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Sample records for variant surface antigen

  1. Genomic environment of variant surface antigen genes of Trypanosoma equiperdum.

    Science.gov (United States)

    Raibaud, A; Gaillard, C; Longacre, S; Hibner, U; Buck, G; Bernardi, G; Eisen, H

    1983-01-01

    Expression of variant antigen genes in Trypanosoma equiperdum is accompanied by the duplication of a silent basic copy gene and the transposition of the copy to an expression site elsewhere in the genome. We have analyzed the genomic locations of both the basic and expression-linked copies of the T. equiperdum gene for variable surface glycoprotein VSG-1. Both copies are situated proximal to termini in both extracted DNA and in chromatin. The regions between the VSG-1 genes and the termini have a very high buoyant density in CsCl and contain an unidentified nucleoside that replaces deoxycytidine. Images PMID:6308614

  2. Genomic environment of variant surface antigen genes of Trypanosoma equiperdum.

    OpenAIRE

    Raibaud, A; Gaillard, C; Longacre, S; Hibner, U; Buck, G; Bernardi, G; Eisen, H

    1983-01-01

    Expression of variant antigen genes in Trypanosoma equiperdum is accompanied by the duplication of a silent basic copy gene and the transposition of the copy to an expression site elsewhere in the genome. We have analyzed the genomic locations of both the basic and expression-linked copies of the T. equiperdum gene for variable surface glycoprotein VSG-1. Both copies are situated proximal to termini in both extracted DNA and in chromatin. The regions between the VSG-1 genes and the termini ha...

  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...... (VSASM) that were better recognized by plasma IgG than VSA expressed by other parasites, but importantly, VSASM-type antigens also appeared to show substantial antigenic homogeneity. Our finding that the repertoire of immunologically distinct VSA in general, and in particular that of VSASM...

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

    DEFF Research Database (Denmark)

    Hviid, Lars

    2010-01-01

    that development of PfEMP1-based vaccines to protect specifically against severe malaria syndromes-in particular PAM-is feasible. This review summarizes the evidence that VSAs are important targets of NAI, discusses why VSA-based vaccines might be feasible despite the extensive intra- and interclonal variation...... 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.......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...

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

    DEFF Research Database (Denmark)

    Hviid, Lars

    2010-01-01

    that development of PfEMP1-based vaccines to protect specifically against severe malaria syndromes-in particular PAM-is feasible. This review summarizes the evidence that VSAs are important targets of NAI, discusses why VSA-based vaccines might be feasible despite the extensive intra- and interclonal variation...... 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.......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...

  6. Malaria-induced acquisition of antibodies to Plasmodium falciparum variant surface antigens

    DEFF Research Database (Denmark)

    Ofori, Michael F; Dodoo, Daniel; Staalsoe, Trine

    2002-01-01

    In areas of intense Plasmodium falciparum transmission, protective immunity is acquired during childhood in parallel with acquisition of agglutinating antibodies to parasite-encoded variant surface antigens (VSA) expressed on parasitized red blood cells. In a semi-immune child in such an area......, clinical disease is caused mainly by parasites expressing VSA not recognized by preexisting VSA-specific antibodies in that child. Such malaria episodes are known to cause an increase in agglutinating antibodies specifically recognizing VSA expressed by the parasite isolate causing the illness, whereas...

  7. Overlapping antigenic repertoires of variant antigens expressed on the surface of erythrocytes infected by Plasmodium falciparum

    DEFF Research Database (Denmark)

    Giha, H A; Staalsoe, T; Dodoo, D

    1999-01-01

    genes and undergoes clonal variation. Using agglutination and mixed agglutination tests and flow cytometry to analyse the recognition of variant antigens on parasitized erythrocytes by plasma antibodies from individuals living in Daraweesh in eastern Sudan, an area of seasonal and unstable malaria...

  8. In vivo switching between variant surface antigens in human Plasmodium falciparum infection

    DEFF Research Database (Denmark)

    Staalsoe, Trine; Hamad, Amel A; Hviid, Lars

    2002-01-01

    A semi-immune individual was retrospectively found to have maintained an apparently monoclonal and genotypically stable asymptomatic infection for months after clinical cure of a Plasmodium falciparum malaria episode. Before the attack, the individual had no antibodies to variant surface antigens...

  9. Plasmodium falciparum parasites expressing pregnancy-specific variant surface antigens adhere strongly to the choriocarcinoma cell line BeWo

    DEFF Research Database (Denmark)

    Haase, Rikke N; Megnekou, Rosette; Lundquist, Maja

    2006-01-01

    Placenta-sequestering Plasmodium falciparum parasites causing pregnancy-associated malaria express pregnancy-specific variant surface antigens (VSA(PAM)). We report here that VSA(PAM)-expressing patient isolates adhere strongly to the choriocarcinoma cell line BeWo and that the BeWo line can...

  10. The sickle cell trait is associated with enhanced immunoglobulin G antibody responses to Plasmodium falciparum variant surface antigens.

    NARCIS (Netherlands)

    Cabrera, G.; Cot, M.; Migot-Nabias, F.; Kremsner, P.G.; Deloron, P.; Luty, A.J.F.

    2005-01-01

    The sickle cell trait (HbAS) protects against severe Plasmodium falciparum malaria in young African children. We investigated the extent of the association between HbAS and antibodies directed to parasite-derived variant surface antigens (VSAs) on the membrane of infected erythrocytes. We measured

  11. Prevalence of naturally occurring surface gene variants of hepatitis B virus in nonimmunized surface antigen-negative Chinese carriers.

    Science.gov (United States)

    Hou, J; Wang, Z; Cheng, J; Lin, Y; Lau, G K; Sun, J; Zhou, F; Waters, J; Karayiannis, P; Luo, K

    2001-11-01

    Previous studies have suggested that hepatitis B virus (HBV) variants may account for the presence of HBV DNA in hepatitis B surface antigen (HBsAg)-negative patients (occult HBV infection). However, it is not known how widespread these variants are and how they influence the course of liver disease. To determine the prevalence of variants within the major hydrophilic region (MHR) of HBsAg, we investigated 2,565 subjects, including subjects with chronic hepatitis, cryptogenic cirrhosis, hemodialysis patients, and blood donors. Fifty-one of them had occult HBV infection. The entire S gene from 46 of these patients was sequenced from amplified serum HBV DNA. Forty-three percent (20 of 46) had mutations in the MHR of HBsAg. Thirty-two amino acid substitutions between positions 100-160 of the MHR of HBsAg were detected in 18 patients, and these ranged from 1 to 4 per patient. These changes involved 11 positions inside and 5 outside of the historical first and second loops of the "a" determinant, and included the following: Q101K, T115A, K122N, T123A, T126N, Q129N, G130R, T131I, M133T, F134L, C138Y, K141E, P142S, G145R, N146S, and C147F/R. Combinations of mutations were detected in 9 patients, and 7 of these have not been described before. Two further patients had insertion mutations immediately before the "a" determinant. Monoclonal antibody binding tests with the Royal Free hepatitis B surface (RFHBs) panel of antibodies revealed decreased immunoreactivity in 6 novel variants of HBsAg. The existence of patients with occult HBV infection caused by HBsAg variants, therefore, has implications for their possible transmission through sexual contact and by blood transfusion.

  12. Induction of endoplasmic reticulum-derived oxidative stress by an occult infection related S surface antigen variant.

    Science.gov (United States)

    Lee, In-Kyung; Lee, Seoung-Ae; Kim, Hong; Won, You-Sub; Kim, Bum-Joon

    2015-06-14

    To investigate the mechanism of endoplasmic reticulum (ER) stress induction by an occult infection related hepatitis B virus S surface antigen (HBsAg) variant. We used an HBsAg variant with lower secretion capacity, which was a KD variant from a Korean subject who was occultly infected with the genotype C. We compared the expression profiles of ER stress-related proteins between HuH-7 cells transfected with HBsAg plasmids of a wild-type and a KD variant using Western blot. Confocal microscopy indicated that the KD variant had higher levels of co-localization with ER than the wild-type HBsAg. The KD variant up-regulated ER stress-related proteins and induced reactive oxygen species (ROS) compared to the wild-type via an increase in calcium. The KD variant also down-regulated anti-oxidant proteins (HO-1, catalase and SOD) compared to the wild-type, which indicates positive amplification loops of the ER-ROS axis. The KD variant also induced apoptotic cell death via the up-regulation of caspase proteins (caspase 6, 9 and 12). Furthermore, the KD variant induced a higher level of nitric oxide than wild-type HBsAg via the up-regulation of the iNOS protein. Our data indicate that occult infection related HBsAg variants can lead to ER-derived oxidative stress and liver cell death in HuH-7 cells.

  13. Antigenic variation of Plasmodium knowlesi malaria: identification of the variant antigen on infected erythrocytes.

    OpenAIRE

    Howard, R J; Barnwell, J W; Kao, V

    1983-01-01

    Erythrocytes infected with mature asexual stages of Plasmodium knowlesi express a new surface antigen such that rhesus monkey antisera specifically agglutinate these cells. Cloned parasites can express different antigenic variants of this antigen. The variant antigen has been identified by comparison of the surface membrane antigens of a clone and of an antigenic variant of that clone of different agglutination phenotype. After lactoperoxidase labeling, 125I-labeled proteins of Mrs 210,000 an...

  14. Hepatitis B surface antigen variants in voluntary blood donors in Nanjing, China

    Directory of Open Access Journals (Sweden)

    Yong-lin Yang

    2012-04-01

    Full Text Available Abstract Background Hepatitis B virus (HBV is still one of the serious infectious risks for the blood transfusion safety in China. One plausible reason is the emergence of the variants in the major antigenic alpha determinant within the major hydrophilic region (MHR of hepatitis B surface antigen (HBsAg, which have been assumed to evade the immune surveillance and pose a challenge to the disease diagnosis. It is well documented that some commercial ELISA kits could detect the wild-type but not the mutant viruses. The high prevalence of HBV in China also impaired the application of nucleic acid testing (NAT in the improvement of blood security. Molecular epidemiological study of HBsAg variations in China is still limited. This study was designed to identify the prevalence of mutations in the HBsAg in voluntary blood donors in Nanjing, China. Methods A total of 20,326 blood units were enrolled in this study, 39 donors were positive for HBV S gene in the nested-PCR. Mutations in the major hydrophilic region (MHR; aa 99-169 were identified by direct sequencing of S region. Results Among of 20,326 blood units in the Red Cross Transfusion Center of Nanjing from October 2008 to April 2009, 296 samples (1.46%, 296/20,326 were HBsAg positive in the 2 successive rounds of the ELISA test. In these HBsAg positive units, HBV S gene could be successfully amplified from 39 donors (13.18%, 39/296 in the nested-PCR. Sequence analysis revealed that 32 strains (82.1%, 32/39 belong to genotype B, 7 strains (17.9%, 7/39 to genotype C. Besides well known G145R, widely dispersed variations in the MHR of S region, were observed in 20 samples of all the strains sequenced. Conclusions HBV/B and HBV/C are dominant in Nanjing, China. The mutations in the MHR of HBsAg associated with disease diagnosis are common.

  15. Nine-year longitudinal study of antibodies to variant antigens on the surface of Plasmodium falciparum-infected erythrocytes

    DEFF Research Database (Denmark)

    Giha, H A; Staalsoe, T; Dodoo, D

    1999-01-01

    PfEMP1 is an antigenically variable molecule which mediates the adhesion of parasitized erythrocytes to a variety of cell types and which is believed to constitute an important target for naturally acquired protective immune responses in malaria. For 9 years we have monitored individuals living...... in an area of low-intensity, seasonal, and unstable malaria transmission in eastern Sudan, and we have used this database to study the acquisition, specificity, and duration of the antibody response to variant parasitized erythrocyte surface antigens. Both the levels and the spectrum of reactivity...... of these antibodies varied considerably among individuals, ranging from low levels of antibodies recognizing only few parasitized erythrocyte surface antigens to high levels of broad-specificity antibodies. In general, episodes of clinical malaria were associated with increases in the levels of parasitized...

  16. Antibodies to variant antigens on the surfaces of infected erythrocytes are associated with protection from malaria in Ghanaian children

    DEFF Research Database (Denmark)

    Dodoo, D; Staalsoe, T; Giha, H

    2001-01-01

    Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is a variant antigen expressed on the surface of infected erythrocytes. Each parasite genome contains about 40 PfEMP1 genes, but only 1 PfEMP1 gene is expressed at a given time. PfEMP1 serves as a parasite-sequestering ligand...... to endothelial cells and enables the parasites to avoid splenic passage. PfEMP1 antibodies may protect from disease by inhibiting sequestration, thus facilitating the destruction of infected erythrocytes in the spleen. In this study, we have measured antibodies in Ghanaian children to a conserved region of PfEMP...... during the season. The prevalences of antibodies to both the conserved PfEMP1 peptide and the variant epitopes were greater than 50%, and the levels of immunoglobulin G (IgG) correlated with age. The levels of antibodies to both the conserved peptide and the variant epitopes were higher in protected than...

  17. Novel Plasmodium falciparum malaria vaccines: evidence-based searching for variant surface antigens as candidates for vaccination against pregnancy-associated malaria

    DEFF Research Database (Denmark)

    Staalsoe, Trine; Jensen, Anja T R; Theander, Thor G

    2002-01-01

    Malaria vaccine development has traditionally concentrated on careful molecular, biochemical, and immunological characterisation of candidate antigens. In contrast, evidence of the importance of identified antigens in immunity to human infection and disease has generally been limited......) in particular, to provide robust evidence of a causal link between the two in order to allow efficient and evidence-based identification of candidate antigens for malaria vaccine development....... to statistically significant co-variation with protection rather than on demonstration of causal relationships. We have studied the relationship between variant surface antigen-specific antibodies and clinical protection from Plasmodium falciparum malaria in general, and from pregnancy-associated malaria (PAM...

  18. Variant surface antigen-specific IgG and protection against clinical consequences of pregnancy-associated Plasmodium falciparum malaria

    DEFF Research Database (Denmark)

    Staalsoe, Trine; Shulman, Caroline E; Bulmer, Judith N

    2004-01-01

    BACKGROUND: Pregnancy-associated malaria caused by Plasmodium falciparum adherence to chondroitin sulfate A in the placental intervillous space is a major cause of low birthweight and maternal anaemia in areas of endemic P falciparum transmission. Adhesion-blocking antibodies that specifically...... recognise parasite-encoded variant surface antigens (VSA) are associated with resistance to pregnancy-associated malaria. We looked for a possible relation between VSA-specific antibody concentrations, placental infection, and protection from low birthweight and maternal anaemia. METHODS: We used flow...... selected or not selected for chondroitin sulfate A adhesiveness in-vitro. FINDINGS: Concentrations of plasma IgG specific for VSA, expressed by chondroitin sulfate A-adhering parasites (VSA in pregnancy-associated malaria or vsa-pam), increased with gravidity and were associated with placental histological...

  19. Determinants of variant surface antigen antibody response in severe Plasmodium falciparum malaria in an area of low and unstable malaria transmission

    DEFF Research Database (Denmark)

    A-Elgadir, T M E; Theander, T G; Elghazali, G

    2006-01-01

    The variant surface antigens (VSA) of infected erythrocytes are important pathogenic markers, a set of variants (VSA(SM)), were assumed to be associated with severe malaria (SM), while SM constitutes clinically diverse forms, such as, severe malarial anemia (SMA) and cerebral malaria (CM). This s......The variant surface antigens (VSA) of infected erythrocytes are important pathogenic markers, a set of variants (VSA(SM)), were assumed to be associated with severe malaria (SM), while SM constitutes clinically diverse forms, such as, severe malarial anemia (SMA) and cerebral malaria (CM...... range of isolates had a higher level of VSA Ab against the recognized isolates (correlation coefficient, 0.727, PSMA (P....001). Parasites obtained from patients with SMA or from children were better recognized than isolates obtained from patients with uncomplicated malaria or from adults, P

  20. Temporal expression and localization patterns of variant surface antigens in clinical Plasmodium falciparum isolates during erythrocyte schizogony.

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    Anna Bachmann

    Full Text Available Avoidance of antibody-mediated immune recognition allows parasites to establish chronic infections and enhances opportunities for transmission. The human malaria parasite Plasmodium falciparum possesses a number of multi-copy gene families, including var, rif, stevor and pfmc-2tm, which encode variant antigens believed to be expressed on the surfaces of infected erythrocytes. However, most studies of these antigens are based on in vitro analyses of culture-adapted isolates, most commonly the laboratory strain 3D7, and thus may not be representative of the unique challenges encountered by P. falciparum in the human host. To investigate the expression of the var, rif-A, rif-B, stevor and pfmc-2tm family genes under conditions that mimic more closely the natural course of infection, ex vivo clinical P. falciparum isolates were analyzed using a novel quantitative real-time PCR approach. Expression patterns in the clinical isolates at various time points during the first intraerythrocytic developmental cycle in vitro were compared to those of strain 3D7. In the clinical isolates, in contrast to strain 3D7, there was a peak of expression of the multi-copy gene families rif-A, stevor and pfmc-2tm at the young ring stage, in addition to the already known expression peak in trophozoites. Furthermore, most of the variant surface antigen families were overexpressed in the clinical isolates relative to 3D7, with the exception of the pfmc-2tm family, expression of which was higher in 3D7 parasites. Immunofluorescence analyses performed in parallel revealed two stage-dependent localization patterns of RIFIN, STEVOR and PfMC-2TM. Proteins were exported into the infected erythrocyte at the young trophozoite stage, whereas they remained inside the parasite membrane during schizont stage and were subsequently observed in different compartments in the merozoite. These results reveal a complex pattern of expression of P. falciparum multi-copy gene families during

  1. 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 blo...

  2. Analysis of IgG with specificity for variant surface antigens expressed by placental Plasmodium falciparum isolates

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    Kremsner Peter G

    2004-07-01

    Full Text Available Abstract Background Pregnancy-associated malaria (PAM is caused by Plasmodium falciparum-infected erythrocytes that can sequester in placental intervillous space by expressing particular variant surface antigens (VSA that can mediate adhesion to chondroitin sulfate A (CSA in vitro. IgG antibodies with specificity for the VSA expressed by these parasites (VSAPAM are associated with protection from maternal anaemia, prematurity and low birth weight, which is the greatest risk factor for death in the first month of life. Methods In this study, the development of anti-VSAPAM antibodies in a group of 151 women who presented to the maternity ward of Albert Schweitzer Hospital in Lambaréné, Gabon for delivery was analysed using flow cytometry assays. Plasma samples from placenta infected primiparous women were also investigated for their capacity to inhibit parasite binding to CSA in vitro. Results In the study cohort, primiparous as well as secundiparous women had the greatest risk of infection at delivery as well as during pregnancy. Primiparous women with infected placentas at delivery showed higher levels of VSAPAM-specific IgG compared to women who had no malaria infections at delivery. Placental isolates of Gabonese and Senegalese origin tested on plasma samples from Gabon showed parity dependency and gender specificity patterns. There was a significant correlation of plasma reactivity as measured by flow cytometry between different placental isolates. In the plasma of infected primiparous women, VSAPAM-specific IgG measured by flow cytometry could be correlated with anti-adhesion antibodies measured by the inhibition of CSA binding. Conclusion Recognition of placental parasites shows a parity- and sex- dependent pattern, like that previously observed in laboratory strains selected to bind to CSA. Placental infections at delivery in primiparous women appear to be sufficient to induce functional antibodies which can both recognize the surface of

  3. Insecticide-treated bed nets reduce plasma antibody levels and limit the repertoire of antibodies to Plasmodium falciparum variant surface antigens

    DEFF Research Database (Denmark)

    Askjaer, N; Maxwell, C; Chambo, W

    2001-01-01

    variant surface antigens (VSA) are important in the development of naturally acquired immunity to Plasmodium falciparum malaria and may thus be good indicators of immune status. We have compared the levels of VSA antibodies in plasma from children who have used ITN for 4 years to levels in plasma from...... children from a nearby village not using ITN. A total of 97 plasma samples were analyzed using 13 different P. falciparum isolates. We found that the children using ITN had significantly lower VSA antibody levels and recognized a smaller proportion of the VSA expressed by the tested parasite isolates than...... children not using ITN....

  4. Analysis of IgG with specificity for variant surface antigens expressed by placental Plasmodium falciparum isolates

    DEFF Research Database (Denmark)

    Khattab, Ayman; Reinhardt, Christina; Staalsoe, Trine

    2004-01-01

    BACKGROUND: Pregnancy-associated malaria (PAM) is caused by Plasmodium falciparum-infected erythrocytes that can sequester in placental intervillous space by expressing particular variant surface antigens (VSA) that can mediate adhesion to chondroitin sulfate A (CSA) in vitro. IgG antibodies......: In the study cohort, primiparous as well as secundiparous women had the greatest risk of infection at delivery as well as during pregnancy. Primiparous women with infected placentas at delivery showed higher levels of VSAPAM-specific IgG compared to women who had no malaria infections at delivery. Placental...

