALB/c mice, the degree of infection and antigen expression in the epithelia went up. As a result, specific immune responses increased correspondingly. Together, these data suggest that polarized budding limits antigen delivery and immune responses, but unpolarized distribution can increase antigen expression and delivery and thus enhance specific immune responses. This conclusion can be used to optimize mucosal HIV vaccine strategies.

  15. Biological variation of total prostate-specific antigen

    DEFF Research Database (Denmark)

    Söletormos, Georg; Semjonow, Axel; Sibley, Paul E C

    2005-01-01

    BACKGROUND: The objectives of this study were to determine whether a single result for total prostate-specific antigen (tPSA) can be used confidently to guide the need for prostate biopsy and by how much serial tPSA measurements must differ to be significant. tPSA measurements include both...... analytical and biological components of variation. The European Group on Tumor Markers conducted a literature survey to determine both the magnitude and impact of biological variation on single, the mean of replicate, and serial tPSA measurements. METHODS: The survey yielded 27 studies addressing the topic......, and estimates for the biological variation of tPSA could be derived from 12 of these studies. RESULTS: The mean biological variation was 20% in the concentration range 0.1-20 microg/L for men over 50 years. The biological variation means that the one-sided 95% confidence interval (CI) of the dispersion...

  16. β-endorphin antigen

    International Nuclear Information System (INIS)

    1981-01-01

    This invention relates to the production of antigens comprising β-endorphin, βsub(h)-endorphin, or βsub(c)-endorphin, in covalent conjugation with human gammaglobulin as immunogenic carrier material, and an antibody having the property of specifically binding β-endorphin or fragments thereof, containing the (6-15) residue sequence. (U.K.)

  17. An in-depth analysis of original antigenic sin in dengue virus infection.

    Science.gov (United States)

    Midgley, Claire M; Bajwa-Joseph, Martha; Vasanawathana, Sirijitt; Limpitikul, Wannee; Wills, Bridget; Flanagan, Aleksandra; Waiyaiya, Emily; Tran, Hai Bac; Cowper, Alison E; Chotiyarnwong, Pojchong; Chotiyarnwon, Pojchong; Grimes, Jonathan M; Yoksan, Sutee; Malasit, Prida; Simmons, Cameron P; Mongkolsapaya, Juthathip; Screaton, Gavin R

    2011-01-01

    The evolution of dengue viruses has resulted in four antigenically similar yet distinct serotypes. Infection with one serotype likely elicits lifelong immunity to that serotype, but generally not against the other three. Secondary or sequential infections are common, as multiple viral serotypes frequently cocirculate. Dengue infection, although frequently mild, can lead to dengue hemorrhagic fever (DHF) which can be life threatening. DHF is more common in secondary dengue infections, implying a role for the adaptive immune response in the disease. There is currently much effort toward the design and implementation of a dengue vaccine but these efforts are made more difficult by the challenge of inducing durable neutralizing immunity to all four viruses. Domain 3 of the dengue virus envelope protein (ED3) has been suggested as one such candidate because it contains neutralizing epitopes and it was originally thought that relatively few cross-reactive antibodies are directed to this domain. In this study, we performed a detailed analysis of the anti-ED3 response in a cohort of patients suffering either primary or secondary dengue infections. The results show dramatic evidence of original antigenic sin in secondary infections both in terms of binding and enhancement activity. This has important implications for dengue vaccine design because heterologous boosting is likely to maintain the immunological footprint of the first vaccination. On the basis of these findings, we propose a simple in vitro enzyme-linked immunosorbent assay (ELISA) to diagnose the original dengue infection in secondary dengue cases.

  18. Mini-review: Strategies for Variation and Evolution of Bacterial Antigens

    Science.gov (United States)

    Foley, Janet

    2015-01-01

    Across the eubacteria, antigenic variation has emerged as a strategy to evade host immunity. However, phenotypic variation in some of these antigens also allows the bacteria to exploit variable host niches as well. The specific mechanisms are not shared-derived characters although there is considerable convergent evolution and numerous commonalities reflecting considerations of natural selection and biochemical restraints. Unlike in viruses, mechanisms of antigenic variation in most bacteria involve larger DNA movement such as gene conversion or DNA rearrangement, although some antigens vary due to point mutations or modified transcriptional regulation. The convergent evolution that promotes antigenic variation integrates various evolutionary forces: these include mutations underlying variant production; drift which could remove alleles especially early in infection or during life history phases in arthropod vectors (when the bacterial population size goes through a bottleneck); selection not only for any particular variant but also for the mechanism for the production of variants (i.e., selection for mutability); and overcoming negative selection against variant production. This review highlights the complexities of drivers of antigenic variation, in particular extending evaluation beyond the commonly cited theory of immune evasion. A deeper understanding of the diversity of purpose and mechanisms of antigenic variation in bacteria will contribute to greater insight into bacterial pathogenesis, ecology and coevolution with hosts. PMID:26288700

  19. Increased levels of IgA antibodies against CRA and FRA recombinant antigens of Trypanosoma cruzi differentiate digestive forms of Chagas disease.

    Science.gov (United States)

    Vasconcelos, Romero H T; Amaral, Fábio N; Cavalcanti, Maria G A M; Silva, Edimilson D; Ferreira, Antonio G P; Morais, Clarice N L; Gomes, Yara M

    2010-10-01

    In the chronic phase of Chagas disease, individuals infected by Trypanosoma cruzi may be asymptomatic or may present cardiac and/or digestive complications. Our aim here was to analyze the relationship between the presence of specific immunoglobulin A antibodies and the different chronic clinical forms of Chagas disease using two recombinant antigens of Trypanosoma cruzi, cytoplasmatic repetitive antigen and flagellar repetitive antigen. The association of this immunoglobulin isotype with the digestive and cardio-digestive forms of the disease determined by indirect enzyme-linked immunosorbent assay, strongly suggests that IgA antibodies against these recombinant antigens of T. cruzi can be used as an immunological marker of the digestive alterations caused by Chagas disease. The tests performed in this study show that it is possible to differentiate digestive forms of Chagas disease. The knowledge provided by these results may help physicians to manage early alterations in the digestive tract of patients with the indeterminate or cardiac forms of Chagas disease. Prospective studies, however, with follow-up of the patients that presenting with high levels of immunoglobulin A against cytoplasmatic repetitive antigen and flagellar repetitive antigen recombinant antigens, need to be conducted to confirm this hypothesis. 2010 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

  20. Long-term Prostate-specific Antigen Velocity in Improved Classification of Prostate Cancer Risk and Mortality

    DEFF Research Database (Denmark)

    Ørsted, David Dynnes; Bojesen, Stig E; Kamstrup, Pia R

    2013-01-01

    BACKGROUND: It remains unclear whether adding long-term prostate-specific antigen velocity (PSAV) to baseline PSA values improves classification of prostate cancer (PCa) risk and mortality in the general population. OBJECTIVE: To determine whether long-term PSAV improves classification of PCa risk...

  1. Diagnosis of Fasciola gigantica infection in cattle using capture-ELISA assay for detecting antigen in faeces

    Directory of Open Access Journals (Sweden)

    Sarwitri Endah Estuningsih

    2006-10-01

    Full Text Available Capture-ELISA assay is a diagnosis for antigen detection in the serum or faeces using polyclonal or monoclonal antibodies. The purpose of this study was to determine the sensitivity and specificity of capture-ELISA assay using polyclonal antibody for diagnosing Fasciola gigantica infection in cattle by detecting antigen in the faeces. In this study, faecal samples and livers were collected from 141 cattle slaughtered in the abattoir in Jakarta. From each animal, liver was processed for liver flukes count and the corresponding faecal sample was analysed for coproantigen. The result of capture-ELISA assay for antigen detection showed that from 85 cattle infected with Fasciola gigantica, 83 had OD > 0.52 (range from 0.52-1.39 and 2 cattle had OD < 0.52 (range from 0.17-0.51. The sensitivity and specificity of the assay were 97.6% and 92.8% respectively. The assay also able to detect 50 ng/ml of antigen in faecal supernatant. It suggests that this assay will have the advantage over the other methods on its ability to detect the active infection. Collection of faeces, rather than serum, will allow a more cost-effective and adaptable method.