  5. Immune selection and within-host competition can structure the repertoire of variant surface antigens in Plasmodium falciparum -a mathematical model

    DEFF Research Database (Denmark)

    van Noort, Sander P; Nunes, Marta C; Weedall, Gareth D

    2010-01-01

    BACKGROUND: The evolutionary mechanisms structuring the expression pattern of variant surface antigen (VSA) families that allow pathogens to evade immune responses and establish chronic and repeated infections pose major challenges to theoretical research. In Plasmodium falciparum, the best......-studied VSA family is erythrocyte membrane protein 1 (PfEMP1). Each parasite genome encodes about 60 PfEMP1 variants, which are important virulence factors and major targets of host antibody responses. Transcriptional switching is the basis of clonal PfEMP1 variation and immune evasion. A relatively conserved......-host and diverse blocks that are favoured by immune selection at the population level. CONCLUSIONS/SIGNIFICANCE: The application of a monotonic dominance profile to VSAs encoded by a gene family generates two opposing selective forces and, consequently, two distinct clusters of genes emerge in adaptation to naïve...

  6. Plasmodium falciparum variant surface antigen expression varies between isolates causing severe and nonsevere malaria and is modified by acquired immunity

    DEFF Research Database (Denmark)

    Nielsen, Morten A; Staalsoe, Trine; Kurtzhals, Jørgen

    2002-01-01

    of immunity, as disease-causing parasites appear to be those not controlled by preexisting VSA-specific Abs. In this work we report that VSA expressed by parasites from young Ghanaian children with P. falciparum malaria were commonly and strongly recognized by plasma Abs from healthy children in the same area...... to limit the risk of severe disease by discriminating against the expression of VSA likely to cause life-threatening complications, such as cerebral malaria and severe anemia. Such VSA seem to be preferred by parasites infecting a nonimmune host, suggesting that VSA expression and switching are not random......In areas of endemic parasite transmission, protective immunity to Plasmodium falciparum malaria is acquired over several years with numerous disease episodes. Acquisition of Abs to parasite-encoded variant surface Ags (VSA) on the infected erythrocyte membrane is important in the development...

  7. Breaking tolerance in hepatitis B surface antigen (HBsAg) transgenic mice by vaccination with cross-reactive, natural HBsAg variants

    DEFF Research Database (Denmark)

    Schirmbeck, Reinhold; Dikopoulos, Nektarios; Kwissa, Marcin

    2003-01-01

    Processing exogenous hepatitis B surface antigen (HBsAg) of the hepatitis B virus (HBV) generates the K(b)-binding S(208-215) epitope 1; processing endogenous HBsAg generates the K(b)-binding S(190-197) epitope 2. Cross-reactive CD8(+) T cell responses were primed to epitope 1 but not epitope 2......-tg mice by HBsAg(adw2); these specific CD8(+) T cells cross-reacted with epitope 1 processed from the transgene-encoded HBsAg(ayw). The liver of vaccinated HBsAg(ayw) transgenic mice showed severe histopathology and contained functional (IFNgamma-producing), cross-reactive CD8(+) T cells, and vaccinated...... HBs-tg mice showed reduced antigenemia. Hence, vaccination with natural HBsAg variants from different HBV sero/genotypes can prime cross-reactive, specific CD8(+) T cell immunity that breaks tolerance to HBsAg....

  8. DNA rearrangements and antigenic variation in Trypanosoma equiperdum: expression-independent DNA rearrangements in the basic copy of a variant surface glycoprotein gene.

    Science.gov (United States)

    Longacre, S; Raibaud, A; Hibner, U; Buck, G; Eisen, H; Baltz, T; Giroud, C; Baltz, D

    1983-03-01

    Antigenic variation in Trypanosoma equiperdum is associated with the sequential expression of variant surface glycoprotein (VSG) genes in a process which involves gene duplication and transposition events. In this paper we present evidence that the genomic environment of the VSG-1 basic copy gene, the template for duplicated, expression-linked VSG-1 genes, differs in every trypanosome clone examined. This variation is thus independent of the expression of the VSG-1 gene, and it also appears to be restricted to the 3' genomic environment. It is also demonstrated that the DNA located 3' to the VSG-1 basic copy gene is moderately sensitive to digestion when the nuclei of either expressor or non-expressor trypanosomes are treated with DNase I.

  9. Serodiagnosis of bovine trypanosomosis caused by non-tsetse transmitted Trypanosoma (Duttonella) vivax parasites using the soluble form of a Trypanozoon variant surface glycoprotein antigen.

    Science.gov (United States)

    Uzcanga, Graciela L; Pérez-Rojas, Yenis; Camargo, Rocío; Izquier, Adriana; Noda, José A; Chacín, Ronny; Parra, Nereida; Ron, Lenin; Rodríguez-Hidalgo, Richar; Bubis, José

    2016-03-15

    Previous studies have shown that a 64-kDa antigen (p64) that was purified from the Venezuelan TeAp-N/D1 isolate of Trypanosoma (Trypanozoon) equiperdum corresponds to the soluble form of its predominant variant surface glycoprotein (VSG), and exhibited cross-reactivity with Trypanosoma (Duttonella) vivax. The course of experimental acute infections of bovines with T. vivax were followed by measuring whole anti-p64 antibodies and specific anti-p64 IgG and IgM antibodies in animal sera by indirect enzyme-linked immunosorbent assay (ELISA). The value of p64 to diagnose bovine trypanosomosis was also examined using 350 sera from healthy and T. vivax-infected cows living in a trypanosomosis-endemic and enzootic stable area, and 48 sera obtained during a trypanosomosis outbreak. Serological assays showed that ∼ 70-80% of the infected sera contained anti-p64 antibodies, based on the comparative immunodetection of the T. equiperdum clarified antigenic fraction used as a reference test. In the absence of a gold standard, Bayesian analysis for multiple testing estimated a sensitivity and specificity of 71.6% and 98.8%, respectively, for the indirect ELISA using p64 as antigen. An apparent prevalence of 37.7% for bovine trypanosomosis infection was also estimated with a Bayesian approach when the p64 ELISA test was used. Employing blood from acute infected cows, the indirect ELISA response against p64 was contrasted with the microhematocrit centrifuge method and analyses by polymerase chain reaction (PCR) using specific primers targeting the inter-specific length variation of the internal transcribed spacer 1 region of the 18S ribosomal gene. The efficiency of p64 for the detection of anti-trypanosome antibodies in acute infected bovines was also corroborated serologically by comparing its response to that of the Indonesian Trypanosoma evansi Rode Trypanozoon antigen type (RoTat) 1.2 VSG, which possesses high specificity and sensitivity. As expected, PCR was the best

  10. Modeling malaria infection and immunity against variant surface antigens in Príncipe island, West Africa

    DEFF Research Database (Denmark)

    Bandeiras, Cátia; Trovoada, Maria Jesus; Gonçalves, Lígia A

    2014-01-01

    After remarkable success of vector control campaigns worldwide, concerns about loss of immunity against Plasmodium falciparum due to lack of exposure to the parasite are relevant since an increase of severe cases in less immune individuals is expected. We present a mathematical model to investigate...... the impact of reducing exposure to the parasite on the immune repertoire against P. falciparum erythrocyte membrane protein 1 (PfEMP1) variants. The model was parameterized with data from Príncipe Island, West Africa, and applied to simulate two alternative transmission scenarios: one where control measures...... are continued to eventually drive the system to elimination; and another where the effort is interrupted after 6 years of its initiation and the system returns to the initial transmission potential. Population dynamics of parasite prevalence predict that in a few years infection levels return to the pre...

  11. Isolation and characterization of human rhinovirus antigenic variants

    Energy Technology Data Exchange (ETDEWEB)

    Watson, D.G.

    1985-01-01

    Isolation of antigenic variants of human rhinovirus types 2, 14, and 17 was attempted by plaquing untreated virus (P-isolates), selecting variants in the presence of homologous antiserum (C-isolates), and by selecting variants in the presence of antibody following 5-fluorouracil mutagenesis (M-isolates). All viruses were triple-plaque purified and purity neutralization tested prior to isolate selection. Based on a fourfold reduction in neutralizing antibody titer to homologous antiserum, no antigenic variation was found in P-isolates from the three serotypes examined. Antigenic variants of all three serotypes could be isolated by the antiserum selection method (C-isolates). However, antigenic variants of RV17 were isolated at a much higher frequency and showed a larger degree of variation than those of RV2 and RV14. At least two of the variants selected, RV17 (C301) and RV2 (M803), failed to be neutralized by the known 89 rhinovirus antiserum. SDS-polyacrylamide gel electrophoresis of (/sup 35/S) methionine-labelled virion polypeptides revealed that each serotype had a characteristic pattern and that selected RV2 and RV17 isolates had patterns identical to those of the prototype strains. By isoelectric focusing an antigenic variant of RV2 was shown to contain altered virion polypeptides VP1 and VP2 whereas two RV17 antigenic variants demonstrated alterations only in the VP1 polypeptide.

  12. Selection of antigenically advanced variants of seasonal influenza viruses

    Science.gov (United States)

    Ozawa, Makoto; Taft, Andrew S.; Das, Subash C.; Hanson, Anthony P.; Song, Jiasheng; Imai, Masaki; Wilker, Peter R.; Watanabe, Tokiko; Watanabe, Shinji; Ito, Mutsumi; Iwatsuki-Horimoto, Kiyoko; Russell, Colin A.; James, Sarah L.; Skepner, Eugene; Maher, Eileen A.; Neumann, Gabriele; Kelso, Anne; McCauley, John; Wang, Dayan; Shu, Yuelong; Odagiri, Takato; Tashiro, Masato; Xu, Xiyan; Wentworth, David E.; Katz, Jacqueline M.; Cox, Nancy J.; Smith, Derek J.; Kawaoka, Yoshihiro

    2016-01-01

    Influenza viruses mutate frequently, necessitating constant updates of vaccine viruses. To establish experimental approaches that may complement the current vaccine strain selection process, we selected antigenic variants from human H1N1 and H3N2 influenza virus libraries possessing random mutations in the globular head of the haemagglutinin protein (which includes the antigenic sites) by incubating them with human and/or ferret convalescent sera to human H1N1 and H3N2 viruses. Further, we selected antigenic escape variants from human viruses treated with convalescent sera and from mice that had been previously immunized against human influenza viruses. Our pilot studies with past influenza viruses identified escape mutants that were antigenically similar to variants that emerged in nature, establishing the feasibility of our approach. Our studies with contemporary human influenza viruses identified escape mutants before they caused an epidemic in 2014–2015. This approach may aid in the prediction of potential antigenic escape variants and the selection of future vaccine candidates before they become widespread in nature. PMID:27572841

  13. Selection of antigenically advanced variants of seasonal influenza viruses

    NARCIS (Netherlands)

    Li, C. (Chengjun); Hatta, M. (Masato); D.F. Burke (David); Ping, J. (Jihui); Zhang, Y. (Ying); Ozawa, M. (Makoto); Taft, A.S. (Andrew S.); Das, S.C. (Subash C.); Hanson, A.P. (Anthony P.); Song, J. (Jiasheng); M. Imai; Wilker, P.R. (Peter R.); Watanabe, T. (Tokiko); Watanabe, S. (Shinji); Ito, M. (Mutsumi); Iwatsuki-Horimoto, K. (Kiyoko); C.A. Russell (Colin); S.L. James (Sarah ); E. Skepner (Eugene); E. Maher (Eileen); G. Neumann (Gabriele); A. Klimov (Alexander); A. Kelso; McCauley, J. (John); D. Wang (Dayan); Y.L. Shu (Yue-Long); T. Odagiri (Takato); Tashiro, M. (Masato); X. Xu (Xiyan); Wentworth, D.E. (David E.); J. Katz (Jacqueline); N.J. Cox (Nancy); D.J. Smith (Derek James); Y. Kawaoka (Yoshihiro)

    2016-01-01

    textabstractInfluenza viruses mutate frequently, necessitating constant updates of vaccine viruses. To establish experimental approaches that may complement the current vaccine strain selection process, we selected antigenic variants from human H1N1 and H3N2 influenza virus libraries possessing

  14. Canine parvovirus: the worldwide occurrence of antigenic variants.

    Science.gov (United States)

    Miranda, Carla; Thompson, Gertrude

    2016-09-01

    The most important enteric virus infecting canids is canine parvovirus type 2 (CPV-2). CPV is the aetiologic agent of a contagious disease, mainly characterized by clinical gastroenteritis signs in younger dogs. CPV-2 emerged as a new virus in the late 1970s, which could infect domestic dogs, and became distributed in the global dog population within 2 years. A few years later, the virus's original type was replaced by a new genetic and antigenic variant, called CPV-2a. Around 1984 and 2000, virus variants with the single change to Asp or Glu in the VP2 residue 426 were detected (sometimes termed CPV-2b and -2c). The genetic and antigenic changes in the variants have also been correlated with changes in their host range; in particular, in the ability to replicate in cats and also host range differences in canine and other tissue culture cells. CPV-2 variants have been circulating among wild carnivores and have been well-documented in several countries around the world. Here, we have reviewed and summarized the current information about the worldwide distribution and evolution of CPV-2 variants since they emerged, as well as the host ranges they are associated with.

  15. Overexpression of squamous cell carcinoma antigen variants in hepatocellular carcinoma.

    Science.gov (United States)

    Pontisso, P; Calabrese, F; Benvegnù, L; Lise, M; Belluco, C; Ruvoletto, M G; Marino, M; Valente, M; Nitti, D; Gatta, A; Fassina, G

    2004-02-23

    Pathogenetic mechanisms of hepatocellular carcinoma (HCC) are still unclear and new tools for diagnostic and therapeutic purposes are ongoing. We have assessed whether squamous cell carcinoma antigen (SCCA), a serpin overexpressed in neoplastic cells of epithelial origin, is also expressed in liver cancer. Squamous cell carcinoma antigen was evaluated by immunohistochemistry in 65 HCCs of different aetiology and in 20 normal livers. Proliferative activity was assessed using MIB-1 antibody. In 18 surgical samples, tumour and nontumour liver tissue was available for SCCA cDNA amplification and sequencing. Squamous cell carcinoma antigen was detected in 55 out of 65 (85%) tumour specimens, but in none of the 20 controls. In the majority of the cases, the positive signal was found in the cytoplasm of more than 50% of the hepatocytes. Low or undetectable SCCA (scoreSCCA score >or=2 (mean+/-s.d.: 2%+/-2.4 vs 7.5%+/-10.3, PSCCA1 variant (G(351) to A) was identified in five cases, while SCCA1 was revealed in six cases and SCCA2 in three cases. In conclusion, SCCA variants are overexpressed in HCC, independently of tumour aetiology. A novel SCCA1 variant has been identified in one third of liver tumours.

  16. Differences in human antibody reactivity to Plasmodium falciparum variant surface antigens are dependent on age and malaria transmission intensity in northeastern Tanzania

    DEFF Research Database (Denmark)

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

    2008-01-01

    at population level, we conducted an immunoepidemiological study in nearby communities in northeastern Tanzania, situated at different altitudes and therefore exposed to different levels of P. falciparum transmission intensity. Samples of plasma and infected red blood cells (IRBC) were collected from 759......-VSA IgG response developed dramatically in individuals at 1 to 2 years of age in the high-transmission area, reaching a maximum level at around 10 years of age; only a modest further increase was observed among older children and adults. In contrast, at lower levels of malaria transmission, anti-VSA Ig...... and functional characteristics of the variant-specific antibody response, which is likely to be important for protection against malaria....

  17. 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...... with a first hydrophobic antigen to provide a first group of bead-antigen conjugates by adsorption of the first hydrophobic antigen on the first group of beads, iii) isolating said bead- antigen conjugates, iv) contacting said bead-antigen conjugates with a sample to bind antibodies therein to provide bead...

  18. A novel category of antigens enabling CTL immunity to tumor escape variants: Cinderella antigens.

    Science.gov (United States)

    Seidel, Ursula J E; Oliveira, Claudia C; Lampen, Margit H; Hall, Thorbald van

    2012-01-01

    Deficiencies in MHC class I antigen presentation are a common feature of tumors and allows escape from cytotoxic T lymphocyte (CTL)-mediated killing. It is crucial to take this capacity of tumors into account for the development of T-cell-based immunotherapy, as it may strongly impair their effectiveness. A variety of escape mechanisms has been described thus far, but progress in counteracting them is poor. Here we review a novel strategy to target malignancies with defects in the antigenic processing machinery (APM). The concept is based on a unique category of CD8+ T-cell epitopes that is associated with impaired peptide processing, which we named TEIPP. We characterized this alternative peptide repertoire emerging in MHC-I on tumors lacking classical antigen processing due to defects in the peptide transporter TAP (transporter associated with peptide processing). These TEIPPs exemplify interesting parallels with the folktale figure Cinderella: they are oppressed and neglected by a stepmother (like functional TAP prevents TEIPP presentation), until the suppression is released and Cinderella/TEIPP achieves unexpected recognition. TEIPP-specific CTLs and their cognate peptide-epitopes provide a new strategy to counteract immune evasion by APM defects and bear potential to targeting escape variants observed in a wide range of cancers.

  19. Improvements and Variants of the Multiple Antigen Blot Assay-MABA: An Immunoenzymatic Technique for Simultaneous Antigen and Antibody Screening.

    Science.gov (United States)

    Noya, Oscar; Losada, Sandra; Toledo, Marilyan; Gauna, Adriana; Lorenzo, María Angelita; Bermúdez, Henry; de Noya, Belkisyolé Alarcón

    2015-01-01

    This simple, versatile, reliable, reproducible, multipurpose, and inexpensive technique is based on the adhesion of different antigens to a single nitrocellulose strip using, as template, an acrylic device containing 28 parallel channels. The inclusion of channels containing normal human serum improves the quality control of this assay. Antigen-sensitized nitrocellulose strips are cut perpendicularly to the antigen-rows, exposed to immune sera followed by the appropriate conjugate. Positive signals are recorded using chemiluminescent or precipitable colorimetric substrates. This assay allows the simultaneous qualitative demonstration of antigenicity and immunogenicity of antigens obtained as synthetic peptides, recombinant molecules, or crude preparations, with high sensitivity and specificity. Its major value is based on the rapid and simultaneous comparative evaluation of various antigenic preparations allowing the diagnosis of a variety of infectious, allergic, and autoimmune diseases. It can in general be used to detect any type of antibody or circulating antigen. Some improvements and variants of the original technique are included.

  20. Structural analysis of hepatitis B surface antigen by monoclonal antibodies.

    Science.gov (United States)

    Ben-Porath, E; Wands, J R; Marciniak, R A; Wong, M A; Hornstein, L; Ryder, R; Canlas, M; Lingao, A; Isselbacher, K J

    1985-10-01

    A method has been developed for the analysis of hepatitis B surface antigen (HBsAg) antigenic structure at the molecular level that creates "fingerprints" or "signatures" of various hepatitis B viral (HBV) strains. This technique employs high affinity IgM and IgG monoclonal antibodies (anti-HBs) directed against distinct and separate determinants on HBsAg. In performing this antigenic structural analysis, separate binding curves for different monoclonal anti-HBs are generated by measuring immunoreactivity in serial dilutions of HBsAg-positive serum by radioimmunoassay. Since the HBsAg concentration in serum is unknown, the binding profiles of groups of samples are aligned by an iterative least-squares procedure to generate the numerical signature characteristic of the viral strain. The numerical signatures are then displayed on a computer-graphic plot. The signature profiles of HBsAg subtypes are a true reflection of their antigenic structure, and in vertical and horizontal transmission studies the molecular characteristics of the viral epitopes are conserved. By signature analysis we found substantial antigenic heterogeneity among the ayw3 strain both in the U.S. and France, as well as in populations of the Far East and Africa. Populations in Ethiopia, Gambia, and the Philippines were infected with two antigenically distinct HBV strains. In some newly identified HBV strains, it was found that epitopes identified by some monoclonal antibodies were absent or substantially reduced, which suggested that a genetic mutation may have occurred. Thus this study suggests that there is far more antigenic heterogeneity in HBV than previously recognized. These variants are antigenically distinct from each other at the epitope level, and were heretofore unrecognized by polyvalent anti-HBsAg antibodies.

  1. Absence of kinetoplast DNA in a late antigenic variant of Trypanosoma equiperdum.

    Science.gov (United States)

    Riou, G; Baltz, T; Gabillot, M; Pautrizel, R

    1980-04-01

    We have analysed by several biochemical techniques the DNA components of two antigenic variants isolated from Trypanosoma equiperdum. We did not observe any kinetoplast DNA (kDNA) structures or networks in the late antigen variant BoTat 28. Furthermore, the results of reassociation kinetics of in vitro labelled kDNA show that neither kDNA minicircle sequences nor kDNA maxicircle sequences of BoTat 1, the basic antigen type, can be detected in the total DNA of BoTat 28.

  2. Mutated epitopes of hepatitis B surface antigen fused to the core antigen of the virus induce antibodies that react with the native surface antigen.

    Science.gov (United States)

    Shiau, A L; Murray, K

    1997-03-01

    Fusion of peptide epitopes to the core antigen (HBcAg) of hepatitis B virus (HBV) enhances their immunogenicity, both quantitatively and qualitatively. In a number of vaccine-induced mutants of HBV, glycine145 of the surface antigen S polypeptide (HBsAg) has been replaced by arginine, resulting in loss of cross-reactivity with antibodies to normal (wild-type) HBsAg. HBcAg fusion proteins carrying the immunodominant epitope of HBsAg, in which glycine145 was replaced by arginine, glutamic acid, or lysine, were produced in Escherichia coli and formed particles that displayed HBc antigenicity and immunogenicity similar to that of HBcAg itself. The fusion proteins also elicited T-cell proliferative responsiveness to HBcAg and HBsAg. Fusions carrying either wild-type or mutated epitopes of HBsAG showed HBs antigenicity in immunoblot analysis and antigen-capture immunoradiometric assay, but both mutant and wild-type derivatives induced antibodies that cross-reacted with wild-type HBsAG. The results emphasise the potential for HBcAg fusion proteins in vaccines by broadening the antibody response in a way that could confer protection against both wild-type and variant form of HBV.

  3. 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...

  4. Characterization of antigenic variants of hepatitis C virus in immune evasion

    Directory of Open Access Journals (Sweden)

    Hershow Ronald

    2011-07-01

    Full Text Available Abstract Background Antigenic variation is an effective way by which viruses evade host immune defense leading to viral persistence. Little is known about the inhibitory mechanisms of viral variants on CD4 T cell functions. Results Using sythetic peptides of a HLA-DRB1*15-restricted CD4 epitope derived from the non-structural (NS 3 protein of hepatitis C virus (HCV and its antigenic variants and the peripheral blood mononuclear cells (PBMC from six HLA-DRB1*15-positive patients chronically infected with HCV and 3 healthy subjects, the in vitro immune responses and the phenotypes of CD4+CD25+ cells of chronic HCV infection were investigated. The variants resulting from single or double amino acid substitutions at the center of the core region of the Th1 peptide not only induce failed T cell activation but also simultaneously up-regulate inhibitory IL-10, CD25-TGF-β+ Th3 and CD4+IL-10+ Tr1 cells. In contrast, other variants promote differentiation of CD25+TGF-β+ Th3 suppressors that attenuate T cell proliferation. Conclusions Naturally occuring HCV antigenic mutants of a CD4 epitope can shift a protective peripheral Th1 immune response into an inhibitory Th3 and/or Tr1 response. The modulation of antigenic variants on CD4 response is efficient and extensive, and is likely critical in viral persistence in HCV infection.

  5. Immunity to Intracellular Salmonella Depends on Surface-associated Antigens

    Science.gov (United States)

    Claudi, Beatrice; Mazé, Alain; Schemmer, Anne K.; Kirchhoff, Dennis; Schmidt, Alexander; Burton, Neil; Bumann, Dirk

    2012-01-01

    Invasive Salmonella infection is an important health problem that is worsening because of rising antimicrobial resistance and changing Salmonella serovar spectrum. Novel vaccines with broad serovar coverage are needed, but suitable protective antigens remain largely unknown. Here, we tested 37 broadly conserved Salmonella antigens in a mouse typhoid fever model, and identified antigen candidates that conferred partial protection against lethal disease. Antigen properties such as high in vivo abundance or immunodominance in convalescent individuals were not required for protectivity, but all promising antigen candidates were associated with the Salmonella surface. Surprisingly, this was not due to superior immunogenicity of surface antigens compared to internal antigens as had been suggested by previous studies and novel findings for CD4 T cell responses to model antigens. Confocal microscopy of infected tissues revealed that many live Salmonella resided alone in infected host macrophages with no damaged Salmonella releasing internal antigens in their vicinity. In the absence of accessible internal antigens, detection of these infected cells might require CD4 T cell recognition of Salmonella surface-associated antigens that could be processed and presented even from intact Salmonella. In conclusion, our findings might pave the way for development of an efficacious Salmonella vaccine with broad serovar coverage, and suggest a similar crucial role of surface antigens for immunity to both extracellular and intracellular pathogens. PMID:23093937

  6. Exposure of the Plasmodium falciparum clonally variant STEVOR proteins on the merozoite surface

    Directory of Open Access Journals (Sweden)

    Meri Seppo

    2011-03-01

    Full Text Available Abstract Background Plasmodium falciparum merozoites are free invasive forms that invade host erythrocytes in iterative cycles in the presence of different arms of the immune system. Variant antigens are known to play a role in immune evasion and several gene families coding for variant antigens have been identified in P. falciparum. However, none of them have been reported to be expressed on the surface of merozoites. Methods Flow cytometry, immunofluorescence microscopy, and immunoblotting assays were performed to assess surface exposure, membrane association and stage specific expression of the STEVOR family of variants proteins, respectively. Results Using a polyclonal antibody (anti-PFL2610w with a broad specificity towards different STEVOR variants, the STEVOR proteins were identified on the surface of non-permeabilized/non-fixed merozoites in flow cytometry assays. Anti-PFL2610w antibody showed that several STEVORs were expressed in the trophozoite stage of the parasite but only one variant was integrated into the merozoite membrane. Moreover, this antibody failed to identify STEVORs on the surface of the parent schizont infected erythrocytes (IE although they were readily identified when schizont IE were permeabilized. Conclusions These data suggest for a role for STEVOR in immune evasion by P. falciparum merozoites to allow successful invasion of erythrocytes. Additionally, the expression of STEVORs in the schizont stage may only represent a step in the biogenesis process of the merozoite surface coat.

  7. 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.

  8. SMIM1 variants rs1175550 and rs143702418 independently modulate Vel blood group antigen expression

    Science.gov (United States)

    Christophersen, Mikael K.; Jöud, Magnus; Ajore, Ram; Vege, Sunitha; Ljungdahl, Klara W.; Westhoff, Connie M.; Olsson, Martin L.; Storry, Jill R.; Nilsson, Björn

    2017-01-01

    The Vel blood group antigen is expressed on the red blood cells of most individuals. Recently, we described that homozygosity for inactivating mutations in SMIM1 defines the rare Vel-negative phenotype. Still, Vel-positive individuals show great variability in Vel antigen expression, creating a risk for Vel blood typing errors and transfusion reactions. We fine-mapped the regulatory region located in SMIM1 intron 2 in Swedish blood donors, and observed a strong correlation between expression and rs1175550 as well as with a previously unreported tri-nucleotide insertion (rs143702418; C > CGCA). While the two variants are tightly linked in Caucasians, we separated their effects in African Americans, and found that rs1175550G and to a lesser extent rs143702418C independently increase SMIM1 and Vel antigen expression. Gel shift and luciferase assays indicate that both variants are transcriptionally active, and we identified binding of the transcription factor TAL1 as a potential mediator of the increased expression associated with rs1175550G. Our results provide insight into the regulatory logic of Vel antigen expression, and extend the set of markers for genetic Vel blood group typing. PMID:28084402

  9. TYPE-SPECIFIC ANTIGENS, M AND T, OF MATT AND GLOSSY VARIANTS OF GROUP A HEMOLYTIC STREPTOCOCCI.

    Science.gov (United States)

    Lancefield, R C

    1940-03-31

    1. Two qualitatively different type-specific antigens, designated M and T, have been found present in matt variants of group A hemolytic streptococci, but only one of these, the T antigen, occurs in the degraded glossy variant. 2. The protein nature of the M antigen, present in matt variants only, has been demonstrated in previous work, but the chemical characteristics of the newly recognized antigenic factor, T, present in both variants, have not been determined. This T factor is identified only by its immunological reactions. It is unknown whether the two type-specific antigenic factors, M and T, occur as separate chemical entities in the matt variant or in conjugation. 3. Antibody to the type-specific protein, M, appears responsible for the M precipitin reaction, for type-specific protection, and, as a rule, for part of the type-specific agglutination of matt variants, but in type 1 it does not cause agglutination. 4. Antibody to the second type-specific antigen, T, seems to be solely responsible for type-specific agglutination of the glossy form and to play a large rôle in type-specific agglutination of the matt form, but apparently it is not involved in protection. This T antibody causes all of the type-specific agglutination of type 1. Consequently, type 1 matt and glossy variants agglutinate and absorb agglutinin alike, and antisera to both are identical in content of type-specific agglutinin though they differ in respect to M antibody. 5. Recognition of the principle underlying type-specific agglutination of glossy variants makes it possible to suggest, with certain reservations, the use of glossy variants for type classification by agglutination. These variants are suitable for preparing type-specific agglutinating antisera, and they form stable suspensions for use in the reaction. Improved methods are needed for deriving glossy from matt variants.

  10. 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.

  11. A LAIR-1 insertion generates broadly reactive antibodies against malaria variant antigens

    OpenAIRE

    Tan, Joshua; Pieper, Kathrin; Piccoli, Luca; Abdi, Abdirahman; Perez, Mathilde Foglierini; Geiger, Roger; Tully, Claire Maria; Jarrossay, David; Maina Ndungu, Francis; Wambua, Juliana; Bejon, Philip; Fregni, Chiara Silacci; Fernandez-Rodriguez, Blanca; Barbieri, Sonia; Bianchi, Siro

    2015-01-01

    Plasmodium falciparum antigens expressed on the surface of infected erythrocytes are important targets of naturally acquired immunity against malaria, but their high number and variability provide the pathogen with a powerful means of escape from host antibodies 1?4 . Although broadly reactive antibodies against these antigens could be useful as therapeutics and in vaccine design, their identification has proven elusive. Here, we report the isolation of human monoclonal antibodies that recogn...

  12. Neutralizing monoclonal antibodies recognize antigenic variants among isolates of infectious hematopoietic necrosis virus

    Science.gov (United States)

    Winton, J.R.; Arakawa, C.N.; Lannan, C.N.; Fryer, J.L.

    1988-01-01

    eutralizing monoclonal antibodies were developed against strains of infectious hematopoietic necrosis virus (IHNV) from steelhead trout Salmo gairdneri in the Deschutes River of Oregon, chinook salmon Oncorhynchus tshawytscha in the Sacramento River of California, and rainbow trout Salmo gairdneri reared in the Hagerman Valley of Idaho, USA. These antibodies were tested for neutralization of 12 IHNV isolates obtained from salmonids in Japan, Alaska, Washington, Oregon, California, and Idaho. The antibodies recognized antigenic variants among the isolates and could be used to separate the viruses into 4 groups. The members of each group tended to be related by geographic area rather than by source host species, virulence, or date of isolation.

  13. Antibodies to variable Plasmodium falciparum-infected erythrocyte surface antigens are associated with protection from novel malaria infections

    DEFF Research Database (Denmark)

    Giha, H A; Staalsoe, T; Dodoo, D

    2000-01-01

    is maintained at low densities. Here, we test the hypothesis that the presence or absence of antibodies against variant antigens on the surface of P. falciparum-infected erythrocytes protect individuals against some infectious challenges and render them susceptible to others. Plasma collected in Daraweesh...

  14. Plasma antibodies from malaria-exposed pregnant women recognize variant surface antigens on Plasmodium falciparum-infected erythrocytes in a parity-dependent manner and block parasite adhesion to chondroitin sulfate A

    DEFF Research Database (Denmark)

    Ricke, C H; Staalsoe, T; Koram, K

    2000-01-01

    In areas of intense Plasmodium falciparum transmission, clinical immunity is acquired during childhood, and adults enjoy substantial protection against malaria. An exception to this rule is pregnant women, in whom malaria is both more prevalent and severe than in nonpregnant women. Pregnancy...... an area of hyperendemic P. falciparum transmission generally possessed low levels of Abs specifically recognizing surface Ags expressed by a CSA-adhering parasite isolate, while unselected isolates were well recognized. In marked contrast, most third-trimester pregnant women from that area had very high...

  15. seroprevalen ce of hepatitis b surface antigen emia among health ...

    African Journals Online (AJOL)

    Objective: The aim of this study was to determine the seroprevalence of Hepatitis B surface antigenaemia among the health care workers at the ... respondents and Hepatitis B surface antigen status determined using the agglutination kits manufactured by .... care workers in Beliza, Central America. Am J. Trop MedHyg.

  16. 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...

  17. Mapping antigenic motifs in the trypomastigote small surface antigen from Trypanosoma cruzi.

    Science.gov (United States)

    Balouz, Virginia; Cámara, María de Los Milagros; Cánepa, Gaspar E; Carmona, Santiago J; Volcovich, Romina; Gonzalez, Nicolás; Altcheh, Jaime; Agüero, Fernán; Buscaglia, Carlos A

    2015-03-01

    The trypomastigote small surface antigen (TSSA) is a mucin-like molecule from Trypanosoma cruzi, the etiological agent of Chagas disease, which displays amino acid polymorphisms in parasite isolates. TSSA expression is restricted to the surface of infective cell-derived trypomastigotes, where it functions as an adhesin and engages surface receptors on the host cell as a prerequisite for parasite internalization. Previous results have established TSSA-CL, the isoform encoded by the CL Brener clone, as an appealing candidate for use in serology-based diagnostics for Chagas disease. Here, we used a combination of peptide- and recombinant protein-based tools to map the antigenic structure of TSSA-CL at maximal resolution. Our results indicate the presence of different partially overlapping B-cell epitopes clustering in the central portion of TSSA-CL, which contains most of the polymorphisms found in parasite isolates. Based on these results, we assessed the serodiagnostic performance of a 21-amino-acid-long peptide that spans TSSA-CL major antigenic determinants, which was similar to the performance of the previously validated glutathione S-transferase (GST)-TSSA-CL fusion molecule. Furthermore, the tools developed for the antigenic characterization of the TSSA antigen were also used to explore other potential diagnostic applications of the anti-TSSA humoral response in Chagasic patients. Overall, our present results provide additional insights into the antigenic structure of TSSA-CL and support this molecule as an excellent target for molecular intervention in Chagas disease. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  18. 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.

  19. Structural Analysis of Histo-Blood Group Antigen Binding Specificity in a Norovirus GII.4 Epidemic Variant: Implications for Epochal Evolution

    Energy Technology Data Exchange (ETDEWEB)

    Shanker, Sreejesh; Choi, Jae-Mun; Sankaran, Banumathi; Atmar, Robert L.; Estes, Mary K.; Prasad, B.V. Venkataram (Baylor); (LBNL)

    2012-03-23

    Susceptibility to norovirus (NoV), a major pathogen of epidemic gastroenteritis, is associated with histo-blood group antigens (HBGAs), which are also cell attachment factors for this virus. GII.4 NoV strains are predominantly associated with worldwide NoV epidemics with a periodic emergence of new variants. The sequence variations in the surface-exposed P domain of the capsid protein resulting in differential HBGA binding patterns and antigenicity are suggested to drive GII.4 epochal evolution. To understand how temporal sequence variations affect the P domain structure and contribute to epochal evolution, we determined the P domain structure of a 2004 variant with ABH and secretor Lewis HBGAs and compared it with the previously determined structure of a 1996 variant. We show that temporal sequence variations do not affect the binding of monofucosyl ABH HBGAs but that they can modulate the binding strength of difucosyl Lewis HBGAs and thus could contribute to epochal evolution by the potentiated targeting of new variants to Lewis-positive, secretor-positive individuals. The temporal variations also result in significant differences in the electrostatic landscapes, likely reflecting antigenic variations. The proximity of some of these changes to the HBGA binding sites suggests the possibility of a coordinated interplay between antigenicity and HBGA binding in epochal evolution. From the observation that the regions involved in the formation of the HBGA binding sites can be conformationally flexible, we suggest a plausible mechanism for how norovirus disassociates from salivary mucin-linked HBGA before reassociating with HBGAs linked to intestinal epithelial cells during its passage through the gastrointestinal tract.

  20. Rational development and characterization of humanized anti–EGFR variant III chimeric antigen receptor T cells for glioblastoma

    Science.gov (United States)

    Johnson, Laura A.; Scholler, John; Ohkuri, Takayuki; Kosaka, Akemi; Patel, Prachi R.; McGettigan, Shannon E.; Nace, Arben K.; Dentchev, Tzvete; Thekkat, Pramod; Loew, Andreas; Boesteanu, Alina C.; Cogdill, Alexandria P.; Chen, Taylor; Fraietta, Joseph A.; Kloss, Christopher C.; Posey, Avery D.; Engels, Boris; Singh, Reshma; Ezell, Tucker; Idamakanti, Neeraja; Ramones, Melissa H.; Li, Na; Zhou, Li; Plesa, Gabriela; Seykora, John T.; Okada, Hideho; June, Carl H.; Brogdon, Jennifer L.; Maus, Marcela V.

    2015-01-01

    Chimeric antigen receptors (CARs) are synthetic molecules designed to redirect T cells to specific antigens. CAR-modified T cells can mediate long-term durable remissions in B cell malignancies, but expanding this platform to solid tumors requires the discovery of surface targets with limited expression in normal tissues. The variant III mutation of the epidermal growth factor receptor (EGFRvIII) results from an in-frame deletion of a portion of the extracellular domain, creating a neoepitope. We chose a vector backbone encoding a second-generation CAR based on efficacy of a murine scFv–based CAR in a xenograft model of glioblastoma. Next, we generated a panel of humanized scFvs and tested their specificity and function as soluble proteins and in the form of CAR-transduced T cells; a low-affinity scFv was selected on the basis of its specificity for EGFRvIII over wild-type EGFR. The lead candidate scFv was tested in vitro for its ability to direct CAR-transduced T cells to specifically lyse, proliferate, and secrete cytokines in response to antigen-bearing targets. We further evaluated the specificity of the lead CAR candidate in vitro against EGFR-expressing keratinocytes and in vivo in a model of mice grafted with normal human skin. EGFRvIII-directed CAR T cells were also able to control tumor growth in xenogeneic subcutaneous and orthotopic models of human EGFRvIII+ glioblastoma. On the basis of these results, we have designed a phase 1 clinical study of CAR T cells transduced with humanized scFv directed to EGFRvIII in patients with either residual or recurrent glioblastoma (NCT02209376). PMID:25696001

  1. Rational development and characterization of humanized anti-EGFR variant III chimeric antigen receptor T cells for glioblastoma.

    Science.gov (United States)

    Johnson, Laura A; Scholler, John; Ohkuri, Takayuki; Kosaka, Akemi; Patel, Prachi R; McGettigan, Shannon E; Nace, Arben K; Dentchev, Tzvete; Thekkat, Pramod; Loew, Andreas; Boesteanu, Alina C; Cogdill, Alexandria P; Chen, Taylor; Fraietta, Joseph A; Kloss, Christopher C; Posey, Avery D; Engels, Boris; Singh, Reshma; Ezell, Tucker; Idamakanti, Neeraja; Ramones, Melissa H; Li, Na; Zhou, Li; Plesa, Gabriela; Seykora, John T; Okada, Hideho; June, Carl H; Brogdon, Jennifer L; Maus, Marcela V

    2015-02-18

    Chimeric antigen receptors (CARs) are synthetic molecules designed to redirect T cells to specific antigens. CAR-modified T cells can mediate long-term durable remissions in B cell malignancies, but expanding this platform to solid tumors requires the discovery of surface targets with limited expression in normal tissues. The variant III mutation of the epidermal growth factor receptor (EGFRvIII) results from an in-frame deletion of a portion of the extracellular domain, creating a neoepitope. We chose a vector backbone encoding a second-generation CAR based on efficacy of a murine scFv-based CAR in a xenograft model of glioblastoma. Next, we generated a panel of humanized scFvs and tested their specificity and function as soluble proteins and in the form of CAR-transduced T cells; a low-affinity scFv was selected on the basis of its specificity for EGFRvIII over wild-type EGFR. The lead candidate scFv was tested in vitro for its ability to direct CAR-transduced T cells to specifically lyse, proliferate, and secrete cytokines in response to antigen-bearing targets. We further evaluated the specificity of the lead CAR candidate in vitro against EGFR-expressing keratinocytes and in vivo in a model of mice grafted with normal human skin. EGFRvIII-directed CAR T cells were also able to control tumor growth in xenogeneic subcutaneous and orthotopic models of human EGFRvIII(+) glioblastoma. On the basis of these results, we have designed a phase 1 clinical study of CAR T cells transduced with humanized scFv directed to EGFRvIII in patients with either residual or recurrent glioblastoma (NCT02209376). Copyright © 2015, American Association for the Advancement of Science.