  2. Antigen processing of glycoconjugate vaccines; the polysaccharide portion of the pneumococcal CRM(197) conjugate vaccine co-localizes with MHC II on the antigen processing cell surface.

    Science.gov (United States)

    Lai, Zengzu; Schreiber, John R

    2009-05-21

    Pneumococcal (Pn) polysaccharides (PS) are T-independent (TI) antigens and do not induce immunological memory or antibodies in infants. Conjugation of PnPS to the carrier protein CRM(197) induces PS-specific antibody in infants, and memory similar to T-dependent (Td) antigens. Conjugates have improved immunogenicity via antigen processing and presentation of carrier protein with MHC II and recruitment of T cell help, but the fate of the PS attached to the carrier is unknown. To determine the location of the PS component of PnPS-CRM(197) in the APC, we separately labeled PS and protein and tracked their location. The PS of types 14-CRM(197) and 19F-CRM(197) was specifically labeled by Alexa Fluor 594 hydrazide (red). The CRM(197) was separately labeled red in a reaction that did not label PS. Labeled antigens were incubated with APC which were fixed, permeabilized and incubated with anti-MHC II antibody labeled green by Alexa Fluor 488, followed by confocal microscopy. Labeled CRM(197) was presented on APC surface and co-localized with MHC II (yellow). Labeled unconjugated 14 or 19F PS did not go to the APC surface, but PS labeled 14-CRM(197) and 19F-CRM(197) was internalized and co-localized with MHC II. Monoclonal antibody to type 14 PS bound to intracellular type 14 PS and PS-CRM(197). Brefeldin A and chloroquine blocked both CRM(197) and PS labeled 14-CRM(197) and 19F-CRM(197) from co-localizing with MHC II. These data suggest that the PS component of the CRM(197) glycoconjugate enters the endosome, travels with CRM(197) peptides to the APC surface and co-localizes with MHC II.

  3. Isocyanate test antigens

    International Nuclear Information System (INIS)

    Karol, M.H.; Alarie, Y.C.

    1980-01-01

    A test antigen for detecting antibodies to a diisocyanate comprises the reaction product of a protein and a monoisocyanate derived from the same radical as the diisocyanate. The diisocyanates most usually encountered and therefore calling for antibody detection are those of toluene, hexamethylene, methylene, isophorone and naphthylene. The preferred protein is human serum albumin. (author)

  4. An approach to the unification of suppressor T cell circuits: a simplified assay for the induction of suppression by T cell-derived, antigen-binding molecules (T-ABM).

    Science.gov (United States)

    Chue, B; Ferguson, T A; Beaman, K D; Rosenman, S J; Cone, R E; Flood, P M; Green, D R

    1989-01-01

    A system is presented in which the in vitro response to sheep red blood cells (SRBC) can be regulated using antigenic determinants coupled to SRBC and T cell-derived antigen-binding molecules (T-ABM) directed against the coupled determinants. T suppressor-inducer factors (TsiF's) are composed of two molecules, one of which is a T-ABM and one which bears I-J determinants (I-J+ molecule). Using two purified T-ABM which have not previously been shown to have in vitro activity, we produced antigen-specific TsiF's which were capable of inducing the suppression of the anti-SRBC response. Suppression was found to require both the T-ABM and the I-J+ molecule, SRBC conjugated with the antigen for which the T-ABM was specific, and a population of Ly-2+ T cells in the culture. Two monoclonal TsiF (or TsF1) were demonstrated to induce suppression of the anti-SRBC response in this system, provided the relevant antigen was coupled to the SRBC in culture. The results are discussed in terms of the general functions of T-ABM in the immune system. This model will be useful in direct, experimental comparisons of the function of T-ABM and suppressor T cell factors under study in different systems and laboratories.

  5. Structural Basis for Antigenic Peptide Recognition and Processing by Endoplasmic Reticulum (ER) Aminopeptidase 2.

    Science.gov (United States)

    Mpakali, Anastasia; Giastas, Petros; Mathioudakis, Nikolas; Mavridis, Irene M; Saridakis, Emmanuel; Stratikos, Efstratios

    2015-10-23

    Endoplasmic reticulum (ER) aminopeptidases process antigenic peptide precursors to generate epitopes for presentation by MHC class I molecules and help shape the antigenic peptide repertoire and cytotoxic T-cell responses. To perform this function, ER aminopeptidases have to recognize and process a vast variety of peptide sequences. To understand how these enzymes recognize substrates, we determined crystal structures of ER aminopeptidase 2 (ERAP2) in complex with a substrate analogue and a peptidic product to 2.5 and 2.7 Å, respectively, and compared them to the apo-form structure determined to 3.0 Å. The peptides were found within the internal cavity of the enzyme with no direct access to the outside solvent. The substrate analogue extends away from the catalytic center toward the distal end of the internal cavity, making interactions with several shallow pockets along the path. A similar configuration was evident for the peptidic product, although decreasing electron density toward its C terminus indicated progressive disorder. Enzymatic analysis confirmed that visualized interactions can either positively or negatively impact in vitro trimming rates. Opportunistic side-chain interactions and lack of deep specificity pockets support a limited-selectivity model for antigenic peptide processing by ERAP2. In contrast to proposed models for the homologous ERAP1, no specific recognition of the peptide C terminus by ERAP2 was evident, consistent with functional differences in length selection and self-activation between these two enzymes. Our results suggest that ERAP2 selects substrates by sequestering them in its internal cavity and allowing opportunistic interactions to determine trimming rates, thus combining substrate permissiveness with sequence bias. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. High affinity antigen recognition of the dual specific variants of herceptin is entropy-driven in spite of structural plasticity.

    Directory of Open Access Journals (Sweden)

    Jenny Bostrom

    Full Text Available The antigen-binding site of Herceptin, an anti-human Epidermal Growth Factor Receptor 2 (HER2 antibody, was engineered to add a second specificity toward Vascular Endothelial Growth Factor (VEGF to create a high affinity two-in-one antibody bH1. Crystal structures of bH1 in complex with either antigen showed that, in comparison to Herceptin, this antibody exhibited greater conformational variability, also called "structural plasticity". Here, we analyzed the biophysical and thermodynamic properties of the dual specific variants of Herceptin to understand how a single antibody binds two unrelated protein antigens. We showed that while bH1 and the affinity-improved bH1-44, in particular, maintained many properties of Herceptin including binding affinity, kinetics and the use of residues for antigen recognition, they differed in the binding thermodynamics. The interactions of bH1 and its variants with both antigens were characterized by large favorable entropy changes whereas the Herceptin/HER2 interaction involved a large favorable enthalpy change. By dissecting the total entropy change and the energy barrier for dual interaction, we determined that the significant structural plasticity of the bH1 antibodies demanded by the dual specificity did not translate into the expected increase of entropic penalty relative to Herceptin. Clearly, dual antigen recognition of the Herceptin variants involves divergent antibody conformations of nearly equivalent energetic states. Hence, increasing the structural plasticity of an antigen-binding site without increasing the entropic cost may play a role for antibodies to evolve multi-specificity. Our report represents the first comprehensive biophysical analysis of a high affinity dual specific antibody binding two unrelated protein antigens, furthering our understanding of the thermodynamics that drive the vast antigen recognition capacity of the antibody repertoire.

  7. Major role for carbohydrate epitopes preferentially recognized by chronically infected mice in the determination of Schistosoma mansoni schistosomulum surface antigenicity

    International Nuclear Information System (INIS)

    Omer-ali, P.; Magee, A.I.; Kelly, C.; Simpson, A.J.G.