  2. Variant surface antigens, virulence genes and the pathogenesis of malaria

    DEFF Research Database (Denmark)

    Deitsch, Kirk W; Hviid, Lars

    2004-01-01

    The first Molecular Approaches to Malaria meeting was held 2-5 February 2000 in Lorne, Australia. Following the meeting, Brian Cooke, Mats Wahlgren and Ross Coppel predicted that research into the molecular details of the mechanisms behind sequestration of parasitized erythrocytes would "become...... increasingly more complicated, with further interactions, receptors, ligands and functional domains". Furthermore, they cautioned that "the challenge will be not to lose ourselves in the molecular detail, but remain focused on the role of [the var genes and other multigene families] in pathogenesis of malaria......". We contemplate on these statements, following the recent second Molecular Approaches to Malaria meeting, which was held at the same venue on 2-5 February 2004....

  3. Sero-prevalence of hepatitis B surface antigen among primary ...

    African Journals Online (AJOL)

    Background and objective: Hepatitis B virus infection is a major public health problem worldwide and in Africa. In the northeastern part of Nigeria. Information on the prevalence of HBV infection in rural communities is scarce. This study was carried out to determine the sero-prevalence of hepatitis B surface antigen amongst ...

  4. Prevalence of Hepatitis B Surface Antigen among Women of ...

    African Journals Online (AJOL)

    This study documents the seroprevalence of hepatitis B surface antigen (HBs Ag) among women of childbearing age attending various family clinics in Lagos, Nigeria. A total of 501 women were screened with Wellcozyme ELISA technique, of which 45(8.9%) were seropositive. Women in occupations related to needle work ...

  5. Prevalence of Hepatitis B surface antigen among pregnant women ...

    African Journals Online (AJOL)

    Prevalence of Hepatitis B surface antigen among pregnant women attending antenatal clinic at Nyamagana ... Journal Home > Vol 18, No 1 (2016) > ... The PDF file you selected should load here if your Web browser has a PDF reader plug-in ...

  6. Prevalence of hepatitis B surface antigen seropositivity among HIV ...

    African Journals Online (AJOL)

    Hepatitis B surface antigen (HBsAg) was tested for using a one step lateral flow rapid chromatographic immunoassay (Acumen labs and diagnostic centre, Bangalore, India) and HIV 1/2 was tested using two kits, Determine (made by Abbot, Japan for Inverness Medical, Japan). Results: A total of 2018 subjects were studied ...

  7. Seroprevalence of hepatitis B Surface antigen among apparently ...

    African Journals Online (AJOL)

    Seroprevalence of hepatitis B Surface antigen among apparently healthy primary school pupils in Batagarawa Local Government area of Katsina State, Nigeria. ... If you would like more information about how to print, save, and work with PDFs, Highwire Press provides a helpful Frequently Asked Questions about PDFs.

  8. Prevalence of hepatitis B surface antigen sero-positivity and ...

    African Journals Online (AJOL)

    2016-07-31

    Jul 31, 2016 ... virus infections. Hepatitis B virus infection was screened using hepatitis B surface antigen by ELISA, while hepatitis C virus infection was screened using anti-HCV antibodies by ELISA. The biodata of the donors were obtained. The prevalence of hepatitis infection among the blood donors was 51 (9.6%).

  9. 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 ...

  10. Seroprevalence of Hepatitis B Surface Antigen (HBsAg) among ...

    African Journals Online (AJOL)

    Seroprevalence of Hepatitis B Surface Antigen (HBsAg) among Children of Primary School Age in a Community, North-Central, Nigeria. ... Furthermore, family history of HBV infection accounted for 3.6%, while male subjects that had traditional method of circumcision recorded a high prevalence of 3.3%. Unfortunately, the ...

  11. Seroprevalence of Hepatitis B surface antigen (HBSAG) among ...

    African Journals Online (AJOL)

    Seroprevalence of Hepatitis B surface antigen (HBSAG) among pregnant women in Aboh Mbaise local government area of Imo State, Nigeria. ... Information on gestational age of the expectant mothers, gravidity, ... B. As a consequence, it is advocated that pregnant women be tested for HBsAg during an early prenatal visit.

  12. Prediction of antigenic epitopes on protein surfaces by consensus scoring

    Directory of Open Access Journals (Sweden)

    Zhang Chi

    2009-09-01

    Full Text Available Abstract Background Prediction of antigenic epitopes on protein surfaces is important for vaccine design. Most existing epitope prediction methods focus on protein sequences to predict continuous epitopes linear in sequence. Only a few structure-based epitope prediction algorithms are available and they have not yet shown satisfying performance. Results We present a new antigen Epitope Prediction method, which uses ConsEnsus Scoring (EPCES from six different scoring functions - residue epitope propensity, conservation score, side-chain energy score, contact number, surface planarity score, and secondary structure composition. Applied to unbounded antigen structures from an independent test set, EPCES was able to predict antigenic eptitopes with 47.8% sensitivity, 69.5% specificity and an AUC value of 0.632. The performance of the method is statistically similar to other published methods. The AUC value of EPCES is slightly higher compared to the best results of existing algorithms by about 0.034. Conclusion Our work shows consensus scoring of multiple features has a better performance than any single term. The successful prediction is also due to the new score of residue epitope propensity based on atomic solvent accessibility.

  13. Glycosylation of the major human Pneumocystis carinii surface antigen

    DEFF Research Database (Denmark)

    Lundgren, Bettina; Koch, C; Mathiesen, Lars Reinhardt

    1993-01-01

    It has recently been shown that the major rat P. carinii surface antigen is important for initial host-organism attachment, possibly through binding to fibronectin, mannose-binding protein, or surfactant protein A. Since a carbohydrate/lectin interaction may be involved in adhesion, we undertook...... this study to characterize the glycosylation of the major human P. carinii surface glycoprotein (gp95). We have used purified gp95 as a source of antigen, and in lectin binding and deglycosylation studies it was found that approximately 9% of gp95 consists of N-linked carbohydrates of mainly high......-mannose and bisected complex-type glycans. Using a polyclonal antibody raised against purified gp95 and crossed affinoimmunoelectrophoresis and the lectins Con A and WGA, gp95 exhibited carbohydrate-dependent microheterogeneity. We therefore suggest that gp95 is composed of subtypes which differ in N...

  14. Exposed epitopes on a Trypanosoma equiperdum variant surface glycoprotein altered by point mutations.

    Science.gov (United States)

    Baltz, T; Giroud, C; Bringaud, F; Eisen, H; Jacquemot, C; Roth, C W

    1991-07-01

    African trypanosomes are covered by a dense protein layer that is immunologically distinct on different trypanosome isolates and is termed the variant surface glycoprotein (VSG). The different VSGs are expressed in a general order, where some VSGs appear preferentially early in infection and others only later. The exposed epitopes on a late antigen, VSG 78, of T.equiperdum were studied by the technique of monoclonal antibody (MAb) escape selection. MAbs that neutralize trypanosomes bearing VSG 78 reacted with the VSG only when it was attached to the trypanosome surface, suggesting that the most immunogenic surface epitopes are conformational. Trypanosome clones resistant to one of the MAbs yet still expressing VSG 78 or 78(20) were isolated in vitro. Two independent variants resistant to MAb H3 changed Ser192 to Arg by a single base change in the VSG gene and a variant resistant to MAb H21 had a single base change that converted Gln172 to Glu. A variant resistant to MAb H7 had several changes in the VSG gene, a gene conversion in the 5' region and an isolated mutation in codon 220 that is proposed to be responsible for the resistance phenotype. The isotypic bias of the MAbs against VSG 78 and an analysis of the natural variants that are resistant to MAb 78H21 suggest that glycosylation plays a role in the immunogenicity of these proteins. The analysis defines some of the exposed amino acid residues and demonstrates that VSG genes are altered by mutations and small gene conversions as well as replaced by large gene conversion-like events. The results provide biological data supporting the model of VSG structure obtained by crystallographic studies.

  15. The Trypomastigote Small Surface Antigen (TSSA) regulates Trypanosoma cruzi infectivity and differentiation.

    Science.gov (United States)

    Cámara, María de Los Milagros; Cánepa, Gaspar E; Lantos, Andrés B; Balouz, Virginia; Yu, Hai; Chen, Xi; Campetella, Oscar; Mucci, Juan; Buscaglia, Carlos A

    2017-08-01

    TSSA (Trypomastigote Small Surface Antigen) is an antigenic, adhesion molecule displayed on the surface of Trypanosoma cruzi trypomastigotes. TSSA displays substantial sequence identity to members of the TcMUC gene family, which code for the trypomastigote mucins (tGPI-mucins). In addition, TSSA bears sequence polymorphisms among parasite strains; and two TSSA variants expressed as recombinant molecules (termed TSSA-CL and TSSA-Sy) were shown to exhibit contrasting features in their host cell binding and signaling properties. Here we used a variety of approaches to get insights into TSSA structure/function. We show that at variance with tGPI-mucins, which rely on their extensive O-glycoslylation to achieve their protective function, TSSA seems to be displayed on the trypomastigote coat as a hypo-glycosylated molecule. This has a functional correlate, as further deletion mapping experiments and cell binding assays indicated that exposition of at least two peptidic motifs is critical for the engagement of the 'adhesive' TSSA variant (TSSA-CL) with host cell surface receptor(s) prior to trypomastigote internalization. These motifs are not conserved in the 'non-adhesive' TSSA-Sy variant. We next developed transgenic lines over-expressing either TSSA variant in different parasite backgrounds. In strict accordance to recombinant protein binding data, trypomastigotes over-expressing TSSA-CL displayed improved adhesion and infectivity towards non-macrophagic cell lines as compared to those over-expressing TSSA-Sy or parental lines. These phenotypes could be specifically counteracted by exogenous addition of peptides spanning the TSSA-CL adhesion motifs. In addition, and irrespective of the TSSA variant, over-expression of this molecule leads to an enhanced trypomastigote-to-amastigote conversion, indicating a possible role of TSSA also in parasite differentiation. In this study we provided novel evidence indicating that TSSA plays an important role not only on the

  16. The Trypomastigote Small Surface Antigen (TSSA regulates Trypanosoma cruzi infectivity and differentiation.

    Directory of Open Access Journals (Sweden)

    María de Los Milagros Cámara

    2017-08-01

    Full Text Available TSSA (Trypomastigote Small Surface Antigen is an antigenic, adhesion molecule displayed on the surface of Trypanosoma cruzi trypomastigotes. TSSA displays substantial sequence identity to members of the TcMUC gene family, which code for the trypomastigote mucins (tGPI-mucins. In addition, TSSA bears sequence polymorphisms among parasite strains; and two TSSA variants expressed as recombinant molecules (termed TSSA-CL and TSSA-Sy were shown to exhibit contrasting features in their host cell binding and signaling properties.Here we used a variety of approaches to get insights into TSSA structure/function. We show that at variance with tGPI-mucins, which rely on their extensive O-glycoslylation to achieve their protective function, TSSA seems to be displayed on the trypomastigote coat as a hypo-glycosylated molecule. This has a functional correlate, as further deletion mapping experiments and cell binding assays indicated that exposition of at least two peptidic motifs is critical for the engagement of the 'adhesive' TSSA variant (TSSA-CL with host cell surface receptor(s prior to trypomastigote internalization. These motifs are not conserved in the 'non-adhesive' TSSA-Sy variant. We next developed transgenic lines over-expressing either TSSA variant in different parasite backgrounds. In strict accordance to recombinant protein binding data, trypomastigotes over-expressing TSSA-CL displayed improved adhesion and infectivity towards non-macrophagic cell lines as compared to those over-expressing TSSA-Sy or parental lines. These phenotypes could be specifically counteracted by exogenous addition of peptides spanning the TSSA-CL adhesion motifs. In addition, and irrespective of the TSSA variant, over-expression of this molecule leads to an enhanced trypomastigote-to-amastigote conversion, indicating a possible role of TSSA also in parasite differentiation.In this study we provided novel evidence indicating that TSSA plays an important role not only on

  17. Antigenicity and immunogenicity of Plasmodium vivax merozoite surface protein-3.

    Directory of Open Access Journals (Sweden)

    Amanda R Bitencourt

    Full Text Available A recent clinical trial in African children demonstrated the potential utility of merozoite surface protein (MSP-3 as a vaccine against Plasmodium falciparum malaria. The present study evaluated the use of Plasmodium vivax MSP-3 (PvMSP-3 as a target antigen in vaccine formulations against malaria caused by P. vivax. Recombinant proteins representing MSP-3α and MSP-3β of P. vivax were expressed as soluble histidine-tagged bacterial fusions. Antigenicity during natural infection was evaluated by detecting specific antibodies using sera from individuals living in endemic areas of Brazil. A large proportion of infected individuals presented IgG antibodies to PvMSP-3α (68.2% and at least 1 recombinant protein representing PvMSP-3β (79.1%. In spite of the large responder frequency, reactivity to both antigens was significantly lower than was observed for the immunodominant epitope present on the 19-kDa C-terminal region of PvMSP-1. Immunogenicity of the recombinant proteins was studied in mice in the absence or presence of different adjuvant formulations. PvMSP-3β, but not PvMSP-3α, induced a TLR4-independent humoral immune response in the absence of any adjuvant formulation. The immunogenicity of the recombinant antigens were also tested in formulations containing different adjuvants (Alum, Salmonella enterica flagellin, CpG, Quil A,TiterMax® and incomplete Freunds adjuvant and combinations of two adjuvants (Alum plus flagellin, and CpG plus flagellin. Recombinant PvMSP-3α and PvMSP-3β elicited higher antibody titers capable of recognizing P. vivax-infected erythrocytes harvested from malaria patients. Our results confirm that P. vivax MSP-3 antigens are immunogenic during natural infection, and the corresponding recombinant proteins may be useful in elucidating their vaccine potential.

  18. Detection of antibodies to variant antigens on Plasmodium falciparum-infected erythrocytes by flow cytometry

    DEFF Research Database (Denmark)

    Staalsoe, T; Giha, H A; Dodoo, D

    1999-01-01

    BACKGROUND: Naturally induced antibodies binding to surface antigens of Plasmodium falciparum-infected erythrocytes can be detected by direct agglutination of infected erythrocytes or by indirect immunofluorescence on intact, unfixed, infected erythrocytes. Agglutinating antibodies have previously...

  19. A Molecular-Level Account of the Antigenic Hantaviral Surface

    Directory of Open Access Journals (Sweden)

    Sai Li

    2016-05-01

    Full Text Available Hantaviruses, a geographically diverse group of zoonotic pathogens, initiate cell infection through the concerted action of Gn and Gc viral surface glycoproteins. Here, we describe the high-resolution crystal structure of the antigenic ectodomain of Gn from Puumala hantavirus (PUUV, a causative agent of hemorrhagic fever with renal syndrome. Fitting of PUUV Gn into an electron cryomicroscopy reconstruction of intact Gn-Gc spike complexes from the closely related but non-pathogenic Tula hantavirus localized Gn tetramers to the membrane-distal surface of the virion. The accuracy of the fitting was corroborated by epitope mapping and genetic analysis of available PUUV sequences. Interestingly, Gn exhibits greater non-synonymous sequence diversity than the less accessible Gc, supporting a role of the host humoral immune response in exerting selective pressure on the virus surface. The fold of PUUV Gn is likely to be widely conserved across hantaviruses.

  20. Expression of hepatitis B surface antigen in transgenic banana plants.

    Science.gov (United States)

    Kumar, G B Sunil; Ganapathi, T R; Revathi, C J; Srinivas, L; Bapat, V A

    2005-10-01

    Embryogenic cells of bananan cv. Rasthali (AAB) have been transformed with the 's' gene of hepatitis B surface antigen (HBsAg) using Agrobacterium mediated transformation. Four different expression cassettes (pHBS, pHER, pEFEHBS and pEFEHER) were utilized to optimize the expression of HBsAg in banana. The transgenic nature of the plants and expression of the antigen was confirmed by PCR, Southern hybridization and reverse transcription (RT)-PCR. The expression levels of the antigen in the plants grown under in vitro conditions as well as the green house hardened plants were estimated by ELISA for all the four constructs. Maximum expression level of 38 ng/g F.W. of leaves was noted in plants transformed with pEFEHBS grown under in vitro conditions, whereas pHER transformed plants grown in the green house showed the maximum expression level of 19.92 ng/g F.W. of leaves. Higher monoclonal antibody binding of 67.87% of the antigen was observed when it was expressed with a C-terminal ER retention signal. The buoyant density in CsCl of HBsAg derived from transgenic banana leaves was determined and found to be 1.146 g/ml. HBsAg obtained from transgenic banana plants is similar to human serum derived one in buoyant density properties. The transgenic plants were grown up to maturity in the green house and the expression of HBsAg in the fruits was confirmed by RT-PCR. These transgenic plants were multiplied under in vitro using floral apex cultures. Attempts were also made to enhance the expression of HBsAg in the leaves of transgenic banana plants by wounding and/or treatment with plant growth regulators. This is the first report on the expression of HBsAg in transgenic banana fruits.

  1. HLA DR and AB surface antigens correlate with cell shape (surface area).

    Science.gov (United States)

    Ende, N; Ritter, A B

    1998-01-01

    In order to help explain some of the various phenomena associated with both benign and malignant cells, this study was undertaken to determine if changes in the shape of the cell could alter the recognition of the cell. Non-transformed Human cells, HEL 299, were evaluated for their shape and surface antigens. A direct statistical correlation was found between the two surface antigens HLA AB and DR and the cell shape (surface area). The possible significance of this phenomena in non transformed human cells to neoplastic proliferation is suggested.

  2. Genetic variation and significance of hepatitis B surface antigen

    Directory of Open Access Journals (Sweden)

    ZHANG Zhenhua

    2013-11-01

    Full Text Available Hepatitis B virus (HBV is prone to genetic variation because there is reverse transcription in the process of HBV replication. The gene mutation of hepatitis B surface antigen may affect clinical diagnosis of HBV infection, viral replication, and vaccine effect. The current research and existing problems are discussed from the following aspects: the mechanism and biological and clinical significance of S gene mutation. Most previous studies focused on S gene alone, so S gene should be considered as part of HBV DNA in the future research on S gene mutation.

  3. Serum anti-hepatitis B surface antigen in hemodialysis patients

    Directory of Open Access Journals (Sweden)

    Rafieian-Kopaei Mahmoud

    2012-01-01

    Full Text Available To evaluate the immune response to hepatitis B vaccination in stable hemodialysis (HD patients, a retro-prospective investigation was conducted on 68 HD patients. Participants were vaccinated against hepatitis B virus with an intramuscular hepatitis B vaccination schedule, 40 micrograms at 0, 1, and 6 months. The serum antibody level against hepatitis B surface antigen (HBs in HD patients was 35±55. In this study, no significant differences of Anti-HBs antibody between diabetic and non-diabetics or male and female subjects were observed. There were not any significant correlation between antibody against HBs-Ag and serum albumin. There was not significant correlation between anti-HBs antibody and age, proportion of HD, duration of HD or dialysis efficacy. In this study, there was not significant correlation between serum antibody level against hepatitis B surface antigen and some demographic indices of HD patients, however, these findings need to re-test in other centers with more participants.

  4. Human Cancer Antigen Globo H Is a Cell-Surface Ligand for Human Ribonuclease 1.

    Science.gov (United States)

    Eller, Chelcie H; Chao, Tzu-Yuan; Singarapu, Kiran K; Ouerfelli, Ouathek; Yang, Guangbin; Markley, John L; Danishefsky, Samuel J; Raines, Ronald T

    2015-07-22

    Pancreatic-type ribonucleases are secretory enzymes that catalyze the cleavage of RNA. Recent efforts have endowed the homologues from cow (RNase A) and human (RNase 1) with toxicity for cancer cells, leading to a clinical trial. The basis for the selective toxicity of ribonuclease variants for cancerous versus noncancerous cells has, however, been unclear. A screen for RNase A ligands in an array of mammalian cell-surface glycans revealed strong affinity for a hexasaccharide, Globo H, that is a tumor-associated antigen and the basis for a vaccine in clinical trials. The affinity of RNase A and RNase 1 for immobilized Globo H is in the low micromolar-high nanomolar range. Moreover, reducing the display of Globo H on the surface of human breast adenocarcinoma cells with a small-molecule inhibitor of biosynthesis or a monoclonal antibody antagonist decreases the toxicity of an RNase 1 variant. Finally, heteronuclear single quantum coherence (HSQC) NMR spectroscopy showed that RNase 1 interacts with Globo H by using residues that are distal from the enzymic active site. The discovery that a systemic human ribonuclease binds to a moiety displayed on human cancer cells links two clinical paradigms and suggests a mechanism for innate resistance to cancer.

  5. Characterisation of centromere (kinetochore) antigen reactive with sera of patients with a scleroderma variant (CREST syndrome).

    Science.gov (United States)

    Nishikai, M; Okano, Y; Yamashita, H; Watanabe, M

    1984-01-01

    Anticentromere (kinetochore) antibody is the marker antibody in CREST syndrome, but the precise molecular composition of the partner antigen has been poorly defined. This report describes for the first time a procedure for the successful extraction and biochemical characterisation of the centromere antigen molecule. The centromere antigen was extracted with 4M NaCl solution. The molecular weight of the partner antigen of the centromere antibody was determined to be 70 000 daltons by the SDS-PAGE and immunoblotting methods. A Sephacryl S-300 column experiment confirmed these results. Centromere antigenic activity was preserved at pHs between 3 and 11 and was resistant to three enzymes, trypsin, RNase, and DNase. Images PMID:6524984

  6. Identification of potential protein partners that bind to the variant surface glycoprotein in Trypanosoma equiperdum.

    Science.gov (United States)

    Carrasquel, Liomary M; Escalona, José L; Acosta-Serrano, Alvaro; Guo, Yurong; Bubis, José

    2017-06-01

    Trypanosoma equiperdum possesses a dense coat of a variant surface glycoprotein (VSG) that is used to evade the host immune response by a process known as antigenic variation. Soluble and membrane forms of the predominant VSG from the Venezuelan T. equiperdum TeAp-N/D1 strain (sVSG and mVSG, respectively) were purified to homogeneity; and antibodies against sVSG and mVSG were raised, isolated, and employed to produce anti-idiotypic antibodies that structurally mimic the VSG surface. Prospective VSG-binding partners were initially detected by far-Western blots, and then by immunoblots using the generated anti-idiotypic antibodies. Polypeptides of ~80 and 55 kDa were isolated when anti-idiotypic antibodies-Sepharose affinity matrixes were used as baits. Mass spectrometry sequencing yielded hits with various proteins from Trypanosoma brucei such as heat-shock protein 70, tryparedoxin peroxidase, VSG variants, expression site associated gene product 6, and two hypothetical proteins. In addition, a possible interaction with a protein homologous to the glutamic acid/alanine-rich protein from Trypanosoma congolense was also found. These results indicate that the corresponding orthologous gene products are candidates for VSG-interacting proteins in T. equiperdum.

  7. Quantification of hepatitis B surface antigen and E antigen: correlation between Elecsys and architect assays.

    Science.gov (United States)

    Zhou, B; Liu, M; Lv, G; Zheng, H; Wang, Y; Sun, J; Hou, J

    2013-06-01

    Quantification of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) and their change model during treatment are emerging as a useful tool for assessing the outcome of hepatitis B virus (HBV) infection and predicting the efficacy of antiviral therapy. The aim of this study was to compare the performance of the Elecsys and Architect assays for HBsAg and HBeAg quantification. Quantification of HBsAg and HBeAg, determined by these two assays, were assessed in 1292 sera from patients with chronic hepatitis B(CHB). HBeAg quantification in serum was performed by calibrating the results through HBeAg Paul-Ehrlich international (PEI) reference standard. The HBV genotype was determined by direct sequencing and phylogenetic analysis. Of 1292 samples, the distribution of genotype was 514 (39.78%) genotype B, 776 (60.06%) genotype C, 2 (0.16%) genotype D. The results of HBsAg and HBeAg quantification between the Architect and Elecsys assays were significantly correlated (HBsAg: r = 0.939; HBeAg: r = 0.987), independent of HBV genotype and treatment phase. The mean differences between the two methods (the log10 [Elecsys] - the log10 [Architect]) were 0.075 log10  IU/mL and -0.149 log10 PE IU/mL in quantifying HBsAg and HBeAg, respectively. This study demonstrates a high correlation between the Elecsys and the Architect assays in quantifying HBsAg and HBeAg, regardless of HBV genotype. Both the two assays can be used to monitor the HBsAg and HBeAg levels in patients with chronic hepatitis B. © 2013 John Wiley & Sons Ltd.

  8. Molecular characterization of atypical antigenic variants of canine rabies virus reveals its reintroduction by wildlife vectors in southeastern Mexico.

    Science.gov (United States)

    Garcés-Ayala, Fabiola; Aréchiga-Ceballos, Nidia; Ortiz-Alcántara, Joanna M; González-Durán, Elizabeth; Pérez-Agüeros, Sandra I; Méndez-Tenorio, Alfonso; Torres-Longoria, Belem; López-Martínez, Irma; Hernández-Rivas, Lucía; Díaz-Quiñonez, José Alberto; Ramírez-González, José Ernesto

    2017-12-01

    Rabies is an infectious viral disease that is practically always fatal following the onset of clinical signs. In Mexico, the last case of human rabies transmitted by dogs was reported in 2006 and canine rabies has declined significantly due to vaccination campaigns implemented in the country. Here we report on the molecular characterization of six rabies virus strains found in Yucatan and Chiapas, remarkably, four of them showed an atypical reaction pattern when antigenic characterization with a reduced panel of eight monoclonal antibodies was performed. Phylogenetic analyses on the RNA sequences unveiled that the three atypical strains from Yucatan are associated with skunks. Analysis using the virus entire genome showed that they belong to a different lineage distinct from the variants described for this animal species in Mexico. The Chiapas atypical strain was grouped in a lineage that was considered extinct, while the others are clustered within classic dog variants.

  9. Stable isotope labeling of oligosaccharide cell surface antigens

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, C.J.; Silks, L.A. III; Martinez, R.A. [and others

    1998-12-31

    The overall goal of this Laboratory Directed Research and Development (LDRD) project was to develop new methods for synthesis of {sup 13}C-labeled oligosaccharides that are required for nuclear magnetic resonance (NMR) studies of their solution conformation. Oligosaccharides are components of the cell`s outer surface and are involved in important processes such as cell-cell recognition and adhesion. Recently, Danishefsky and coworkers at Slone-Kettering Cancer Center developed a method for the solid-phase chemical synthesis of oligosaccharides. The specific goal of this LDRD project was to prepare uniform {sup 13}C-labeled aldohexose precursors required for the solid-phase synthesis of the Lewis blood-group antigenic determinants. We report the synthesis of {sup 13}C-labeled D-glucal, D-galactal and Fucosyl precursors. We have been collaborating with the Danishefsky group on the synthesis of the Lewis oligosaccharides and the NMR analysis of their solution conformation.

  10. Novel Non-Histocompatibility Antigen Mismatched Variants Improve the Ability to Predict Antibody-Mediated Rejection Risk in Kidney Transplant

    Directory of Open Access Journals (Sweden)

    Silvia Pineda

    2017-12-01

    Full Text Available Transplant rejection is the critical clinical end-point limiting indefinite survival after histocompatibility antigen (HLA mismatched organ transplantation. The predominant cause of late graft loss is antibody-mediated rejection (AMR, a process whereby injury to the organ is caused by donor-specific antibodies, which bind to HLA and non-HLA (nHLA antigens. AMR is incompletely diagnosed as donor/recipient (D/R matching is only limited to the HLA locus and critical nHLA immunogenic antigens remain to be identified. We have developed an integrative computational approach leveraging D/R exome sequencing and gene expression to predict clinical post-transplant outcome. We performed a rigorous statistical analysis of 28 highly annotated D/R kidney transplant pairs with biopsy-confirmed clinical outcomes of rejection [either AMR or T-cell-mediated rejection (CMR] and no-rejection (NoRej, identifying a significantly higher number of mismatched nHLA variants in AMR (ANOVA—p-value = 0.02. Using Fisher’s exact test, we identified 123 variants associated mainly with risk of AMR (p-value < 0.001. In addition, we applied a machine-learning technique to circumvent the issue of statistical power and we found a subset of 65 variants using random forest, that are predictive of post-tx AMR showing a very low error rate. These variants are functionally relevant to the rejection process in the kidney and AMR as they relate to genes and/or expression quantitative trait loci (eQTLs that are enriched in genes expressed in kidney and vascular endothelium and underlie the immunobiology of graft rejection. In addition to current D/R HLA mismatch evaluation, additional mismatch nHLA D/R variants will enhance the stratification of post-tx AMR risk even before engraftment of the organ. This innovative study design is applicable in all solid organ transplants, where the impact of mitigating AMR on graft survival may be greater, with considerable benefits on

  11. CARbodies: Human Antibodies Against Cell Surface Tumor Antigens Selected From Repertoires Displayed on T Cell Chimeric Antigen Receptors

    OpenAIRE

    Alonso-Camino, Vanesa; Sánchez-Martín, David; Compte, Marta; Nuñez-Prado, Natalia; Diaz, Rosa M; Vile, Richard; Alvarez-Vallina, Luis

    2013-01-01

    A human single-chain variable fragment (scFv) antibody library was expressed on the surface of human T cells after transduction with lentiviral vectors (LVs). The repertoire was fused to a first-generation T cell receptor ζ (TCRζ)-based chimeric antigen receptor (CAR). We used this library to isolate antibodies termed CARbodies that recognize antigens expressed on the tumor cell surface in a proof-of-principle system. After three rounds of activation-selection there was a clear repertoire res...

  12. Characterization of surface antigens of reticulated immature platelets.

    Science.gov (United States)

    Lador, Adi; Leshem-Lev, Dorit; Spectre, Galia; Abelow, Aryeh; Kornowski, Ran; Lev, Eli I

    2017-10-01

    Reticulated platelets (RPs) are immature platelets with high dense granules content and a residual amount of megakaryocyte-derived of mRNA. Increased level of RPs has been found to be an independent predictor of cardiovascular ischemic events, and has been associated with impaired response to various anti-platelet drugs. The study aimed to characterize and compare the surface antigenic properties of reticulated versus mature platelets. Platelets from healthy individuals and diabetic patients were tested at rest and after activation with adenosine diphosphate (ADP). For each patient, we calculated the proportion of RPs and mature platelets using flow cytometry analysis with thiazole orange staining (for RPs) and CD42b platelet-specific antibody. We also tested the surface expression of P-selectin and Annexin V, by double staining flow cytometry in RPs versus mature platelets. A total of 20 subjects were recruited (10 healthy individuals, 10 diabetics). Activation with ADP did not cause a significant change in the proportion of RPs. Following activation, RPs demonstrated a significant increase in the expression of both P-selectin and Annexin V, while mature platelets exhibited a non-significant increase in both markers. These findings were consistent in both healthy subjects and patients with diabetes. In conclusion, RPs have a significantly higher capacity to increase the expression of platelet activation markers compared with mature platelets.

  13. A unique variant of streptococcal group O-antigen (C-polysaccharide) that lacks phosphocholine

    DEFF Research Database (Denmark)

    Bergström, N; Jansson, P.-E.; Kilian, Mogens

    2003-01-01

    Streptococcus mitis strain SK598, which represents a subgroup of biovar 1, possesses a unique variant of the C-polysaccharide found in the cell wall of all strains of Streptococcus pneumoniae and in some strains of S. mitis. This new variant lacks the choline methyl groups in contrast to the prev......Streptococcus mitis strain SK598, which represents a subgroup of biovar 1, possesses a unique variant of the C-polysaccharide found in the cell wall of all strains of Streptococcus pneumoniae and in some strains of S. mitis. This new variant lacks the choline methyl groups in contrast...... to the previously characterized forms of C-polysaccharide, which all contain one or two choline residues per repeat. The following structure of the repeating unit of the SK598 polysaccharide was established: where AAT is 2-acetamido-4-amino-2,4,6-trideoxy-d-galactose. This structure is identical to the double...... choline-substituted form of C-polysaccharide, except that it is substituted with ethanolamine instead of choline. This extends the number of recognized C-polysaccharide variants to four....

  14. sero-prevalence of hepatitis b surface antigen (hbsag) among blood ...

    African Journals Online (AJOL)

    DR. AMINU

    ABSTRACT. A study on the sero-prevalence of Hepatitis B surface Antigen among 100 blood donors attending. Ahmadu Bello University Teaching Hospital Shika, Zaria, Kaduna Nigeria was carried out in June,. 2008 using hepatitis B surface Antigen latex. Higher prevalence rate was observed between the age range of 30 ...

  15. Effect of hepatitis B immunisation in newborn infants of mothers positive for hepatitis B surface antigen

    DEFF Research Database (Denmark)

    Lee, Chuanfang; Gong, Yan; Brok, Jesper

    2006-01-01

    To evaluate the effects of hepatitis B vaccine and immunoglobulin in newborn infants of mothers positive for hepatitis B surface antigen.......To evaluate the effects of hepatitis B vaccine and immunoglobulin in newborn infants of mothers positive for hepatitis B surface antigen....

  16. Sero-prevalence of hepatitis b surface antigen (hbsag) among blood ...

    African Journals Online (AJOL)

    A study on the sero-prevalence of Hepatitis B surface Antigen among 100 blood donors attending. Ahmadu Bello University Teaching Hospital Shika, Zaria, Kaduna Nigeria was carried out in June, 2008 using hepatitis B surface Antigen latex. Higher prevalence rate was observed between the age range of 30 – 39years.

  17. Surface co-expression of two different PfEMP1 antigens on single Plasmodium falciparum-infected erythrocytes facilitates binding to ICAM1 and PECAM1

    DEFF Research Database (Denmark)

    Joergensen, Louise; Bengtsson, Dominique C; Bengtsson, Anja

    2010-01-01

    The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) antigens play a major role in cytoadhesion of infected erythrocytes (IE), antigenic variation, and immunity to malaria. The current consensus on control of variant surface antigen expression is that only one PfEMP1 encoded by one var...... gene is expressed per cell at a time. We measured var mRNA transcript levels by real-time Q-PCR, analysed var gene transcripts by single-cell FISH and directly compared these with PfEMP1 antigen surface expression and cytoadhesion in three different antibody-selected P. falciparum 3D7 sub-lines using......-line was found to bind both CD31/PECAM1 and CD54/ICAM1 and to adhere twice as efficiently to human endothelial cells, compared to infected cells having only one PfEMP1 variant on the surface. These new results on PfEMP1 antigen expression indicate that a re-evaluation of the molecular mechanisms involved in P...

  18. Using multiple linear regression and physicochemical changes of amino acid mutations to predict antigenic variants of influenza A/H3N2 viruses.

    Science.gov (United States)

    Cui, Haibo; Wei, Xiaomei; Huang, Yu; Hu, Bin; Fang, Yaping; Wang, Jia

    2014-01-01

    Among human influenza viruses, strain A/H3N2 accounts for over a quarter of a million deaths annually. Antigenic variants of these viruses often render current vaccinations ineffective and lead to repeated infections. In this study, a computational model was developed to predict antigenic variants of the A/H3N2 strain. First, 18 critical antigenic amino acids in the hemagglutinin (HA) protein were recognized using a scoring method combining phi (ϕ) coefficient and information entropy. Next, a prediction model was developed by integrating multiple linear regression method with eight types of physicochemical changes in critical amino acid positions. When compared to other three known models, our prediction model achieved the best performance not only on the training dataset but also on the commonly-used testing dataset composed of 31878 antigenic relationships of the H3N2 influenza virus.

  19. Surface epitope localization and gene structure of a Babesia bovis 44-kilodalton variable merozoite surface antigen.

    Science.gov (United States)

    Jasmer, D P; Reduker, D W; Hines, S A; Perryman, L E; McGuire, T C

    1992-10-01

    Variation of Babesia bovis merozoite surface antigens occurs among geographic strains of the parasite. In this and a concurrent report, we investigate this variation at the gene and protein level. Using a monoclonal antibody (mAb 23/70.174), B. bovis gene sequences were identified that encoded a surface epitope of a 44-kDa merozoite surface antigen (MSA-2). This epitope is variably expressed among geographic isolates of B. bovis. Here, we describe the MSA-2 protein gene sequence, localize this surface epitope to a repeated amino acid sequence, and investigate the genomic organization of the gene in B. bovis strains from Mexico and Australia. The predicted protein sequence had hydrophobic regions at its amino and carboxy termini consistent with a signal peptide and a membrane anchor via glycosylphosphatidyl inositol, respectively. The surface epitope recognized by mAb 23/70.174 was localized within a 24-amino acid sequence which is repeated twice in tandem. Six different EcoRI bands hybridized to the MSA-2 gene sequence with varying intensities in genomic Southern blots of the homologous strain. Two of these appear to be alleles of the MSA-2 gene. Whereas 5' and 3' sequences of the MSA-2 gene sequence were detected in an Australia strain of B. bovis, internal gene sequences encoding the surface epitope were not. The 3' sequences of the MSA-2 gene also had significant sequence similarity with the MSA-1 gene of the Mexico strain B. bovis and a gene from the previously described BabR locus. These data indicate that the MSA-2 protein gene belongs to the BabR locus which encodes variable merozoite surface antigens.

  20. Hepatitis B Virus DNA in Blood Samples Positive for Antibodies to Core Antigen and Negative for Surface Antigen

    Science.gov (United States)

    Gutiérrez, C.; León, G.; Loureiro, C. L.; Uzcátegui, N.; Liprandi, F.; Pujol, F. H.

    1999-01-01

    Anti-hepatitis B core antigen (HBcAg)-positive hepatitis B surface antigen (HBsAg)-negative plasma samples from blood donors were tested by nested PCR. DNA positivity was more significantly associated with high levels of anti-HBcAg than with low levels of anti-HBsAg antibodies. Analysis of a dilution of anti-HBcAg antibodies might result in a more rational exclusion of anti-HBcAg-positive HBsAg-negative samples, reducing the number of donations discarded and enabling more countries to incorporate anti-HBcAg testing. PMID:10473534

  1. Major surface antigen, P30, of Toxoplasma gondii is anchored by a glycolipid

    Energy Technology Data Exchange (ETDEWEB)

    Nagel, S.D.; Boothroyd, J.C.

    1989-04-05

    P30, the major surface antigen of the parasitic protozoan Toxoplasma gondii, can be specifically labeled with (/sup 3/H)palmitic acid and with myo-(2-/sup 3/H)inositol. The fatty acid label can be released by treatment of P30 with phosphatidylinositol-specific phospholipase C (PI-PLC). Such treatment exposes an immunological cross-reacting determinant first described on Trypanosoma brucei variant surface glycoprotein. PI-PLC cleavage of intact parasites metabolically labeled with (/sup 35/S)methionine results in the release of intact P30 polypeptide in a form which migrates faster in polyacrylamide gel electrophoresis. These results argue that P30 is anchored by a glycolipid. Results from thin layer chromatography analysis of purified (/sup 3/H) palmitate-labeled P30 treated with PI-PLC, together with susceptibility to mild alkali hydrolysis and to cleavage with phospholipase A2, suggest that the glycolipid anchor of T. gondii P30 includes a 1,2-diacylglycerol moiety.

  2. JC Polyomavirus Infection Is Strongly Controlled by Human Leucocyte Antigen Class II Variants

    DEFF Research Database (Denmark)

    Sundqvist, Emilie; Buck, Dorothea; Warnke, Clemens

    2014-01-01

    mark infection occur only in 50-60% of infected individuals, and high JCV-antibody titers seem to increase the risk of developing PML. We here investigated the role of human leukocyte antigen (HLA), instrumental in immune defense in JCV antibody response. Anti-JCV antibody status, as a surrogate...... antibody response and human leucocyte antigens supports the notion that CD4+ T cells are crucial in the immune defence to JCV and lays the ground for risk stratification for PML and development of therapy and prevention.......JC polyomavirus (JCV) carriers with a compromised immune system, such as in HIV, or subjects on immune-modulating therapies, such as anti VLA-4 therapy may develop progressive multifocal leukoencephalopathy (PML) which is a lytic infection of oligodendrocytes in the brain. Serum antibodies to JCV...

  3. Recruitment of PfSET2 by RNA polymerase II to variant antigen encoding loci contributes to antigenic variation in P. falciparum.

    Directory of Open Access Journals (Sweden)

    Uchechi E Ukaegbu

    2014-01-01

    Full Text Available Histone modifications are important regulators of gene expression in all eukaryotes. In Plasmodium falciparum, these epigenetic marks regulate expression of genes involved in several aspects of host-parasite interactions, including antigenic variation. While the identities and genomic positions of many histone modifications have now been cataloged, how they are targeted to defined genomic regions remains poorly understood. For example, how variant antigen encoding loci (var are targeted for deposition of unique histone marks is a mystery that continues to perplex the field. Here we describe the recruitment of an ortholog of the histone modifier SET2 to var genes through direct interactions with the C-terminal domain (CTD of RNA polymerase II. In higher eukaryotes, SET2 is a histone methyltransferase recruited by RNA pol II during mRNA transcription; however, the ortholog in P. falciparum (PfSET2 has an atypical architecture and its role in regulating transcription is unknown. Here we show that PfSET2 binds to the unphosphorylated form of the CTD, a property inconsistent with its recruitment during mRNA synthesis. Further, we show that H3K36me3, the epigenetic mark deposited by PfSET2, is enriched at both active and silent var gene loci, providing additional evidence that its recruitment is not associated with mRNA production. Over-expression of a dominant negative form of PfSET2 designed to disrupt binding to RNA pol II induced rapid var gene expression switching, confirming both the importance of PfSET2 in var gene regulation and a role for RNA pol II in its recruitment. RNA pol II is known to transcribe non-coding RNAs from both active and silent var genes, providing a possible mechanism by which it could recruit PfSET2 to var loci. This work unifies previous reports of histone modifications, the production of ncRNAs, and the promoter activity of var introns into a mechanism that contributes to antigenic variation by malaria parasites.