    1986-01-01

    A radioimmunoassay that makes use of whole Schistosomula and 125 I-labeled protein A has been used to characterize and to quantify the binding of antisera to the surface of 3 hr mechanically transformed schistosomula of Schistosoma mansoni. This technique facilitates the determination of epitopes on the schistosomula in addition to those detected by surface labeling and immunoprecipitation. By using this technique, it has been demonstrated that there is a much greater binding to the parasite surface of antibodies from chronically infected mice (CMS) than of antibodies from mice infected with highly irradiated cercariae (VMS), and CMS recognizes epitopes that VMS does not. Treatment of the surface of the schistosomula with trifluoromethanesulphonic acid and sodium metaperiodate has suggested that the discrepancy of the binding between the two sera is due to the recognition of a large number of additional epitopes by CMS, which are carbohydrate in nature. Some of the carbohydrate epitopes are expressed on the previously described surface glycoprotein antigens of M/sub r/ 200,000, 38,000, and 17,000

  8. Vaccination and the TAP-independent antigen processing pathways.

    Science.gov (United States)

    López, Daniel; Lorente, Elena; Barriga, Alejandro; Johnstone, Carolina; Mir, Carmen

    2013-09-01

    The cytotoxic CD8(+) T lymphocyte-mediated cellular response is important for the elimination of virus-infected cells and requires the prior recognition of short viral peptide antigens previously translocated to the endoplasmic reticulum by the transporter associated with antigen processing (TAP). However, individuals with nonfunctional TAP complexes or infected cells with TAP molecules blocked by specific viral proteins, such as the cowpoxvirus, a component of the first source of early empirical vaccination against smallpox, are still able to present several HLA class I ligands generated by the TAP-independent antigen processing pathways to specific cytotoxic CD8(+) T lymphocytes. Currently, bioterrorism and emerging infectious diseases have renewed interest in poxviruses. Recent works that have identified HLA class I ligands and epitopes in virus-infected TAP-deficient cells have implications for the study of both the effectiveness of early empirical vaccination and the analysis of HLA class I antigen processing in TAP-deficient subjects.

  9. ONCOLYTIC VIRUS-MEDIATED REVERSAL OF IMPAIRED TUMOR ANTIGEN PRESENTATION

    Directory of Open Access Journals (Sweden)

    Shashi Ashok Gujar

    2014-04-01

    Full Text Available Anti-tumor immunity can eliminate existing cancer cells and also maintain a constant surveillance against possible relapse. Such an antigen-specific adaptive response begins when tumor-specific T cells become activated. T cell activation requires two signals on antigen presenting cells (APCs: antigen presentation through MHC molecules and co-stimulation. In the absence of one or both of these signals, T cells remain inactivated or can even become tolerized. Cancer cells and their associated microenvironment strategically hinder the processing and presentation of tumor antigens and consequently prevent the development of anti-tumor immunity. Many studies, however, demonstrate that interventions that overturn tumor-associated immune evasion mechanisms can establish anti-tumor immune responses of therapeutic potential. One such intervention is oncolytic virus (OV-based anti-cancer therapy. Here we discuss how OV-induced immunological events override tumor-associated antigen presentation impairment and promote appropriate T cell:APC interaction. Detailed understanding of this phenomenon is pivotal for devising the strategies that will enhance the efficacy of OV-based anti-cancer therapy by complementing its inherent oncolytic

  10. I-125 input into antibodies molecules specific to australian antigen

    International Nuclear Information System (INIS)

    Abdukayumov, A. M.; Chistyakov, P.G.; Garajshina, G. R.

    1999-01-01

    There are experimental data on I-125 input into antibodies molecules specific to superficial antigen of hepatitis B virus (australian antigen). Three ways of input are submitted: with the help of T chloramine usage, Bolton-Hunter Reagent and with the help of iodogen. There are also comparative characteristics of iodized products obtained: molar radioactivity, radiochemical frequency, immuno - reactivity. The report also discusses advantages and disadvantages of the used methods for inputting I-125 into antibodies to australian antigen in order to study the possibility of creating radio immunological test system for detecting superficial antigen of B hepatitis

  11. Effects of roasting, blanching, autoclaving, and microwave heating on antigenicity of almond (Prunus dulcis L.) proteins.

    Science.gov (United States)

    Venkatachalam, M; Teuber, S S; Roux, K H; Sathe, S K

    2002-06-05

    Whole, unprocessed Nonpareil almonds were subjected to a variety of heat processing methods that included roasting (280, 300, and 320 degrees F for 20 and 30 min each; and 335 and 350 degrees F for 8, 10, and 12 min each), autoclaving (121 degrees C, 15 psi, for 5, 10, 15, 20, 25, and 30 min), blanching (100 degrees C for 1, 2, 3, 4, 5, and 10 min), and microwave heating (1, 2, and 3 min). Proteins were extracted from defatted almond flour in borate saline buffer, and immunoreactivity of the soluble proteins (normalized to 1 mg protein/mL for all samples) was determined using enzyme linked immunosorbent assay (ELISA). Antigenic stability of the almond major protein (amandin) in the heat-processed samples was determined by competitive inhibition ELISA using rabbit polyclonal antibodies raised against amandin. Processed samples were also assessed for heat stability of total antigenic proteins by sandwich ELISA using goat and rabbit polyclonal antibodies raised against unprocessed Nonpareil almond total protein extract. ELISA assays and Western blotting experiments that used both rabbit polyclonal antibodies and human IgE from pooled sera indicated antigenic stability of almond proteins when compared with that of the unprocessed counterpart.

  12. Cysteine proteases as potential antigens in antiparasitic DNA vaccines

    DEFF Research Database (Denmark)

    Jørgensen, Louise von Gersdorff; Buchmann, Kurt

    2011-01-01

    En litteraturgennemgang af muligheder for at bruge cystein proteaser som antigener i antiparasitære vacciner.......En litteraturgennemgang af muligheder for at bruge cystein proteaser som antigener i antiparasitære vacciner....

  13. Chagas disease-specific antigens: characterization of epitopes in CRA/FRA by synthetic peptide mapping and evaluation by ELISA-peptide assay.

    Science.gov (United States)

    Bottino, Carolina G; Gomes, Luciano P; Pereira, José B; Coura, José R; Provance, David William; De-Simone, Salvatore G

    2013-12-03

    The identification of epitopes in proteins recognized by medically relevant antibodies is useful for the development of peptide-based diagnostics and vaccines. In this study, epitopes in the cytoplasmic repetitive antigen (CRA) and flagellar repetitive antigen (FRA) proteins from Trypanosoma cruzi were identified using synthetic peptide techniques and pooled sera from Chagasic patients. The epitopes were further assayed with an ELISA assay based on synthetic peptides. Twenty-two overlapping synthetic peptides representing the coding sequence of the T. cruzi CRA and FRA proteins were assessed by a Spot-synthesis array analysis using sera donated by patients with Chagas disease. Shorter peptides were selected that represented the determined epitopes and synthesized by solid phase synthesis to evaluate the patterns of cross-reactivities and discrimination through an ELISA-diagnostic assay. The peptide Spot-synthesis array successfully identified two IgG antigenic determinants in the CRA protein and four in FRA. Bioinformatics suggested that the CRA antigens were unique to T. cruzi while the FRA antigen showed similarity with sequences present within various proteins from Leishmania sp. Subsequently, shorter peptides representing the CRA-1, CRA-2 and FRA-1 epitopes were synthesized by solid phase synthesis and assayed by an ELISA-diagnostic assay. The CRA antigens gave a high discrimination between Chagasic, Leishmaniasis and T. cruzi-uninfected serum. A sensitivity and specificity of 100% was calculated for CRA. While the FRA antigen showed a slightly lower sensitivity (91.6%), its specificity was only 60%. The epitopes recognized by human anti-T. cruzi antibodies have been precisely located in two biomarkers of T. cruzi, CRA and FRA. The results from screening a panel of patient sera through an ELISA assay based on peptides representing these epitopes strongly suggest that the sequences from CRA would be useful for the development of diagnostic reagents that could

  14. Binding of monoclonal antibody to protein antigen in fluid phase or bound to solid supports

    Energy Technology Data Exchange (ETDEWEB)