  4. Artificial neural network accurately predicts hepatitis B surface antigen seroclearance.

    Directory of Open Access Journals (Sweden)

    Ming-Hua Zheng

    Full Text Available BACKGROUND & AIMS: Hepatitis B surface antigen (HBsAg seroclearance and seroconversion are regarded as favorable outcomes of chronic hepatitis B (CHB. This study aimed to develop artificial neural networks (ANNs that could accurately predict HBsAg seroclearance or seroconversion on the basis of available serum variables. METHODS: Data from 203 untreated, HBeAg-negative CHB patients with spontaneous HBsAg seroclearance (63 with HBsAg seroconversion, and 203 age- and sex-matched HBeAg-negative controls were analyzed. ANNs and logistic regression models (LRMs were built and tested according to HBsAg seroclearance and seroconversion. Predictive accuracy was assessed with area under the receiver operating characteristic curve (AUROC. RESULTS: Serum quantitative HBsAg (qHBsAg and HBV DNA levels, qHBsAg and HBV DNA reduction were related to HBsAg seroclearance (P<0.001 and were used for ANN/LRM-HBsAg seroclearance building, whereas, qHBsAg reduction was not associated with ANN-HBsAg seroconversion (P = 0.197 and LRM-HBsAg seroconversion was solely based on qHBsAg (P = 0.01. For HBsAg seroclearance, AUROCs of ANN were 0.96, 0.93 and 0.95 for the training, testing and genotype B subgroups respectively. They were significantly higher than those of LRM, qHBsAg and HBV DNA (all P<0.05. Although the performance of ANN-HBsAg seroconversion (AUROC 0.757 was inferior to that for HBsAg seroclearance, it tended to be better than those of LRM, qHBsAg and HBV DNA. CONCLUSIONS: ANN identifies spontaneous HBsAg seroclearance in HBeAg-negative CHB patients with better accuracy, on the basis of easily available serum data. More useful predictors for HBsAg seroconversion are still needed to be explored in the future.

  5. Partial determination of the primary structure of a variant surface glycoprotein from Trypanosoma equiperdum. Composition and location of a carbohydrate moiety.

    Science.gov (United States)

    Duvillier, G; Richet, C; Briand, G; Baltz, T; Degand, P

    1983-05-01

    Salivarian trypanosomes have the ability to evade the immune response of their hosts by the sequential expression of different cell surface glycoproteins. Among the isolated specific antigens from cloned variants of Trypanosoma equiperdum, a structural study was undertaken on two immunologically cross-reacting variant surface glycoproteins, and results concerning the basic antigenic type are reported. The glycoprotein was cleaved by cyanogen bromide, and amino acids of several purified fractions obtained by gel filtration chromatography of this cleavage mixture were sequenced by automated Edman degradation. Sequencing in particular allowed the identification of the N-terminal portion of the molecule (residues 1-74). Sugar compositions of the fractions have demonstrated the presence of at least two carbohydrate moieties in the glycoprotein. Using a subsequent enzymatic subcleavage we were able to locate the first glycosylation site in position 57. An important observation was that the first oligosaccharide identified was rich in mannose and devoid of galactose.

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

    Energy Technology Data Exchange (ETDEWEB)

    Tomavo, S.; Schwarz, R.T.; Dubremetz, J.F. (Institut National de la Recheche Medicale, Villeneuve d' Ascq (France))

    1989-10-01

    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 {sup 3}H-fatty acids, ({sup 3}H)ethanolamine, and ({sup 3}H)carbohydrates. Treatment of {sup 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. Method to conjugate polysaccharide antigens to surfaces for the detection of antibodies

    DEFF Research Database (Denmark)

    Boas, Ulrik; Lind, Peter; Riber, Ulla

    2014-01-01

    A new generic method for the conjugation of lipopolysaccharide (LPS)-derived polysaccharide antigens from gram-negative bacteria has been developed using Salmonella as a model. After removal of lipid A from the LPS by mild acidolysis, the polysaccharide antigen was conjugated to polystyrene...... against Salmonella typhimurium and Salmonella dublin. The presented method was compared with a similar method for conjugation of Salmonella polysaccharide antigens to surfaces. Here, the new method showed higher antigen coupling efficiency by detecting low concentrations of antibodies. Furthermore......, the polysaccharide-conjugated beads showed preserved bioactivity after 1 year of use....

  8. Sero-prevalence of hepatitis B surface antigen among pregnant ...

    African Journals Online (AJOL)

    There were no significant differences between prevalence recorded and agegroups but these were significant associations between risk factors namely tribal marks/tattooing, types of marriages and prevalence of hepatitis infection. The prevalence of hepatitis B antigen among pregnant women suggests that vertical ...

  9. Prevalence Of Hepatitis B Surface Antigen In Primary School ...

    African Journals Online (AJOL)

    Objective; To study the prevalence of hepatitis B virus infection among primary school children in Nnewi Subjects / Methods: Two hundred and thirty seven randomly selected schools children (127 females, 110 males) in Nnewi Nigeria were assayed for Hepatitis antigen (HbsAg) using ELISA technique. Results: Eighteen ...

  10. Surface Plasmon Resonance is an Analytically Sensitive Method for Antigen Profiling of Extracellular Vesicles.

    Science.gov (United States)

    Gool, Elmar L; Stojanovic, Ivan; Schasfoort, Richard B M; Sturk, Auguste; van Leeuwen, Ton G; Nieuwland, Rienk; Terstappen, Leon W M M; Coumans, Frank A W

    2017-10-01

    Identification, enumeration, and characterization of extracellular vesicles (EVs) are hampered by the small size of EVs, a low refractive index, and low numbers of antigens on their surface. We investigated the potential of a 48-multiplex surface plasmon resonance imaging (SPRi) system to perform EV phenotyping. Antigen surface density of 11 antigens was measured on the human breast cancer cell lines HS578T, MCF7, and SKBR3 and their EVs by use of both SPRi and the widely used flow cytometry (FCM). For cells, the SPRi and FCM signals for antigen exposure correlated (RHS578T cells2 = 0.66, RMCF7 cells2 = 0.78, RSKBR3 cells2 = 0.60). With regard to EVs, SPRi detected 31 out of 33 tested antibody-EV pairs, whereas our flow cytometer detected 5 antibody-EV pairs because of high blank and isotype control signals. For HS578T-derived EVs, the SPRi and FCM signals correlated (R2HS578T EVs = 0.98). However, on MCF7- and SKBR3-derived EVs, insufficient antigens were detected by our flow cytometer. To confirm that the SPRi responses correlated with mean antigen density on EVs, the SPRi responses of EVs were correlated with antigen density on parental cells as measured by FCM (RHS578T2 = 0.77, RMCF72 = 0.49, RSKBR32 = 0.52). SPRi responses correlate with mean antigen density. Moreover, SPRi detects lower antigen-exposure levels than FCM because SPRi measures an ensemble of EVs binding to the sensor surface, whereas FCM detects antigens of single EV. © 2017 American Association for Clinical Chemistry.

  11. The immune responses to bacterial antigens encountered in vivo at mucosal surfaces.

    OpenAIRE

    Dougan, G; Ghaem-Maghami, M; Pickard, D; Frankel, G; Douce, G; Clare, S; Dunstan, S; Simmons, C

    2000-01-01

    Mammals have evolved a sophisticated immune system for handling antigens encountered at their mucosal surfaces. The way in which mucosally delivered antigens are handled influences our ability to design effective mucosal vaccines. Live attenuated derivatives of pathogens are one route towards the development of mucosal vaccines. However, some molecules, described as mucosal immunogens, are inherently immunogenic at mucosal surfaces. Studies on mucosal immunogens may facilitate the identificat...

  12. Expression of a cell surface immobilization antigen during serotype transformation in Tetrahymena thermophila.

    OpenAIRE

    Williams, N. E.; Doerder, F. P.; Ron, A

    1985-01-01

    A temperature shift from 40 to 28 degrees C rapidly induced expression of a specific immobilization antigen at the cell surface in Tetrahymena thermophila. This transformation was inhibited by actinomycin D and cycloheximide but not by colchicine or cytochalasin B. The major surface antigen expressed at 28 degrees C in cells homozygous for the SerH3 allele was partially purified, and an antiserum against this preparation was raised in rabbits. Electrophoresis, immunoblot, and [35S]methionine ...

  13. JC polyomavirus infection is strongly controlled by human leucocyte antigen class II variants.

    Directory of Open Access Journals (Sweden)

    Emilie Sundqvist

    2014-04-01

    Full Text Available JC polyomavirus (JCV carriers with a compromised immune system, such as in HIV, or subjects on immune-modulating therapies, such as anti VLA-4 therapy may develop progressive multifocal leukoencephalopathy (PML which is a lytic infection of oligodendrocytes in the brain. Serum antibodies to JCV mark infection occur only in 50-60% of infected individuals, and high JCV-antibody titers seem to increase the risk of developing PML. We here investigated the role of human leukocyte antigen (HLA, instrumental in immune defense in JCV antibody response. Anti-JCV antibody status, as a surrogate for JCV infection, were compared to HLA class I and II alleles in 1621 Scandinavian persons with MS and 1064 population-based Swedish controls and associations were replicated in 718 German persons with MS. HLA-alleles were determined by SNP imputation, sequence specific (SSP kits and a reverse PCR sequence-specific oligonucleotide (PCR-SSO method. An initial GWAS screen displayed a strong HLA class II region signal. The HLA-DRB1*15 haplotype was strongly negatively associated to JCV sero-status in Scandinavian MS cases (OR = 0.42, p = 7×10(-15 and controls (OR = 0.53, p = 2×10(-5. In contrast, the DQB1*06:03 haplotype was positively associated with JCV sero-status, in Scandinavian MS cases (OR = 1.63, p = 0.006, and controls (OR = 2.69, p = 1×10(-5. The German dataset confirmed these findings (OR = 0.54, p = 1×10(-4 and OR = 1.58, p = 0.03 respectively for these haplotypes. HLA class II restricted immune responses, and hence CD4+ T cell immunity is pivotal for JCV infection control. Alleles within the HLA-DR1*15 haplotype are associated with a protective effect on JCV infection. Alleles within the DQB1*06:03 haplotype show an opposite association. These associations between JC virus antibody response and human leucocyte antigens supports the notion that CD4+ T cells are crucial in the immune defence to JCV and

  14. Expression of a cell surface immobilization antigen during serotype transformation in Tetrahymena thermophila.

    Science.gov (United States)

    Williams, N E; Doerder, F P; Ron, A

    1985-08-01

    A temperature shift from 40 to 28 degrees C rapidly induced expression of a specific immobilization antigen at the cell surface in Tetrahymena thermophila. This transformation was inhibited by actinomycin D and cycloheximide but not by colchicine or cytochalasin B. The major surface antigen expressed at 28 degrees C in cells homozygous for the SerH3 allele was partially purified, and an antiserum against this preparation was raised in rabbits. Electrophoresis, immunoblot, and [35S]methionine incorporation studies are reported which support the conclusion that the H3 antigen is an acidic protein with an Mr of approximately 52,000 daltons. An induced synthesis of the H3 immobilization antigen was detected within 30 min after a shift from 40 to 28 degrees C. This protein appeared to be synthesized in the microsomal fraction and transferred without cleavage to the cell surface, where it was inserted first into nonciliated regions.

  15. Lenalidomide enhances the function of chimeric antigen receptor T cells against the epidermal growth factor receptor variant III by enhancing immune synapses.

    Science.gov (United States)

    Kuramitsu, S; Ohno, M; Ohka, F; Shiina, S; Yamamichi, A; Kato, A; Tanahashi, K; Motomura, K; Kondo, G; Kurimoto, M; Senga, T; Wakabayashi, T; Natsume, A

    2015-10-01

    The epidermal growth factor receptor variant III (EGFRvIII) is exclusively expressed on the cell surface in ~50% of glioblastoma multiforme (GBM). This variant strongly and persistently activates the phosphatidylinositol 3-kinase-Akt signaling pathway in a ligand-independent manner resulting in enhanced tumorigenicity, cellular motility and resistance to chemoradiotherapy. Our group generated a recombinant single-chain variable fragment (scFv) antibody specific to the EGFRvIII, referred to as 3C10-scFv. In the current study, we constructed a lentiviral vector transducing the chimeric antigen receptor (CAR) that consisted of 3C10-scFv, CD3ζ, CD28 and 4-1BB (3C10-CAR). The 3C10-CAR-transduced peripheral blood mononuclear cells (PBMCs) and CD3(+) T cells specifically lysed the glioma cells that express EGFRvIII. Moreover, we demonstrated that CAR CD3(+) T cells migrated to the intracranial xenograft of GBM in the mice treated with 3C10-CAR PBMCs. An important and novel finding of our study was that a thalidomide derivative lenalidomide induced 3C10-CAR PBMC proliferation and enhanced the persistent antitumor effect of the cells in vivo. Lenalidomide also exhibited enhanced immunological synapses between the effector cells and the target cells as determined by CD11a and F-actin polymerization. Collectively, lentiviral-mediated transduction of CAR effectors targeting the EGFRvIII showed specific efficacy, and lenalidomide even intensified CAR cell therapy by enhanced formation of immunological synapses.

  16. CARbodies: Human Antibodies Against Cell Surface Tumor Antigens Selected From Repertoires Displayed on T Cell Chimeric Antigen Receptors

    Directory of Open Access Journals (Sweden)

    Vanesa Alonso-Camino

    2013-01-01

    Full Text Available A human single-chain variable fragment (scFv antibody library was expressed on the surface of human T cells after transduction with lentiviral vectors (LVs. The repertoire was fused to a first-generation T cell receptor ζ (TCRζ-based chimeric antigen receptor (CAR. We used this library to isolate antibodies termed CARbodies that recognize antigens expressed on the tumor cell surface in a proof-of-principle system. After three rounds of activation-selection there was a clear repertoire restriction, with the emergence dominant clones. The CARbodies were purified from bacterial cultures as soluble and active proteins. Furthermore, to validate its potential application for adoptive cell therapy, human T cells were transduced with a LV encoding a second-generation costimulatory CAR (CARv2 bearing the selected CARbodies. Transduced human primary T cells expressed significant levels of the CARbodies-based CARv2 fusion protein on the cell surface, and importantly could be specifically activated, after stimulation with tumor cells. This approach is a promising tool for the generation of antibodies fully adapted to the display format (CAR and the selection context (cell synapse, which could extend the scope of current adoptive cell therapy strategies with CAR-redirected T cells.

  17. Production of immunologically active surface antigens of hepatitis B virus by Escherichia coli.

    Science.gov (United States)

    MacKay, P; Pasek, M; Magazin, M; Kovacic, R T; Allet, B; Stahl, S; Gilbert, W; Schaller, H; Bruce, S A; Murray, K

    1981-01-01

    Several plasmids have been constructed which direct the synthesis of hepatitis B virus surface antigens in Escherichia coli either as the native polypeptide or fused to other plasmid encoded polypeptides. When injected into rabbits, extracts from bacteria carrying some of these plasmids induced the synthesis of antibodies to the antigens even though the extracts did not give satisfactory positive results in radioimmunoassay for them. Either the NH2-terminal segment or the COOH-terminal segment of the surface antigens alone was sufficient to elicit the immune response, but antibodies against the two segments showed different specificities. The results emphasize the value of an in vivo assay for the presence of antigens in crude cell extracts and illustrate the feasibility of this type of screening with laboratory animals. PMID:6170067

  18. Transgenic Expression of IL15 Improves Antiglioma Activity of IL13Rα2-CAR T Cells but Results in Antigen Loss Variants.

    Science.gov (United States)

    Krenciute, Giedre; Prinzing, Brooke L; Yi, Zhongzhen; Wu, Meng-Fen; Liu, Hao; Dotti, Gianpietro; Balyasnikova, Irina V; Gottschalk, Stephen

    2017-07-01

    Glioblastoma (GBM) is the most aggressive primary brain tumor in adults and is virtually incurable with conventional therapies. Immunotherapy with T cells expressing GBM-specific chimeric antigen receptors (CAR) is an attractive approach to improve outcomes. Although CAR T cells targeting GBM antigens, such as IL13 receptor subunit α2 (IL13Rα2), HER2, and EGFR variant III (EGFRvIII), have had antitumor activity in preclinical models, early-phase clinical testing has demonstrated limited antiglioma activity. Transgenic expression of IL15 is an appealing strategy to enhance CAR T-cell effector function. We tested this approach in our IL13Rα2-positive glioma model in which limited IL13Rα2-CAR T-cell persistence results in recurrence of antigen-positive gliomas. T cells were genetically modified with retroviral vectors encoding IL13Rα2-CARs or IL15 (IL13Rα2-CAR.IL15 T cells). IL13Rα2-CAR.IL15 T cells recognized glioma cells in an antigen-dependent fashion, had greater proliferative capacity, and produced more cytokines after repeated stimulations in comparison with IL13Rα2-CAR T cells. No autonomous IL13Rα2-CAR.IL15 T-cell proliferation was observed; however, IL15 expression increased IL13Rα2-CAR T-cell viability in the absence of exogenous cytokines or antigen. In vivo, IL13Rα2-CAR.IL15 T cells persisted longer and had greater antiglioma activity than IL13Rα2-CAR T cells, resulting in a survival advantage. Gliomas recurring after 40 days after T-cell injection had downregulated IL13Rα2 expression, indicating that antigen loss variants occur in the setting of improved T-cell persistence. Thus, CAR T cells for GBM should not only be genetically modified to improve their proliferation and persistence, but also to target multiple antigens.Summary: Glioblastoma responds imperfectly to immunotherapy. Transgenic expression of IL15 in T cells expressing CARs improved their proliferative capacity, persistence, and cytokine production. The emergence of antigen loss

  19. Relation between laboratory test results and histological hepatitis activity in individuals positive for hepatitis B surface antigen and antibodies to hepatitis B e antigen

    NARCIS (Netherlands)

    ter Borg, F.; ten Kate, F. J.; Cuypers, H. T.; Leentvaar-Kuijpers, A.; Oosting, J.; Wertheim-van Dillen, P. M.; Honkoop, P.; Rasch, M. C.; de Man, R. A.; van Hattum, J.; Chamuleau, R. A.; Reesink, H. W.; Jones, E. A.

    1998-01-01

    BACKGROUND: Hepatitis B surface antigen (HBsAg) and antibodies to hepatitis B e antigen (anti-HBe) commonly coexist, and laboratory tests are often requested to assess histological hepatitis activity. An optimum panel of tests has not been found and the usefulness of hepatitis B virus (HBV) DNA

  20. Quasispecies variant of pre-S/S gene in HBV-related hepatocellular carcinoma with HBs antigen positive and occult infection.