    Kennel, S J

    1982-01-01

    Rat monoclonal antibody (MoAb) to fragment D (FgD) of human fibrinogen was used to characterize the direct binding of antibody to protein in solution or bound to solid supports. Purified IgG, F(ab')/sub 2/ and Fab' were prepared from ascites fluid of hybridoma 104-14B which is a fusion product of spleen cells from a rat immunized with FgD and the mouse myeloma cell line, P3-X63-Ag8. Two-dimensional electrophoresis of radioiodinated antibody preparations demonstrated the presence of hybrid immunoglobulin molecules, but only structures having rat heavy and rat light chains had active antibody combinig sites. The affinity constant for IgG as well as F(ab')/sub 2/ and Fab', 6x10/sup 9/ M/sup -1/, was identical when tested using fluid phase antigen (/sup 125/I-labeled FgD). Affinity constants determined for direct binding of iodinated IgG using FgD immobilized on solid supports showed a slight dependence on the antigen concentration used in the measurement. These values ranged from 0.5x10/sup 9/ M/sup -1/ at high antigen concentrations (1.3x10/sup -7/ M) to 9x10/sup 9/ M/sup -1/ at low antigen concentration (1.3x10/sup -10/ M). Binding constants for F(ab')/sub 2/ and Fab' gave similar results indicating that binding was homogeneous and univalent. The capacity of solid state antigen to bind antibody varied with the method used to bind FgD to the solid support. FgD bound directly to polystyrene plates was least efficient at binding labeled antibody; FgD bound to plates through intermediate carriers poly(L-lysine) was only slightly more efficient, while antigen bound to Sepharose beads by cyanogen bromide activation was the most active.

  15. Insect-cell expression, crystallization and X-ray data collection of the bradyzoite-specific antigen BSR4 from Toxoplasma gondii

    International Nuclear Information System (INIS)

    Grujic, Ognjen; Grigg, Michael E.; Boulanger, Martin J.

    2008-01-01

    Preliminary X-ray diffraction studies of the bradyzoite-specific surface antigen BSR4 from T. gondii are described. Toxoplasma gondii is an important global pathogen that infects nearly one third of the world’s adult population. A family of developmentally expressed structurally related surface-glycoprotein adhesins (SRSs) mediate attachment to and are utilized for entry into host cells. The latent bradyzoite form of T. gondii persists for the life of the host and expresses a distinct family of SRS proteins, of which the bradyzoite-specific antigen BSR4 is a prototypical member. Structural studies of BSR4 were initiated by first recombinantly expressing BSR4 in insect cells, which was followed by crystallization and preliminary X-ray data collection to 1.95 Å resolution. Data processing showed that BSR4 crystallized with one molecule in the asymmetric unit of the P4 1 2 1 2 or P4 3 2 1 2 space group, with a solvent content of 60% and a corresponding Matthews coefficient of 2.98 Å 3 Da −1

  16. PCR Assays for Identification of Coccidioides posadasii Based on the Nucleotide Sequence of the Antigen 2/Proline-Rich Antigen

    Science.gov (United States)

    Bialek, Ralf; Kern, Jan; Herrmann, Tanja; Tijerina, Rolando; Ceceñas, Luis; Reischl, Udo; González, Gloria M.

    2004-01-01

    A conventional nested PCR and a real-time LightCycler PCR assay for detection of Coccidioides posadasii DNA were designed and tested in 120 clinical strains. These had been isolated from 114 patients within 10 years in Monterrey, Nuevo Leon, Mexico, known to be endemic for coccidioidomycosis. The gene encoding the specific antigen 2/proline-rich antigen (Ag2/PRA) was used as a target. All strains were correctly identified, whereas DNA from related members of the family Onygenaceae remained negative. Melting curve analysis by LightCycler and sequencing of the 526-bp product of the first PCR demonstrated either 100% identity to the GenBank sequence of the Silveira strain, now known to be C. posadasii (accession number AF013256), or a single silent mutation at position 1228. Length determination of two microsatellite-containing loci (GAC and 621) identified all 120 isolates as C. posadasii. Specific DNA was amplified by conventional nested PCR from three microscopically spherule-positive paraffin-embedded tissue samples, whereas 20 human tissue samples positive for other dimorphic fungi remained negative. Additionally, the safety of each step of a modified commercially available DNA extraction procedure was evaluated by using 10 strains. At least three steps of the protocol were demonstrated to sufficiently kill arthroconidia. This safe procedure is applicable to cultures and to clinical specimens. PMID:14766853

  17. Correlation of hepatospleno-scintigraphic findings with HBe antigenicity in chronic hepatitis B

    Energy Technology Data Exchange (ETDEWEB)

    Song, K S; Chun, K S; Chung, S K; Bahk, Y W [Catholic Medical College, Seoul (Korea, Republic of)

    1983-12-15

    Radioimmunoassay plays an important role in diagnosing the hepatitis B and in clinical assessment of the course of the disease as well. Among a number of antigens, antibodies and enzymes related with hepatitis, HBe Ag, DNA polymerase, IgM-HBc-antibody, {delta} Ag have been known as useful indicators of ongoing infectivity of hepatitis B. The present study has been undertaken to correlate the HBe antigenicity with hepatospleno-scintigraphic findings in hepatitis. The study covered a 10 month period from September 1982 through to July 1983. We reviewed and analyzed the hepatospeno-scintgraphic findings and the results of radioimmuassays in 32 patients of chronic hepatitis seen at St. Mary Hospital, Catholic Medical College. Hepatitis B was diagnosed either when HBs Ag was positive or Anti-HBc wa positive even if HBs Ag was negative. We classified the HBe antigenicity into two groups of HBe Ag(+) and HBe(-) and analyzed the scintgraphic findings in terms of liver size, motting, splenomegaly and splenic shift. From the present study, it is concluded that the activity of hepatitis B can not be assessed by the findings of hepatospleno-scintigram so far as the activity is determined on the basis of the positive HBe Ag, which has been believed to indicated continuing viral replication.

  18. Correlation of hepatospleno-scintigraphic findings with HBe antigenicity in chronic hepatitis B

    International Nuclear Information System (INIS)

    Song, K. S.; Chun, K. S.; Chung, S. K.; Bahk, Y. W.

    1983-01-01

    Radioimmunoassay plays an important role in diagnosing the hepatitis B and in clinical assessment of the course of the disease as well. Among a number of antigens, antibodies and enzymes related with hepatitis, HBe Ag, DNA polymerase, IgM-HBc-antibody, δ Ag have been known as useful indicators of ongoing infectivity of hepatitis B. The present study has been undertaken to correlate the HBe antigenicity with hepatospleno-scintigraphic findings in hepatitis. The study covered a 10 month period from September 1982 through to July 1983. We reviewed and analyzed the hepatospeno-scintgraphic findings and the results of radioimmuassays in 32 patients of chronic hepatitis seen at St. Mary Hospital, Catholic Medical College. Hepatitis B was diagnosed either when HBs Ag was positive or Anti-HBc wa positive even if HBs Ag was negative. We classified the HBe antigenicity into two groups of HBe Ag(+) and HBe(-) and analyzed the scintgraphic findings in terms of liver size, motting, splenomegaly and splenic shift. From the present study, it is concluded that the activity of hepatitis B can not be assessed by the findings of hepatospleno-scintigram so far as the activity is determined on the basis of the positive HBe Ag, which has been believed to indicated continuing viral replication

  19. MYCN: from oncoprotein to tumor-associated antigen

    International Nuclear Information System (INIS)

    Pistoia, Vito; Morandi, Fabio; Pezzolo, Annalisa; Raffaghello, Lizzia; Prigione, Ignazia

    2012-01-01

    MYCN is a well-known oncogene over-expressed in different human malignancies including neuroblastoma (NB), rhabdomyosarcoma, medulloblastoma, astrocytoma, Wilms’ tumor, and small cell lung cancer. In the case of NB, MYCN amplification is an established biomarker of poor-prognosis. MYCN belongs to a family of transcription factors (the most important of which is C-MYC) that show a high degree of homology. Down-regulation of MYC protein expression leads to tumor regression in animal models, indicating that MYC proteins represent interesting therapeutic targets. Pre-requisites for a candidate tumor-associated antigen (TAA) to be targeted by immunotherapeutic approaches are the following, (i) expression should be tumor-restricted, (ii) the putative TAA should be up-regulated in cancer cells, and (iii) protein should be processed into immunogenic peptides capable of associating to major histocompatibility complex molecules with high affinity. Indeed, the MYCN protein is not expressed in human adult tissues and up-regulated variably in NB cells, and MYCN peptides capable of associating to HLA-A1 or HLA-A2 molecules with high affinity have been identified. Thus the MYCN protein qualifies as putative TAA in NB. Additional issues that determine the feasibility of targeting a putative TAA with cytotoxic T lymphocytes (CTLs) and will be here discussed are the following, (i) the inadequacy of tumor cells per se to act as antigen-presenting cells witnessed, in the case of NB cells, by the low to absent expression of HLA class I molecules, the lack of co-stimulatory molecules and multiple defects in the HLA class I related antigen processing machinery, and (ii) the immune evasion mechanisms operated by cancer cells to fool the host immune system, such as up-regulation of soluble immunosuppressive molecules (e.g., soluble MICA and HLA-G in the case of NB) or generation of immunosuppressive cells in the tumor microenvironment. A final issue that deserves consideration is the

  20. MYCN: From Oncoprotein To Tumor-Associated Antigen

    Directory of Open Access Journals (Sweden)

    Vito ePistoia

    2012-11-01

    Full Text Available MYCN is a well known oncogene overexpressed in different human malignancies including neuroblastoma, rhabdomyosarcoma, medulloblastoma, astrocytoma, Wilms’ tumor and small cell lung cancer. In the case of neuroblastoma (NB, MYCN amplification is an established biomarker of poor prognosis. MYCN belongs to a family of transcription factors (the most important of which is CMYC that show a high degree of homology. Downregulation of MYC protein expression leads to tumor regression in animal models, indicating that MYC proteins represent interesting therapeutic targets.Pre-requisites for a candidate tumor-associated antigen (TAA to be targeted by immunotherapeutic approaches are the following, i expression should be tumor-restricted, ii the putative TAA should be up-regulated in cancer cells and iii protein should be processed into immunogenic peptides capable of associating to MHC molecules with high affinity. Indeed, the MYCN protein is not expressed in human adult tissues and upregulated variably in NB cells, and MYCN peptides capable of associating to HLA-A1 or –A2 molecules with high affinity have been identified. Thus the MYCN protein qualifies as putative TAA in NB.Additional issues that determine the feasibility of targeting a putative TAA with cytotoxic T lymphocytes (CTL and will be here discussed are the following, i the inadequacy of tumor cells per se to act as antigen-presenting cells witnessed, in the case of NB cells, by the low to absent expression of HLA- class I molecules, the lack of costimulatory molecules and multiple defects in the HLA class I related antigen processing machinery, and ii the immune evasion mechanisms operated by cancer cells to fool the host immune system, such as up-regulation of soluble immunosuppressive molecules (e.g. soluble MICA and HLA-G in the case of NB or generation of immunosuppressive cells in the tumor microenvironment. A final issue that deserves consideration is the strategy used to generate

  1. MYCN: from oncoprotein to tumor-associated antigen

    Energy Technology Data Exchange (ETDEWEB)

    Pistoia, Vito; Morandi, Fabio; Pezzolo, Annalisa; Raffaghello, Lizzia; Prigione, Ignazia, E-mail: vitopistoia@ospedale-gaslini.ge.it [Laboratory of Oncology, Translational Research and Laboratory Medicine, G. Gaslini Institute, Genoa (Italy)

    2012-11-16

    MYCN is a well-known oncogene over-expressed in different human malignancies including neuroblastoma (NB), rhabdomyosarcoma, medulloblastoma, astrocytoma, Wilms’ tumor, and small cell lung cancer. In the case of NB, MYCN amplification is an established biomarker of poor-prognosis. MYCN belongs to a family of transcription factors (the most important of which is C-MYC) that show a high degree of homology. Down-regulation of MYC protein expression leads to tumor regression in animal models, indicating that MYC proteins represent interesting therapeutic targets. Pre-requisites for a candidate tumor-associated antigen (TAA) to be targeted by immunotherapeutic approaches are the following, (i) expression should be tumor-restricted, (ii) the putative TAA should be up-regulated in cancer cells, and (iii) protein should be processed into immunogenic peptides capable of associating to major histocompatibility complex molecules with high affinity. Indeed, the MYCN protein is not expressed in human adult tissues and up-regulated variably in NB cells, and MYCN peptides capable of associating to HLA-A1 or HLA-A2 molecules with high affinity have been identified. Thus the MYCN protein qualifies as putative TAA in NB. Additional issues that determine the feasibility of targeting a putative TAA with cytotoxic T lymphocytes (CTLs) and will be here discussed are the following, (i) the inadequacy of tumor cells per se to act as antigen-presenting cells witnessed, in the case of NB cells, by the low to absent expression of HLA class I molecules, the lack of co-stimulatory molecules and multiple defects in the HLA class I related antigen processing machinery, and (ii) the immune evasion mechanisms operated by cancer cells to fool the host immune system, such as up-regulation of soluble immunosuppressive molecules (e.g., soluble MICA and HLA-G in the case of NB) or generation of immunosuppressive cells in the tumor microenvironment. A final issue that deserves consideration is the

  2. Rational design of protamine nanocapsules as antigen delivery carriers.

    Science.gov (United States)

    González-Aramundiz, José Vicente; Presas, Elena; Dalmau-Mena, Inmaculada; Martínez-Pulgarín, Susana; Alonso, Covadonga; Escribano, José M; Alonso, María J; Csaba, Noemi Stefánia

    2017-01-10

    Current challenges in global immunization indicate the demand for new delivery strategies, which could be applied to the development of new vaccines against emerging diseases, as well as to improve safety and efficacy of currently existing vaccine formulations. Here, we report a novel antigen nanocarrier consisting of an oily core and a protamine shell, further stabilized with pegylated surfactants. These nanocarriers, named protamine nanocapsules, were rationally designed to promote the intracellular delivery of antigens to immunocompetent cells and to trigger an efficient and long-lasting immune response. Protamine nanocapsules have nanometric size, positive zeta potential and high association capacity for H1N1 influenza hemagglutinin, a protein that was used here as a model antigen. The new formulation shows an attractive stability profile both, as an aqueous suspension or a freeze-dried powder formulation. In vitro studies showed that protamine nanocapsules were efficiently internalized by macrophages without eliciting significant toxicity. In vivo studies indicate that antigen-loaded nanocapsules trigger immune responses comparable to those achieved with alum, even when using significantly lower antigen doses, thus indicating their adjuvant properties. These promising in vivo data, alongside with their versatility for the loading of different antigens and oily immunomodulators and their excellent stability profile, make these nanocapsules a promising platform for the delivery of antigens. Protamine sulphate (PubChem SID: 7849283), Sodium Cholate (PubChem CID: 23668194), Miglyol (PubChem CID: 53471835), α tocopherol (PubChem CID: 14985), Tween® 20(PubChem CID: 443314), Tween® 80(PubChem CID: 5281955), TPGS (PubChem CID: 71406). Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Screening of random peptide library of hemagglutinin from pandemic 2009 A(H1N1 influenza virus reveals unexpected antigenically important regions.