    Science.gov (United States)

    Hatazawa, Yuri; Yano, Yoshihiko; Okada, Rina; Tanahashi, Toshihito; Hayashi, Hiroki; Hirano, Hirotaka; Minami, Akihiro; Kawano, Yuki; Tanaka, Motofumi; Fukumoto, Takumi; Murakami, Yoshiki; Yoshida, Masaru; Hayashi, Yoshitake

    2018-01-01

    Hepatocellular carcinoma (HCC) can develop in patients who are negative for the hepatitis B surface antigen (HBsAg) in serum but positive for hepatitis B virus (HBV) DNA in the liver, referred to as occult HBV infection (OBI). Previous reports showed that HBV variants in OBI-related HCC are different from those in HBsAg-positive HCC. In the present study, HBV quasispecies based on the pre-S/S gene in OBI-related HCC patients were examined by high throughput sequencing and compared with those in HBsAg-positive HCC. Nineteen tissue samples (9 OBI-related and 10 HBsAg-positive non-cancerous tissues) were collected at the time of surgery at Kobe University Hospital. The quasispecies with more than 1% variation in the pre-S/S region were isolated and analysed by ultra-deep sequencing. There were no significant differences in the major HBV populations, which exhibit more than 20% variation within the entire pre-S/S region, between OBI-related HCC and HBsAg-positive HCC. However, the prevalences of major populations with pre-S2 region mutations and of minor populations with polymerized human serum albumin-binding domain mutations were significantly higher in OBI-related HCC than in HBsAg-positive HCC. Moreover, the major variant populations associated with the B-cell epitope, located within the pre-S1 region, and the a determinant domain, located in the S region, were detected frequently in HBsAg-positive HCC. The minor populations of variants harbouring the W4R, L30S, Q118R/Stop, N123D and S124F/P mutations in the pre-S region and the L21F/S and L42F/S mutations in the S region were detected more frequently in OBI-related HCC than in HBsAg-positive HCC. Ultra-deep sequencing revealed that the B-cell epitope domain in the pre-S1 region and alpha determinant domain in the S region were variable in HBsAg-positive HCC, although the quasispecies associated with the pre-S2 region were highly prevalent in OBI-related HCC. Ref: R000034382/UMIN000030113; Retrospectively

  1. The Effect of Superparamagnetic Iron Oxide Nanoparticle Surface Charge on Antigen Cross-Presentation

    Science.gov (United States)

    Mou, Yongbin; Xing, Yun; Ren, Hongyan; Cui, Zhihua; Zhang, Yu; Yu, Guangjie; Urba, Walter J.; Hu, Qingang; Hu, Hongming

    2017-01-01

    Magnetic nanoparticles (NPs) of superparamagnetic iron oxide (SPIO) have been explored for different kinds of applications in biomedicine, mechanics, and information. Here, we explored the synthetic SPIO NPs as an adjuvant on antigen cross-presentation ability by enhancing the intracellular delivery of antigens into antigen presenting cells (APCs). Particles with different chemical modifications and surface charges were used to study the mechanism of action of antigen delivery. Specifically, two types of magnetic NPs, γFe2O3/APTS (3-aminopropyltrimethoxysilane) NPs and γFe2O3/DMSA (meso-2, 3-Dimercaptosuccinic acid) NPs, with the same crystal structure, magnetic properties, and size distribution were prepared. Then, the promotion of T-cell activation via dendritic cells (DCs) was compared among different charged antigen coated NPs. Moreover, the activation of the autophagy, cytosolic delivery of the antigens, and antigen degradation mediated by the proteasome and lysosome were measured. Our results indicated that positive charged γFe2O3/APTS NPs, but not negative charged γFe2O3/DMSA NPs, enhanced the cross-presentation ability of DCs. Increased cross-presentation ability induced by γFe2O3/APTS NPs was associated with increased cytosolic antigen delivery. On the contrary, γFe2O3/DMSA NPs was associated with rapid autophagy. Overall, our results suggest that antigen delivered in cytoplasm induced by positive charged particles is beneficial for antigen cross-presentation and T-cell activation. NPs modified with different chemistries exhibit diverse biological properties and differ greatly in their adjuvant potentials. Thus, it should be carefully considered many different effects of NPs to design effective and safe adjuvants.

  2. Mendelian and non-mendelian mutations affecting surface antigen expression in Paramecium tetraurelia

    Energy Technology Data Exchange (ETDEWEB)

    Epstein, L.M.; Forney, J.D.

    1984-08-01

    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.

  3. Mendelian and non-mendelian mutations affecting surface antigen expression in Paramecium tetraurelia.

    Science.gov (United States)

    Epstein, L M; Forney, J D

    1984-08-01

    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.

  4. Correlation of presentation of insulin with surface I-Ad and A alpha and A beta mRNA expression by cloned B lymphoma hybridoma variants.

    Science.gov (United States)

    Bernard, N F; Reid, P C; Phillips, M L; Delovitch, T L

    1988-10-01

    We investigated the relationship between Ia expression and antigen presentation in cloned B cells, using variants of TA3 antigen presenting cells. Two TA3 subclones were selected as high presenters and 5 as low presenters of insulin to pork insulin/I-Ad restricted T cells. All TA3 subclones express the surface I-Ak, I-Ek, I-Ad and I-Ed Ia antigens characteristic of the parental cell line. However, surface I-Ad levels correlated best with the ability to present insulin, since high presenters express 2- to 4-fold more I-Ad than low presenters. High presenters possess 2- to 4-times more A alpha and A beta Ia mRNA than low presenters and also transcribe these mRNAs 2- to 5-fold faster than most low presenters. Thus, the correlation noted between I-Ad surface density and capacity to present insulin by our panel of TA3 variants is regulated at the level of transcription and not translation of I-Ad specific mRNAs.

  5. INFLUENCE OF DIFFERENT SOIL TILLAGE VARIANTS ON QUALITY OF SEED DISTRIBUTION QUALITY PER SURFACE AND DEPTH

    Directory of Open Access Journals (Sweden)

    Luka Šumanovac

    2000-12-01

    Full Text Available The paper aims to present influence of different soil tillage variants on quality of seed distribution per surface and depth. A sowing experiment was set up in three soil tillage variants: a repeated disking, b conventional tillage, c ploughing + harrowing. Soil levelling, effects of longitudinal and transverzal distribution as well as seed distribution per depth were analyzed by the investigation. The research results indicate the fact that no soil tillage variant is characterized by achieving satisfactory sowing quality . It means that reduced soil tillage application is justifiable in terms of energetic aspect, work productivity increase, sowing performance in optimal agricultural terms etc., compared to conventional soil tillage.

  6. Antigenic variants of influenza A virus, PR8 strain. I. Their development during serial passage in the lungs of partially immune mice.

    Science.gov (United States)

    GERBER, P; LOOSLI, C G; HAMBRE, D

    1955-06-01

    Antigenically different strains of mouse-adapted PR8 influenza A virus have been produced by 17 serial passages of the virus in the lungs of mice immunized with the homologous agent. Comparative serological tests show that the variant strains share antigenic components with the parent strain but the dominant antigen is different. By means of antibody absorption it was shown that the "new" antigenic component of the variant was already present in minor amounts up to the eighth passage and thereafter gained prominence with continued passage in vaccinated mice. Groups of mice vaccinated with either the PR8-S or T(21) virus and having comparable antibody titers showed no growth of virus in the lungs following aid-borne challenge with homologous strains. On the other hand, following heterologous air-borne challenge no deaths occurred, but virus grew in the lungs of both groups of vaccinated mice. Almost unrestricted virus multiplication took place in the lungs of mice vaccinated with the parent strain and challenged with the PR8-T(21) virus which resulted in extensive consolidation. Less virus grew in the lungs of the mice vaccinated with the variant strains and challenged with the PR8-S virus. In these animals only microscopic evidence of changes due to virus growth in the lungs was observed. The successful serial passage of PR8 influenza A virus in immunized animals was dependent on the initial selection of mice with uniformly low H.I. antibody titers as determined on tail blood, and the intranasal instillation of sufficient virus to favor the survival of those virus particles least related to the antibodies present. The epidemiological implications of these observations are discussed briefly.

  7. Trypanosoma evansi: identification and characterization of a variant surface glycoprotein lacking cysteine residues in its C-terminal domain.

    Science.gov (United States)

    Jia, Yonggen; Zhao, Xinxin; Zou, Jingru; Suo, Xun

    2011-01-01

    African trypanosomes are flagellated unicellular parasites which proliferate extracellularly in the mammalian host blood-stream and tissue spaces. They evade the hosts' antibody-mediated lyses by sequentially changing their variant surface glycoprotein (VSG). VSG tightly coats the entire parasite body, serving as a physical barrier. In Trypanosoma brucei and the closely related species Trypanosoma evansi, Trypanosoma equiperdum, each VSG polypeptide can be divided into N- and C-terminal domains, based on cysteine distribution and sequence homology. N-terminal domain, the basis of antigenic variation, is hypervariable and contains all the exposed epitopes; C-terminal domain is relatively conserved and a full set of four or eight cysteines were generally observed. We cloned two genes from two distinct variants of T. evansi, utilizing RT-PCR with VSG-specific primers. One contained a VSG type A N-terminal domain followed a C-terminal domain lacking cysteine residues. To confirm that this gene is expressed as a functional VSG, the expression and localization of the corresponding gene product were characterized using Western blotting and immunofluorescent staining of living trypanosomes. Expression analysis showed that this protein was highly expressed, variant-specific, and had a ubiquitous cellular surface localization. All these results indicated that it was expressed as a functional VSG. Our finding showed that cysteine residues in VSG C-terminal domain were not essential; the conserved C-terminal domain generally in T. brucei like VSGs would possibly evolve for regulating the VSG expression. Copyright © 2010 Elsevier Inc. All rights reserved.

  8. Identification of Babesia bigemina infected erythrocyte surface antigens containing epitopes conserved among strains.

    Science.gov (United States)

    Shompole, S; McElwain, T F; Jasmer, D P; Hines, S A; Katende, J; Musoke, A J; Rurangirwa, F R; McGuire, T C

    1994-03-01

    The presence of previously uncharacterized antigens (new antigens) on the surface of intact erythrocytes infected with three strains of Babesia bigemina from Kenya and one each from Puerto Rico, Mexico, St. Croix, and Texcoco-Mexico was demonstrated by indirect immunofluorescent antibody (IFA) reactions. These antigens were not strain specific because antibodies in bovine immune serum to either the Mexico or Kenya isolates reacted with all seven strains tested. Homologous and heterologous immune serum antibodies bound a maximum of 83% and 55%, respectively, of intact erythrocytes infected with the Kenya-Ngong strain but not uninfected erythrocytes. Both sera caused agglutination of only infected erythrocytes. Antibodies eluted from the surface of glutaraldehyde (0.25%) fixed infected erythrocytes had IFA reaction patterns among strains similar to those of immune sera before elution. Eluted antibodies were used to determine if these antigens were protein and encoded by B. bigemina. Eluted antibodies bound seven parasite-encoded proteins of 240, 220, 66, 62, 58, 52 and 38 kDa in an erythrocyte surface-specific immunoprecipitation reaction of 35S-methionine labelled proteins. It was concluded that the surface of B. bigemina infected erythrocytes had parasite-encoded proteins and that these proteins had surface exposed epitopes that were conserved among the seven strains examined which were from two continents.

  9. Characterization of surface associated antigens of axenic Giardia lamblia trophozoites & their recognition by human sera.

    Science.gov (United States)

    Kum, K; Khanna, R; Vinayak, V K

    1991-01-01

    Two surface associated antigens (GLSA-82 and GLSA-56) of axenically grown G. lamblia trophozoites (PI strain) were affinity purified from its sonic extract. Both GLSA-82 and GLSA-56 were heat labile, sensitive to treatment with pronase, trypsin and were also sodium metaperiodate modifiable as assessed by micro ELISA. Lectin binding studies revealed that GLSA-82 specifically bound concanavalin A and pokeweed mitogen, and had alpha-methyl mannoside and n-acetyl-B-d-glucosamine sugar moieties. However, GLSA-56 selectively bound Ricinus communis agglutinin and phytohaemagglutinin, and had B-d-galactose and n-acetyl-B-d-galastosamine as sugar moieties. Human sera obtained during acute G. lamblia infection recognised GLSA-82 and GLSA-56 antigens. However, the antibody levels to GLSA-82 were significantly lower (P less than 0.05) during active giardiasis infection. Such surface associated antigens may be target of antiparasitic immune responses and thus, may modulate disease processes.

  10. Utility of immunohistochemical analysis of KAI1, epithelial-specific antigen, and epithelial-related antigen for distinction of chromophobe renal cell carcinoma, an eosinophilic variant from renal oncocytoma.

    Science.gov (United States)

    Ohe, Chisato; Kuroda, Naoto; Takasu, Kosho; Senzaki, Hideto; Shikata, Nobuaki; Yamaguchi, Tadanori; Miyasaka, Chika; Nakano, Yorika; Sakaida, Noriko; Uemura, Yoshiko

    2012-06-01

    Distinction of renal oncocytoma (RO) from chromophobe renal cell carcinoma (ChRCC) is important because their clinical behavior is different. As part of a search for the best available immunohistochemical markers to distinguish ChRCC from RO, we investigated the immunohistochemical profiles of these tumors. We selected 30 renal tumors consisting of ChRCC, typical variant (n = 14), ChRCC, eosinophilic variant (n = 6), and RO (n = 10). Their expression of cytokeratin (CK) 7, KAI1, epithelial-specific antigen (ESA), epithelial-related antigen (ERA), Claudin- 7, and Claudin-8 was studied using an autostainer. Immunoreactivity was assessed based on a combined score of the extent and intensity of staining. Compared to RO, a significantly higher percentage of the total ChRCCs stained positive for CK7 (85% vs. 10%, respectively), KAI1 (90% vs. 10%), ESA (95% vs. 10%), ERA (95% vs. 10%), and Claudin-7 (95% vs. 20%) (P < 0.001). Additionally, there was a significant difference between the percentage of ChRCC eosinophilic variant (ChRCC-E) and RO that stained positive for KAI1 (100% vs. 10%, respectively), ESA (83% vs. 10%), and ERA (83% vs. 10%) (P < 0.001). We recommend immunohistochemical analysis of KAI1, ESA, and ERA to distinguish ChRCC-E from RO.

  11. Microfluidic cell surface antigen expression analysis using a single antibody type.

    Science.gov (United States)

    Zhang, Ye; Pappas, Dimitri

    2016-02-21

    Antigen expression plays a significant role in clinical studies, pathology, biology and chemistry. The type and degree of antigen expression can provide information for disease diagnosis/monitoring and is used for phenotype analysis of cells. In this work, an affinity capture method was developed to capture cells based on antigen expression differences in a single microfluidic chip. Microfluidic chips with two affinity regions-at different antibody concentrations-captured two cell types based on differences in the expression of a single antigen. Using herringbone-modified capture channels, a separation purity of 95% and a capture efficiency of 15% were achieved under continuous-flow conditions. We observed that the capture ratio of Ramos B lymphocytes and HuT 78 T lymphocytes matched the expression ratio of CD71 for the two cell lines (R(2) = 0.94). To further validate our analytical method, Ramos B lymphocytes were spiked into blood samples to demonstrate performance with a complex sample. Expression ratios matched conventional flow cytometry measurements over a 40-fold difference, and the sample enrichment was 9.5×. This method has proven to be a robust system to measure the differences in antigen expression, and can be used to distinguish cells without having a unique surface antigen if the expression level is sufficiently high in one cell type.

  12. Optimisation of Production of Immunogenic Malaria Surface Antigens in Plants

    OpenAIRE

    KARTIKA MAILLIKA SETYABUDI

    2017-01-01

    Vaccines against malaria could serve a useful purpose in malaria control, prevention and eradication efforts. P. falciparum merozoite surface proteins (MSP): MSP119, MSP4 and MSP5 are promising targets to establish a protective immune response against malaria. Plant-based expression system presents potential as a cost-effective, scalable and safe production platform. This study provides evidence of the feasibility of plants to produce immunogenic P. falciparum MSP119, MSP4, MSP5. The demonstr...

  13. Recombination-driven generation of the largest pathogen repository of antigen variants in the protozoan Trypanosoma cruzi.

    Science.gov (United States)

    Weatherly, D Brent; Peng, Duo; Tarleton, Rick L

    2016-09-13

    The protozoan parasite Trypanosoma cruzi, causative agent of Chagas disease, depends upon a cell surface-expressed trans-sialidase (ts) to avoid activation of complement-mediated lysis and to enhance intracellular invasion. However these functions alone fail to account for the size of this gene family in T. cruzi, especially considering that most of these genes encode proteins lacking ts enzyme activity. Previous whole genome sequencing of the CL Brener clone of T. cruzi identified ~1400 ts variants, but left many partially assembled sequences unannotated. In the current study we reevaluated the trans-sialidase-like sequences in this reference strain, identifying an additional 1779 full-length and partial ts genes with their important features annotated, and confirming the expression of previously annotated "pseudogenes" and newly annotated ts family members. Multiple EM for Motif Elicitation (MEME) analysis allowed us to generate a model T. cruzi ts (TcTS) based upon the most conserved motif patterns and demonstrated that a common motif order is highly conserved among ts family members. Using a newly developed pipeline for the analysis of recombination within large gene families, we further demonstrate that TcTS family members are undergoing frequent recombination, generating new variants from the thousands of functional and non-functional ts gene segments but retaining the overall structure of the core TcTS family members. The number and variety as well as high recombination frequency of TcTS family members supports strong evolutionary pressure, probably exerted by immune selection, for continued variation in ts sequences in T. cruzi, and thus for a unique immune evasion mechanism for the large ts gene family.

  14. Yeast expressing hepatitis B virus surface antigen determinants on its surface: Implications for a possible oral vaccine

    NARCIS (Netherlands)

    Schreuder, M.P.; Deen, C.; Boersma, W.J.A.; Pouwels, P.H.; Klis, F.M.

    1996-01-01

    The two major hydrophilic regions of the hepatitis B virus surface antigen (HBsAg) have been expressed in the outer mannoprotein layer of the cell wall of 'Bakers Yeast', Saccharomyces cerevisiae, by fusing them between the yeast invertase signal sequence and the yeast α-agglutinin carboxyterminal

  15. Prevalence of Hepatitis-B Surface Antigen (HbsAg), Hepatitis C ...

    African Journals Online (AJOL)

    The prevalence of Hepatitis-B surface antigen (HBsAg), Hepatitis C virus (HCV) and Human immunodeficiency virus (HIV) was determined among apparently healthy male blood donors in Aminu Kano Teaching Hospital, Kano, between January and December, 2002. A total of 2,288 blood samples from the blood donors ...

  16. Hepatitis B-Surface Antigen In Ascitic Fluid Of Patients With Chronic ...

    African Journals Online (AJOL)

    A prospective evaluation of eleven consecutive cases of chronic liver disease over a twelve-month period was carried out clinically and ultrasonographically. By the use of the method of reverse passive haemagglutination, sera and ascitic fluid of the patients were tested for the presence of the Hepatitis B surface antigen.

  17. Follow-up of blood donors positive for hepatitis B surface antigen

    NARCIS (Netherlands)

    Reesink, H. W.; Wesdorp, I. C.; Grijm, R.; Hengeveld, P.; Jöbsis, A. C.; Aay, C.; Reerink-Brongers, E. E.

    1980-01-01

    From 1973 to 1977 in Amsterdam the incidence of hepatitis B surface antigen (HBsAg) in blood donations from new donors was 0.224 and from known donors 0.034%. 65 donors, previously found positive for HBsAg, were re-examined. Persistence of HBsAg in new donors (28 of 31) occurred significantly (p

  18. 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 ...

  19. Hepatitis B virus surface antigen and antibody markers in children at ...

    African Journals Online (AJOL)

    Hepatitis B virus surface antigen and antibody markers in children at a major paediatric hospital after the pentavalent DTP-HBV-Hib vaccination. ... Ghana Medical Journal ... Abstract. Objectives: The knowledge about outcomes of infant vaccination against HBV infections using the DPT-HepB-Hib vaccine in Ghana is limited.

  20. Surface antigen-negative hepatitis B virus infection in Dutch blood donors

    NARCIS (Netherlands)

    Lieshout-Krikke, R. W.; Molenaar-de Backer, M. W. A.; van Swieten, P.; Zaaijer, H. L.

    2014-01-01

    Hepatitis B virus (HBV) surface antigen (HBsAg) is a reliable marker for HBV infection, but HBsAg-negative forms of HBV infection occur. The introduction of HBV DNA screening of Dutch blood donors, which were not preselected for absence of HBV core antibodies, enabled the characterization of

  1. Sero-prevalence of Hepatitis B Surface Antigen (HBsAg), in Sexually ...

    African Journals Online (AJOL)

    Sero-prevalence of Hepatitis B Surface Antigen (HBsAg), in Sexually Transmitted Disease Patients. ... partners in the past 6 months, a history of a number of episodes of STDs, history of heterosexual exposure to partners at risk, for example prostitutes; a history of symptoms of an STD at the commencement of the study.

  2. Purification and characterization of a major human Pneumocystis carinii surface antigen

    DEFF Research Database (Denmark)

    Lundgren, B; Lipschik, G Y; Kovacs, J A

    1991-01-01

    . To evaluate humoral immune responses to the human P. carinii protein, an enzyme-linked immunosorbent assay using purified protein was developed. Some, but not all, patients who subsequently developed P. carinii pneumonia demonstrated a serum antibody response to the surface antigen. Nearly all subjects...

  3. Immunization of cattle with recombinant Babesia bovis merozoite surface antigen-1.

    Science.gov (United States)

    Hines, S A; Palmer, G H; Jasmer, D P; Goff, W L; McElwain, T F

    1995-01-01

    Cattle immunized with a recombinant merozoite surface antigen-1 molecule (MSA-1) produced high-titered antibody that reacted with the surface of the parasite and neutralized merozoite infectivity in vitro. However, recombinant MSA-1 immunization did not confer protection against challenge with virulent Babesia bovis. These results indicate that antibody-mediated neutralization of merozoite infectivity in vitro, at least for MSA-1-specific antibody, does not reflect in vivo protective immunity to babesiosis.

  4. Virological Determinants of Spontaneous Postpartum e Antigen Seroconversion and Surface Antigen Seroclearance in Pregnant Women Infected with Hepatitis B Virus.