    Directory of Open Access Journals (Sweden)

    Wanghui Xu

    Full Text Available The antigenic structure of the membrane protein hemagglutinin (HA from the 2009 A(H1N1 influenza virus was dissected with a high-throughput screening method using complex antisera. The approach involves generating yeast cell libraries displaying a pool of random peptides of controllable lengths on the cell surface, followed by one round of fluorescence-activated cell sorting (FACS against antisera from mouse, goat and human, respectively. The amino acid residue frequency appearing in the antigenic peptides at both the primary sequence and structural level was determined and used to identify "hot spots" or antigenically important regions. Unexpectedly, different antigenic structures were seen for different antisera. Moreover, five antigenic regions were identified, of which all but one are located in the conserved HA stem region that is responsible for membrane fusion. Our findings are corroborated by several recent studies on cross-neutralizing H1 subtype antibodies that recognize the HA stem region. The antigenic peptides identified may provide clues for creating peptide vaccines with better accessibility to memory B cells and better induction of cross-neutralizing antibodies than the whole HA protein. The scheme used in this study enables a direct mapping of the antigenic regions of viral proteins recognized by antisera, and may be useful for dissecting the antigenic structures of other viral proteins.

  4. Expression of the Gastrin-Releasing Peptide Receptor, the Prostate Stem Cell Antigen and the Prostate-Specific Membrane Antigen in Lymph Node and Bone Metastases of Prostate Cancer

    NARCIS (Netherlands)

    Ananias, Hildo J. K.; van den Heuvel, Marius C.; Helfrich, Wijnand; de Jong, Igle J.

    2009-01-01

    OBJECTIVE. Cell membrane antigens like the gastrin-releasing peptide receptor (GRPR), the prostate stem cell antigen (PSCA), and the prostate-specific membrane antigen (PSMA), expressed in prostate cancer, are attractive targets for new therapeutic and diagnostic applications. Therefore, we

  5. The potential for induction of autoimmune disease by a randomly-mutated self-antigen

    DEFF Research Database (Denmark)

    Pedersen, Anders Elm

    2007-01-01

    -antigens can be immunogenic and lead to autoimmunity against wildtype self-antigens. In theory, modified self-antigens can arise by random errors and mutations during protein synthesis and would be recognized as foreign antigens by naïve B and T lymphocytes. Here, it is postulated that the initial auto......, a relation to an infectious disease is described, and it is thought that microbes can play a direct role in induction of autoimmunity, for instance by molecular mimicry or bystander activation of autoreactive T cells. In contrast, less attention has been given to the possibility that modified self......-antigen is not a germline self-antigen, but rather a mutated self-antigen. This mutated self-antigen might interfere with peripheral tolerance if presented to the immune system during an infection. The infection lead to bystander activation of naïve T and B cells with specificity for mutated self-antigen and this can lead...

  6. Helminth antigens counteract a rapid high-fat diet-induced decrease in adipose tissue eosinophils.

    Science.gov (United States)

    van den Berg, Susan M; van Dam, Andrea D; Kusters, Pascal J H; Beckers, Linda; den Toom, Myrthe; van der Velden, Saskia; Van den Bossche, Jan; van Die, Irma; Boon, Mariëtte R; Rensen, Patrick C N; Lutgens, Esther; de Winther, Menno P J

    2017-10-01

    Brown adipose tissue (BAT) activation and white adipose tissue (WAT) beiging can increase energy expenditure and have the potential to reduce obesity and associated diseases. The immune system is a potential target in mediating brown and beige adipocyte activation. Type 2 and anti-inflammatory immune cells contribute to metabolic homeostasis within lean WAT, with a prominent role for eosinophils and interleukin (IL)-4-induced anti-inflammatory macrophages. We determined eosinophil numbers in epididymal WAT (EpAT), subcutaneous WAT (ScAT) and BAT after 1 day, 3 days or 1 week of high-fat diet (HFD) feeding in C57Bl/6 mice. One day of HFD resulted in a rapid drop in eosinophil numbers in EpAT and BAT, and after 3 days, in ScAT. In an attempt to restore this HFD-induced drop in adipose tissue eosinophils, we treated 1-week HFD-fed mice with helminth antigens from Schistosoma mansoni or Trichuris suis and evaluated whether the well-known protective metabolic effects of helminth antigens involves BAT activation or beiging. Indeed, antigens of both helminth species induced high numbers of eosinophils in EpAT, but failed to induce beiging. In ScAT, Schistosoma mansoni antigens induced mild eosinophilia, which was accompanied by slightly more beiging. No effects were observed in BAT. To study type 2 responses on brown adipocytes directly, T37i cells were stimulated with IL-4. This increased Ucp1 expression and strongly induced the production of eosinophil chemoattractant CCL11 (+26-fold), revealing that brown adipocytes themselves can attract eosinophils. Our findings indicate that helminth antigen-induced eosinophilia fails to induce profound beiging of white adipocytes. © 2017 Society for Endocrinology.

  7. Quantitative radioimmunoassay for membranous and soluble H-Y antigen typing

    Energy Technology Data Exchange (ETDEWEB)

    Casanova-Bettane, M.; Latron, F.; Jakob, H.; Fellous, M.

    1981-01-01

    Two sensitive and quantitative methods for membranous or soluble H-Y antigen typing using rat anti-H-Y immune sera and /sup 125/I labelled protein A were carried out. These techniques were used to study H-Y antigen expression in human cell lines, and to refine the hypothesis that ..beta../sub 2/m serves as an anchorage point for H-Y antigen.

  8. Efficient Capsid Antigen Presentation From Adeno-Associated Virus Empty Virions In Vivo.

    Science.gov (United States)

    Pei, Xiaolei; Earley, Lauriel Freya; He, Yi; Chen, Xiaojing; Hall, Nikita Elexa; Samulski, Richard Jude; Li, Chengwen

    2018-01-01

    Adeno-associated virus (AAV) vectors have been successfully applied in clinical trials for hemophilic patients. Although promising, the clinical results suggest that the capsid-specific CD8+T cell response has a negative effect on therapeutic success. In an in vitro analysis using an engineered AAV virus carrying immune-dominant SIINFEKL peptide in the capsid backbone, we have previously demonstrated that capsid antigen presentation from full (genome containing) AAV capsids requires endosome escape and is proteasome dependent and that no capsid antigen presentation is induced from empty virions. In the present study, we examined capsid antigen presentation from administration of empty virions in animal models. In wild-type mice, similar to AAV full particles, capsid antigen presentation from AAV empty virion infection was dose dependent, and the kinetics studies showed that antigen presentation was detected from 2 to 40 days after AAV empty virion administration. In the transporter associated with antigen processing 1 deficient (TAP-/-) mice, capsid antigen presentation was inhibited from both AAV full and empty virions, but higher inhibition was achieved from AAV full particle administration than that from empty virions. This indicates that the pathway of capsid antigen presentation from AAV transduction is dependent on proteasome-mediated degradation of AAV capsids (mainly for full particles) and that the endosomal pathway may also play a role in antigen presentation from empty particles but not full virions. The capsid antigen presentation efficiency from AAV preparations was positively correlated with the amount of empty virions contaminated with full particles. Collectively, the results indicate that contamination of AAV empty virions induces efficient antigen presentation in vivo and the mechanism of capsid antigen presentation from empty virions involves both endosomal and proteasomal pathways. The elucidation of capsid antigen presentation from AAV empty

  9. Genetic distinctions between autoimmune hepatitis in Italy and North America.

    Science.gov (United States)

    Muratori, Paolo; Czaja, Albert-J; Muratori, Luigi; Pappas, Georgios; Maccariello, Silvana; Cassani, Fabio; Granito, Alessandro; Ferrari, Rodolfo; Mantovani, Vilma; Lenzi, Marco; Bianchi, Francesco-B

    2005-03-28

    Our goals were to analyze the known genetic predispositions for autoimmune hepatitis (AIH) in AIH Italian population and to compare them with North American counterparts. Human leukocyte antigens (HLA) B8, C7, DR3, DR4, DR7, DR11, DR13, DQ2 and the B8-DR3-DQ2 phenotype were determined by microlymphocytotoxicity and polymerase chain reaction in 74 Italian patients (57 with type 1 and 17 with type 2 AIH) and 149 North American patients with type 1 AIH, and in adequate controls. B8-DR3-DQ2 occurred more frequently in Italian patients with type 1 AIH than in Italian controls (30% vs 7%, P<0.0001), but less frequently than in North American counterparts (30% vs 48%, P = 0.02). DR4 occurred less frequently in Italian patients with type 1 AIH (23% vs 43%, P = 0.01) and in controls (16% vs 34%, P = 0.0003) than in North American counterparts. No differences were found in alleles' frequency between type 1 and type 2 Italian AIH patients. DR11 had a frequency lower in type 1 Italian AIH patients than controls (17% vs 35%, P = 0.01). HLA DR4 is not associated with AIH in Italy. The known HLA risk factors for AIH occur similarly in Italian patients with type 1 and type 2 AIH, and they are less frequent than in North American patients. B8-DR3-DQ2 is the predominant phenotype of type 1 AIH also in Italy, and HLA DR11 may be a regionally distinctive protective factor against type 1 AIH.

  10. Shedding of leukemia-associated P24 antigen by lymphoblastoid cell lines.

    Science.gov (United States)

    Komada, Y; Ochiai, H; Shimizu, K; Azuma, E; Kamiya, H; Sakurai, M

    1987-12-01

    We report the development of a unique enzyme-linked immunosorbent assay (ELISA) which makes possible the detection of leukemia-associated P24 antigen, utilizing its ability to bind the Ricinus communis agglutinin (RCA1) and a monoclonal antibody, SJ-9A4 simultaneously. Using the RCA1/SJ-9A4-ELISA, P24 antigen, as few as 50 X 10(3) cells from a common acute lymphoblastic leukemia (C-ALL) cell line could be detected. The presence of D-galactose gave complete and specific inhibition of P24 antigen binding to RCA1. Matched concentrations of D-glucose and D-sucrose had no effect on binding. The release of the P24 antigen into the culture medium by a C-ALL cell line maintained at 37 degrees C could be detected; however, no P24 antigen was present in the culture medium when the cells were maintained at 4 degrees C. Sequential analysis of the culture medium for soluble P24 antigen revealed that release of the P24 antigen associated with cell growth. Molecular sieve chromatography of concentrated culture medium indicated that shed P24 antigen was eluted in the macromolecule fraction. P24 antigen was detected in the cerebrospinal fluid (CSF) of four patients with P24 positive ALL at the time of relapse of the central nervous system (CNS) and was undetectable while in complete remission. The CSF from three patients with P24 negative ALL and three patients with aseptic meningitis had no detectable activity.

  11. Selection of antigenic markers on a GFP-C{kappa} fusion scaffold with high sensitivity by eukaryotic ribosome display

    Energy Technology Data Exchange (ETDEWEB)

    Yongmin, Yang [Institute of Genetics, San Diego, CA 92121-2233 (United States); IgE Therapeutics, Inc., San Diego, CA 92121-2233 (United States); Barankiewicz, Teresa J [Institute of Genetics, San Diego, CA 92121-2233 (United States); IgE Therapeutics, Inc., San Diego, CA 92121-2233 (United States); Mingyue, He [Babraham Institute, Cambridge CB2 4AT (United Kingdom); Taussig, Michael J [Babraham Institute, Cambridge CB2 4AT (United Kingdom); Chen, Swey-Shen [Institute of Genetics, San Diego, CA 92121-2233 (United States) and IgE Therapeutics, Inc., San Diego, CA 92121-2233 (United States)

    2007-07-27

    Ribosome display is a cell-free system permitting gene selection through the physical association of genetic material (mRNA) and its phenotypic (protein) product. While often used to select single-chain antibodies from large libraries by panning against immobilized antigens, we have adapted ribosome display for use in the 'reverse' format in order to select high affinity antigenic determinants against solid-phase antibody. To create an antigenic scaffold, DNA encoding green fluorescent protein (GFP) was fused to a light chain constant domain (C{kappa}) with stop codon deleted, and with 5' signals (T7 promoter, Kozak) enabling coupled transcription/translation in a eukaryotic cell-free system. Epitopes on either GFP (5') or C{kappa} (3') were selected by anti-GFP or anti-C{kappa} antibodies, respectively, coupled to magnetic beads. After selection, mRNA was amplified directly from protein-ribosome-mRNA (PRM) complexes by in situ PCR followed by internal amplification and reassembly PCR. As little as 10 fg of the 1 kb DNA construct, i.e. approximately 7500 molecules, could be recovered following a single round of interaction with solid-phase anti-GFP antibody. This platform is highly specific and sensitive for the antigen-antibody interaction and may permit selection and reshaping of high affinity antigenic variants of scaffold proteins.

  12. Expression of cathepsins B, L, S, and D by gastric epithelial cells implicates them as antigen presenting cells in local immune responses.

    Science.gov (United States)

    Barrera, C; Ye, G; Espejo, R; Gunasena, S; Almanza, R; Leary, J; Crowe, S; Ernst, P; Reyes, V E

    2001-10-01

    Helicobacter pylori infection is linked to chronic gastritis, peptic ulcer and gastric carcinoma. During H. pylori infection, class II MHC expression by the gastric epithelium increases, as does the number of local CD4(+) T cells, which appear to be important in the associated pathogenesis. These observations suggested that the epithelium might present antigens to T cells. Thus, we sought to determine whether gastric epithelial cells process antigens to establish their function as local antigen presenting cells (APC). We examined a panel of gastric epithelial cell lines for expression of the antigen processing cathepsins B (CB), L (CL), S (CS), and D (CD). The mRNA for these enzymes were detected by RT-PCR and the enzymes in the gastric epithelial cells were identified by various independent methods. We corroborated the expression of CB and CD on gastric epithelial cells from human biopsy samples. The functions of these proteases were confirmed by assessing their ability to digest ovalbumin, a conventional dietary antigen, and proteins from H. pylori. In summary, multiple lines of evidence suggest gastric epithelial cells process antigens for presentation to CD4(+) T cells. To our knowledge, these are the first studies to document the antigen processing capacity of human gastric epithelial cells.

  13. Development of recombinant antigen array for simultaneous detection of viral antibodies.

    Directory of Open Access Journals (Sweden)

    Yi Liu

    Full Text Available Protein microarrays have been developed to study antibody reactivity against a large number of antigens, demonstrating extensive perspective for clinical application. We developed a viral antigen array by spotting four recombinant antigens and synthetic peptide, including glycoprotein G of herpes simplex virus (HSV type 1 and 2, phosphoprotein 150 of cytomegalovirus (CMV, Rubella virus (RV core plus glycoprotein E1 and E2 as well as a E1 peptide with the optimal concentrations on activated glass slides to simultaneously detect IgG and IgM against HSV1, HSV2, CMV and RV in clinical specimens of sera and cerebrospinal fluids (CSFs. The positive reference sera were initially used to measure the sensitivity and specificity of the array with the optimal conditions. Then clinical specimens of 144 sera and 93 CSFs were tested for IgG and IgM antibodies directed against HSV1, HSV2, CMV and RV by the antigen array. Specificity of the antigen array for viral antibodies detection was satisfying compared to commercial ELISA kits but sensitivity of the array varied relying on quality and antigenic epitopes of the spotting antigens. In short, the recombinant antigen array has potential to simultaneous detect multiple viral antibodies using minute amount (3 µl of samples, which holds the particularly advantage to detect viral antibodies in clinical CSFs being suspicious of neonatal meningitis and encephalitis.

  14. 76 FR 58867 - Endangered and Threatened Species; Determination of Nine Distinct Population Segments of...

    Science.gov (United States)

    2011-09-22

    ... Northern District of California modified the February 19, 2010, deadline to March 8, 2010. On March 16... markedly separated from other populations of the same taxon (an organism or group of organisms) as a... to identify two genetically distinct nesting populations in the Pacific--a northern hemisphere...

  15. Prostate-specific antigen lowering effect of metabolic syndrome is influenced by prostate volume.

    Science.gov (United States)

    Choi, Woo Suk; Heo, Nam Ju; Paick, Jae-Seung; Son, Hwancheol

    2016-04-01

    To investigate the influence of metabolic syndrome on prostate-specific antigen levels by considering prostate volume and plasma volume. We retrospectively analyzed 4111 men who underwent routine check-ups including prostate-specific antigen and transrectal ultrasonography. The definition of metabolic syndrome was based on the modified Adult Treatment Panel III criteria. Prostate-specific antigen mass density (prostate-specific antigen × plasma volume / prostate volume) was calculated for adjusting plasma volume and prostate volume. We compared prostate-specific antigen and prostate-specific antigen mass density levels of participants with metabolic syndrome (metabolic syndrome group, n = 1242) and without metabolic syndrome (non-prostate-specific antigen metabolic syndrome group, n = 2869). To evaluate the impact of metabolic syndrome on prostate-specific antigen, linear regression analysis for the natural logarithm of prostate-specific antigen was used. Patients in the metabolic syndrome group had significantly older age (P prostate volume (P prostate-specific antigen (non-metabolic syndrome group vs metabolic syndrome group; 1.22 ± 0.91 vs 1.15 ± 0.76 ng/mL, P = 0.006). Prostate-specific antigen mass density in the metabolic syndrome group was still significantly lower than that in the metabolic syndrome group (0.124 ± 0.084 vs 0.115 ± 0.071 μg/mL, P = 0.001). After adjusting for age, prostate volume and plasma volume using linear regression model, the presence of metabolic syndrome was a significant independent factor for lower prostate-specific antigen (prostate-specific antigen decrease by 4.1%, P = 0.046). Prostate-specific antigen levels in patients with metabolic syndrome seem to be lower, and this finding might be affected by the prostate volume. © 2016 The Japanese Urological Association.

  16. Mendelian and non-mendelian mutations affecting surface antigen expression in Paramecium tetraurelia

    International Nuclear Information System (INIS)

    Epstein, L.M.; Forney, J.D.

    1984-01-01

    A screening procedure was devised for the isolation of X-ray-induced mutations affecting the expression of the A immobilization antigen (i-antigen) in Paramecium tetraurelia. Two of the mutations isolated by this procedure proved to be in modifier genes. The two genes are unlinked to each other and unlinked to the structural A i-antigen gene. These are the first modifier genes identified in a Paramecium sp. that affect surface antigen expression. Another mutation was found to be a deletion of sequences just downstream from the A i-antigen gene. In cells carrying this mutation, the A i-antigen gene lies in close proximity to the end of a macronuclear chromosome. The expression of the A i-antigen is not affected in these cells, demonstrating that downstream sequences are not important for the regulation and expression of the A i-antigen gene. A stable cell line was also recovered which shows non-Mendelian inheritance of a macronuclear deletion of the A i-antigen gene. This mutant does not contain the gene in its macronucleus, but contains a complete copy of the gene in its micronucleus. In the cytoplasm of wild-type animals, the micronuclear gene is included in the developing macronucleus; in the cytoplasm of the mutant, the incorporation of the A i-antigen gene into the macronucleus is inhibited. This is the first evidence that a mechanism is available in ciliates to control the expression of a gene by regulating its incorporation into developing macronuclei

  17. Prevalence of Diego blood group antigen and the antibody in three ethnic population groups in Klang valley of Malaysia

    OpenAIRE

    Cheong Tar Wei; Faisal Muti Al-Hassan; Norris Naim; Aishah Knight; Sanmukh R Joshi

    2013-01-01

    Background: Diego blood group antigen, Di(a), is very rare among Caucasians and Blacks, but relatively common among the South American Indians and Asians of Mongolian origin. The antibody to Di(a) is clinically significant to cause hemolytic disease in a new-born or hemolytic transfusion reaction. Objectives: This study was designed to determine the prevalence of Di(a) antigen among the blood donors from the three major ethnic groups in Klang Valley of Malaysia as well as to find an incidence...

  18. Role of antigen in migration patterns of T cell subsets arising from gut-associated lymphoid tissue

    International Nuclear Information System (INIS)

    Dunkley, M.L.; Husband, A.J.

    1989-01-01

    Studies of the migration of antigen-specific regulatory T cell subsets responding to gut immunization were undertaken to clarify their migratory potential and the role of antigen in their localization. In initial experiments, lymphocytes collected from the thoracic duct of rats after immunization of Peyer's patches (PP) with keyhole limpet hemocyanin (KLH), were enriched for T helper (Th) cells and labelled with the fluorochrome H33342. In other experiments, a higher frequency of antigen-specific T cells was achieved by short-term culture of the enriched Th cells in the presence of KLH and the blast cells labelled with 3H-thymidine. The distribution of both populations was determined after injection into immunized and unimmunized syngeneic recipients. Whereas the uncultured population (predominantly small Th cells) localized almost exclusively in follicular lymphoid tissues, the cells expanded by secondary culture (predominantly Th blasts) appeared in the gut lamina propria (LP) initially, then in PP and mesenteric lymph nodes. The Th blasts in the LP were almost always seen in close proximity to the gut epithelium. However, the migration of neither population appeared to be influenced significantly by antigen, in contrast to previous findings with regard to IgA-committed B cells. The initial subepithelial location of Th blasts in the gut LP and their subsequent appearance in PP may provide a mechanism by which antigen presented by epithelial cells could influence B cell differentiation in PP through modulation of signals expressed by these T cells

  19. Role of antigen in migration patterns of T cell subsets arising from gut-associated lymphoid tissue

    Energy Technology Data Exchange (ETDEWEB)

    Dunkley, M.L.; Husband, A.J. (Univ. of Newcastle, N.S.W. (Australia))

    1989-07-01

    Studies of the migration of antigen-specific regulatory T cell subsets responding to gut immunization were undertaken to clarify their migratory potential and the role of antigen in their localization. In initial experiments, lymphocytes collected from the thoracic duct of rats after immunization of Peyer's patches (PP) with keyhole limpet hemocyanin (KLH), were enriched for T helper (Th) cells and labelled with the fluorochrome H33342. In other experiments, a higher frequency of antigen-specific T cells was achieved by short-term culture of the enriched Th cells in the presence of KLH and the blast cells labelled with 3H-thymidine. The distribution of both populations was determined after injection into immunized and unimmunized syngeneic recipients. Whereas the uncultured population (predominantly small Th cells) localized almost exclusively in follicular lymphoid tissues, the cells expanded by secondary culture (predominantly Th blasts) appeared in the gut lamina propria (LP) initially, then in PP and mesenteric lymph nodes. The Th blasts in the LP were almost always seen in close proximity to the gut epithelium. However, the migration of neither population appeared to be influenced significantly by antigen, in contrast to previous findings with regard to IgA-committed B cells. The initial subepithelial location of Th blasts in the gut LP and their subsequent appearance in PP may provide a mechanism by which antigen presented by epithelial cells could influence B cell differentiation in PP through modulation of signals expressed by these T cells.

  20. Identification and characterization of surface antigens in parasites, using radiolabelling techniques

    International Nuclear Information System (INIS)

    Ramasamy, R.

    1982-04-01

    Surface proteins of Schistosoma sp and Leishmania sp were studied using 125-Iodine as tracer. The surface proteins were labelled by the Lactoperoxidase method and the proteins then separated using SDS PAG electrophoresis and autoradiography. The possible immunogens were then separated using immunoprecipitation and Fluorescent Antibody techniques using sera from patients or from artificially immunized rabbits. Four common antigens were identified from the surfaces of male and female adult worms, cercariae and schistosomulae of S.mansoni. These antigens, which had molecular weights of 150,000, 78,000, 45,000, and 22,000 were also isolated from the surfaces of S.haematobium adults. The surface antigens on promastigotes of a Kenyan strain of Leishmania donovani were separated into three protein antigens with molecular weights of 66,000, 59,000 and 43,000 respectively. The 59,000 molecular weight antigen was a glycoprotein and was common to promastigotes of an American and Indian strain of L.donovani and to L.braziliensis mexicana. None of the isolated antigens have been shown to have a protective effect when vaccinated into mice, but the study illustrates the value of radionuclide tracers in the unravelling of the mosaic of antigens which parasites possess