    Science.gov (United States)

    Hu, Yali; Feng, Zhenhua; Liu, Jingli; Chen, Jie; Zhang, Shu; Zhou, Yi-Hua

    2016-04-01

    We investigated the virological factors predicting spontaneous postpartum hepatitis B e antigen (HBeAg) seroconversion and hepatitis B surface antigen (HBsAg) seroclearance in pregnant women infected with hepatitis B virus (HBV). We invited 419 HBV infected women whose sera had been collected during their pregnancy from August 2002-July 2004 and archived at -30°C, to participate the follow-up in October 2009-March 2010. Various virological factors were determined and compared in women with or without the seroconversion and seroclearance. A total of 264 (63.0%) antiviral naive women participated in the follow-up with an average observation period of 6.4 years (5.4-7.4). Of 76 women who were HBeAg positive during pregnancy, 42 (55.3%) seroconverted to anti-HBe during follow-up. Compared to pregnant women with HBV DNA ≥3 × 10(7) IU/mL or HBeAg ≥770 S/CO, those with HBV DNA women cleared HBsAg; pregnant women with HBsAg levels of 100-999 and 1000 IU/mL. HBeAg-positive pregnant women with HBV DNA <3 × 10(7) IU/mL or HBeAg <770 S/CO are more likely to undergo postpartum HBeAg seroconversion. HBsAg <100 IU/mL is a strong predictor of spontaneous postpartum HBsAg seroclearance. Copyright © 2016 IMSS. Published by Elsevier Inc. All rights reserved.

  5. Surface differentiation antigens of human mammary epithelial cells carried on the human milk fat globule.

    Science.gov (United States)

    Ceriani, R L; Thompson, K; Peterson, J A; Abraham, S

    1977-02-01

    Rabbit antibodies against components of the human milk fat globule bind specifically to normal human breast epithelial cells and cell lines derived from breast carcinomas, as well as to the outer surface of the human milk fat globule. Variation in indirect immunofluorescence staining in both intensity per cell and percentage of cells stained is observed for the different brest cell lines. Cells derived from other epithelial and other ectodermal tissues, fetal fibroblasts, cells of the blood buffy coat, and even fibroblasts of the breast itself do not bind the antibodies. This suggests that these antibodies are detecting cell-type-specific antigens. These normal breast epithelial cell antigens are on the cell surface and their expression is stable in long-term cultured cell lines, even after much chromosomal variation in a given line. By affinity chromatography, three distinct antigenic components can be isolated from the milk fat globule, one of which contains carbohydrate. These differentiation antigens of the human breast epithelial cell are not only useful as specific cell-type markers, but also can provide a tool to study the role of the cell surface in normal and neoplastic mammary development.

  6. Identification of Bacterial Surface Antigens by Screening Peptide Phage Libraries Using Whole Bacteria Cell-Purified Antisera

    Science.gov (United States)

    Hu, Yun-Fei; Zhao, Dun; Yu, Xing-Long; Hu, Yu-Li; Li, Run-Cheng; Ge, Meng; Xu, Tian-Qi; Liu, Xiao-Bo; Liao, Hua-Yuan

    2017-01-01

    Bacterial surface proteins can be good vaccine candidates. In the present study, we used polyclonal antibodies purified with intact Erysipelothrix rhusiopthiae to screen phage-displayed random dodecapeptide and loop-constrained heptapeptide libraries, which led to the identification of mimotopes. Homology search of the mimotope sequences against E. rhusiopthiae-encoded ORF sequences revealed 14 new antigens that may localize on the surface of E. rhusiopthiae. When these putative surface proteins were used to immunize mice, 9/11 antigens induced protective immunity. Thus, we have demonstrated that a combination of using the whole bacterial cells to purify antibodies and using the phage-displayed peptide libraries to determine the antigen specificities of the antibodies can lead to the discovery of novel bacterial surface antigens. This can be a general approach for identifying surface antigens for other bacterial species. PMID:28184219

  7. Identification and characterization of a putative agglutination/immobilization antigen on the surface of Cryptocaryon irritans.

    Science.gov (United States)

    Hatanaka, A; Umeda, N; Yamashita, S; Hirazawa, N

    2007-08-01

    The ciliated protozoan Cryptocaryon irritans, a parasite of seawater fishes, was found to express an antigen that elicits antibodies in rabbits and tiger puffer (Takifugu ruburipes). Serum from rabbits and fish immunized with theronts had agglutination/immobilization activity against theronts in vitro; fish serum antibody levels (measured by enzyme-linked immunosorbent assays: ELISA) correlated with this activity. Anti-theront antibody levels in fish were significantly higher in the immunized group as compared with control fish at 2 weeks after booster immunization (injection of bovine serum albumin; Student's t-test, P<0.01). Biochemical analyses indicated that a Triton X-114-soluble 32 kDa theront integral membrane protein may be the agglutination/immobilization antigen. Indirect immunofluorescence staining of theronts suggested that this 32 kDa antigen was expressed on the surface of cilia. The full-length 32 kDa antigen cDNA contained 1147 basepairs, encoding a 328-amino acid protein including hydrophobic N- and C-termini. As with Tetrahymena and Paramecium spp., TAA and TAG appear to be used as glutamine codons in the 32 kDa antigen gene.

  8. The promoter for a variant surface glycoprotein gene expression site in Trypanosoma brucei

    NARCIS (Netherlands)

    Zomerdijk, J. C.; Ouellette, M.; ten Asbroek, A. L.; Kieft, R.; Bommer, A. M.; Clayton, C. E.; Borst, P.

    1990-01-01

    The variant-specific surface glycoprotein (VSG) gene 221 of Trypanosoma brucei is transcribed as part of a 60 kb expression site (ES). We have identified the promoter controlling this multigene transcription unit by the use of 221 chromosome-enriched DNA libraries and VSG gene 221 expression site

  9. Antibodies produced by mice immunized with recombinant vaccinia viruses expressing two different types of a major Theileria sergenti surface antigen (p32) react with the native surface antigen.

    Science.gov (United States)

    Takasima, Y; Xuan, X; Matsumoto, Y; Onuma, M; Otsuka, H

    1999-07-01

    A 32 kDa major surface antigen, p32, of Theileria sergenti at the piroplasm stage is the main target of the host immune response. The immunogenic property of the p32 varies in some strains among the population of Theileria sergenti in Japan where the Chitose type and the Ikeda type are the most common varieties. We have constructed vaccinia virus recombinants vv/p32C and vv/p32I which harbor the Chitose and Ikeda types of p32 gene, respectively. It was found that vv/p32C and vv/p32I produced type-specific p32 which did not cross react with the monoclonal antibodies (mAbs) against the other type of p32. When mice were immunized with vv/p32C and vv/p32I, antibodies against p32 were detectable 2 weeks after the immunization, and these antibodies reacted with the native surface antigen in purified T. sergenti merozoite.

  10. Neospora caninum surface antigen (p40) is a potential diagnostic marker for cattle neosporosis.

    Science.gov (United States)

    He, Pengfei; Li, Jianhua; Gong, Pengtao; Liu, Chengwu; Zhang, Guocai; Yang, Ju; Tuo, Wenbin; Yang, Bintong; Zhang, Xichen

    2013-05-01

    Neospora caninum is an intracellular protozoan that infects domestic and wild canids as well as many warm-blooded animals as shown by the isolation of viable parasites. The effectiveness of diagnostic tests for detecting specific antibodies against N. caninum is hampered by potential cross-reaction with other Coccidia. So, there is currently an urgent need for a sensitive and specific diagnostic assay for detecting N. caninum in animals. The N. caninum 40-kD surface antigen (p40), similar to NcSAG1 and NcSRS2, was shown to belong to surface antigen super family and thus represents an excellent marker for the diagnosis of neosporosis. In order to test the hypothesis, recombinant Ncp40 (rNcp40) was expressed in Escherichia coli, and an indirect ELISA test was developed using recombinant NCp40 antigen for N. caninum serodiagnosis. The antigen used in this study did not have cross-reactivity with anti-Toxoplasma gondii serum. Anti-p40 antibodies were detected by ELISA in the sera of Yellow cattle and were compared with (IFAT). Optimal sensitivity and specificity (98.2 and 98.6 %) were identified by IFAT. Additionally, 37 positive sera of T. gondii were detected and there was no significant difference with the negative serum of N. caninum. The rNcp40 ELISA developed here provides a specific and sensitive assay for detecting neosporosis in cattle.

  11. KIR content genotypes associate with carriage of hepatitis B surface antigen, e antigen and HBV viral load in Gambians.

    Directory of Open Access Journals (Sweden)

    Louis-Marie Yindom

    Full Text Available Hepatocellular carcinoma (HCC causes over 800,000 deaths worldwide annually, mainly in low income countries, and incidence is rising rapidly in the developed world with the spread of hepatitis B (HBV and C (HCV viruses. Natural Killer (NK cells protect against viral infections and tumours by killing abnormal cells recognised by Killer-cell Immunoglobulin-like Receptors (KIR. Thus genes and haplotypes encoding these receptors may be important in determining both outcome of initial hepatitis infection and subsequent chronic liver disease and tumour formation. HBV is highly prevalent in The Gambia and the commonest cause of liver disease. The Gambia Liver Cancer Study was a matched case-control study conducted between September 1997 and January 2001 where cases with liver disease were identified in three tertiary referral hospitals and matched with out-patient controls with no clinical evidence of liver disease.We typed 15 KIR genes using the polymerase chain reaction with sequence specific primers (PCR-SSP in 279 adult Gambians, 136 with liver disease (HCC or Cirrhosis and 143 matched controls. We investigated effects of KIR genotypes and haplotypes on HBV infection and associations with cirrhosis and HCC.Homozygosity for KIR group A gene-content haplotype was associated with HBsAg carriage (OR 3.7, 95% CI 1.4-10.0 whilst telomeric A genotype (t-AA was associated with reduced risk of e antigenaemia (OR 0.2, 95% CI 0.0-0.6 and lower viral loads (mean log viral load 5.2 vs. 6.9, pc = 0.022. One novel telomeric B genotype (t-ABx2 containing KIR3DS1 (which is rare in West Africa was also linked to e antigenaemia (OR 8.8, 95% CI 1.3-60.5. There were no associations with cirrhosis or HCC.Certain KIR profiles may promote clearance of hepatitis B surface antigen whilst others predispose to e antigen carriage and high viral load. Larger studies are necessary to quantify the effects of individual KIR genes, haplotypes and KIR/HLA combinations on long

  12. Hepatitis B Surface Antigen Testing Among Pregnant Women, United States 2014.

    Science.gov (United States)

    Kolasa, Maureen S; Tsai, Yuping; Xu, Jing; Fenlon, Nancy; Schillie, Sarah

    2017-07-01

    Post-exposure prophylaxis administered to infants shortly after birth prevents approximately 90% of cases of perinatal hepatitis B virus (HBV) transmission. The Advisory Committee on Immunization Practices recommends that all pregnant women be tested for hepatitis B surface antigen (HBsAg) at an early prenatal visit during each pregnancy to detect active infection with HBV. This study sought to determine the proportion and characteristics of pregnant women tested\

  13. Chronic hepatitis B in pregnant women: is hepatitis B surface antigen quantification useful for viral load prediction?

    Science.gov (United States)

    Fujiko, Masita; Chalid, Maisuri T; Turyadi; Ie, Susan I; Maghfira; Syafri; Wahyuni, Ridha; Roni, Martono; Patellongi, Ilhamjaya; Massi, M Nasrum; Muljono, David H

    2015-12-01

    New cases of hepatitis B virus (HBV) infection continue to occur worldwide. Most of these are due to mother-to-child transmission (MTCT), with maternal viraemia as the most important contributing factor. The hepatitis B surface antigen (HBsAg) level, which correlates positively with viral load, has been used for treatment monitoring in chronic hepatitis B. This study evaluated the usefulness of quantitative HBsAg for viral load prediction in HBsAg-positive pregnant women. A total of 943 pregnant women in Makassar, Indonesia, were screened for HBsAg. Sixty-four women were HBsAg-positive and investigated. HBsAg level and hepatitis B e antigen (HBeAg)/hepatitis B e antibody (anti-HBe) status were determined serologically. Viral load was measured by real-time PCR. HBV DNA was sequenced and analysed for identification of genotype and basal core promoter (BCP)/precore (PC) mutations. Of 64 subjects, 12 (18.8%) were HBeAg-positive and 52 (81.3%) were HBeAg-negative. HBsAg and HBV DNA levels were significantly higher in the HBeAg-positive group (ppregnant women. The HBeAg-negative group had a frequent occurrence of BCP/PC variants, which may have contributed to the lack of correlation observed. Samples with a low HBsAg level, which is associated with a low risk of MTCT, do not require HBV DNA measurement. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. Evaluation the Surface Antigen of the Salmonella typhimurium ATCC 14028 Ghosts Prepared by “SLRP”

    Directory of Open Access Journals (Sweden)

    Amara A. Amro

    2014-01-01

    Full Text Available Recently, bacterial ghosts (BGs were prepared using a protocol based on critical chemical concentrations. It has been given the name “sponge like” (SL protocol and used in its reduced form “sponge like reduced protocol” (SLRP. While specific antibody for Salmonella is available on the market under the commercial names (of some kits such as Febrile Antigen Kit (N.S. BIO-TEC, we used the described Kit to investigate the validity of the SLRP. In this study, using SLRP we succeeded to prepare STGs with correct surface antigens could interact with their specific antibodies. Additionally the study has included oral vaccination with STGs with challenge test. The rats serums have been evaluated against both of the O and H antigens. The antigen-antibody interaction (agglutination results of both the SLRP and the animal experiments prove that we have correct STGs able to immunize the rats against viable Salmonella. STGs could be used as vaccine and as adjuvant and in the antibodies and in the diagnostic kits production. This study is an additional step for the establishment of correct BGs for immunological purposes.

  15. Immuno disc assay for screening duck hepatitis B surface antigen in serum, liver tissue and cultured hepatocytes

    NARCIS (Netherlands)

    G.A. de Wilde (G.); R.A. Heijtink

    1993-01-01

    textabstractAn immuno disc assay (IDA) for semi-quantitative analysis of the surface antigen (DHBsAg) of duck hepatitis B virus (DHBV) is described. Unpurified antigen preparations were adsorbed onto punched-out nitrocellulose membrane discs. Rabbit antiserum raised against serum-derived

  16. Simulation and Theory of Antibody Binding to Crowded Antigen-Covered Surfaces.

    Directory of Open Access Journals (Sweden)

    Cristiano De Michele

    2016-03-01

    Full Text Available In this paper we introduce a fully flexible coarse-grained model of immunoglobulin G (IgG antibodies parametrized directly on cryo-EM data and simulate the binding dynamics of many IgGs to antigens adsorbed on a surface at increasing densities. Moreover, we work out a theoretical model that allows to explain all the features observed in the simulations. Our combined computational and theoretical framework is in excellent agreement with surface-plasmon resonance data and allows us to establish a number of important results. (i Internal flexibility is key to maximize bivalent binding, flexible IgGs being able to explore the surface with their second arm in search for an available hapten. This is made clear by the strongly reduced ability to bind with both arms displayed by artificial IgGs designed to rigidly keep a prescribed shape. (ii The large size of IgGs is instrumental to keep neighboring molecules at a certain distance (surface repulsion, which essentially makes antigens within reach of the second Fab always unoccupied on average. (iii One needs to account independently for the thermodynamic and geometric factors that regulate the binding equilibrium. The key geometrical parameters, besides excluded-volume repulsion, describe the screening of free haptens by neighboring bound antibodies. We prove that the thermodynamic parameters govern the low-antigen-concentration regime, while the surface screening and repulsion only affect the binding at high hapten densities. Importantly, we prove that screening effects are concealed in relative measures, such as the fraction of bivalently bound antibodies. Overall, our model provides a valuable, accurate theoretical paradigm beyond existing frameworks to interpret experimental profiles of antibodies binding to multi-valent surfaces of different sorts in many contexts.

  17. Blocking Synthesis of the Variant Surface Glycoprotein Coat in Trypanosoma brucei Leads to an Increase in Macrophage Phagocytosis Due to Reduced Clearance of Surface Coat Antibodies.

    Directory of Open Access Journals (Sweden)

    Jackie L Y Cheung

    2016-11-01

    Full Text Available The extracellular bloodstream form parasite Trypanosoma brucei is supremely adapted to escape the host innate and adaptive immune system. Evasion is mediated through an antigenically variable Variant Surface Glycoprotein (VSG coat, which is recycled at extraordinarily high rates. Blocking VSG synthesis triggers a precytokinesis arrest where stalled cells persist for days in vitro with superficially intact VSG coats, but are rapidly cleared within hours in mice. We therefore investigated the role of VSG synthesis in trypanosome phagocytosis by activated mouse macrophages. T. brucei normally effectively evades macrophages, and induction of VSG RNAi resulted in little change in phagocytosis of the arrested cells. Halting VSG synthesis resulted in stalled cells which swam directionally rather than tumbling, with a significant increase in swim velocity. This is possibly a consequence of increased rigidity of the cells due to a restricted surface coat in the absence of VSG synthesis. However if VSG RNAi was induced in the presence of anti-VSG221 antibodies, phagocytosis increased significantly. Blocking VSG synthesis resulted in reduced clearance of anti-VSG antibodies from the trypanosome surface, possibly as a consequence of the changed motility. This was particularly marked in cells in the G2/ M cell cycle stage, where the half-life of anti-VSG antibody increased from 39.3 ± 4.2 seconds to 99.2 ± 15.9 seconds after induction of VSG RNAi. The rates of internalisation of bulk surface VSG, or endocytic markers like transferrin, tomato lectin or dextran were not significantly affected by the VSG synthesis block. Efficient elimination of anti-VSG-antibody complexes from the trypanosome cell surface is therefore essential for trypanosome evasion of macrophages. These experiments highlight the essentiality of high rates of VSG recycling for the rapid removal of host opsonins from the parasite surface, and identify this process as a key parasite

  18. [Functional analysis of hepatitis B virus immune escape mutants with insertion mutations in the surface antigen].

    Science.gov (United States)

    Yu, Shu-li; Yu, De-min; Zhang, Dong-hua; Jiang, Jie-hong; Chen, Jia; Deng, Lin; Zhang, Xin-xin

    2013-07-01

    To evaluate the influence of insertion mutations occurring in the hydrophobic region, between amino acids 114 and 115, of the hepatitis B surface antigen (HBsAg) on viral antigenicity and replication. Hepatitis B virus (HBV) DNA was obtained from patients with HBsAg-positive chronic hepatitis B (CHB) infection and subjected to sequence analysis and comparison to GenBank reference sequences for HBV genotype B (AB073826) and genotype C (AF286594). Insertion mutations detected in the HBsAg region were used to make recombinant expression plasmids via site-directed mutagenesis. After transfecting the recombinant HBsAg into Huh7 cells, the mutants' effects on viral antigenicity and replication were evaluated by chemiluminescence microparticle immunoassay (CMIA) and Southern blot hybridization, respectively. The viral antigenicity of each mutant was predicted by bioinformatic analysis, using the Jameson-Wolf method to predict the antigenic index, the Hopp-Woods method to predict hydrophilicity, the Emini method to predict the probability of a region lying of the protein's surface, and the Karplus-Schulz method to predict the flexibility of the protein backbone. Two CHB patients harbored HBV with insertion mutations in HBsAg: one with two (NT) and one with three (NTT) inserted amino acids between 114 and 115. The NTT recombinant HBsAg mutant showed no impact on viral replication and reacted weakly with anti-HBs in CMIA (P = 0.02). The antigen indices for the insertion of NTT were 1.00, -0.16, and 0.18, and insertion of the three amino acids affected the index values of five proximal amino acid sites (with an average increase of 0.13). The hydrophilic indices for the insertion of NTT were 0.2, -0.4, and -0.4, with no significant effect on the proximal amino acids. The insertion of the three amino acids changed both the surface probability (range: -0.55 to 2.97; affecting eight proximal amino acids) and the flexibility (range: -0.01 to 1.1; affecting five proximal amino

  19. Phenotypic and genetic characterization of a unique variant of serogroup C ET-15 meningococci (with the antigenic formula C:2a:P1.7,1) causing invasive meningococcal disease in Quebec, Canada.

    Science.gov (United States)

    Tsang, Raymond S W; Tsai, Chao Ming; Zhu, Peixuan; Ringuette, Louise; Lorange, Manon; Law, Dennis K S

    2004-04-01

    Serogroup C Neisseria meningitidis belonging to the electrophoretic type (ET) ET-15, a variant of ET-37, is endemic in Canada. Like other serogroup C ET-37 meningococci, the endemic ET-15 strains are usually found to carry the serotype and serosubtype antigens of 2a:P1.5,2. In 2001, a sudden increase in the number of cases of serogroup C meningococcal disease in Quebec, Canada, was caused by an antigenic variant of the ET-15 strain. This antigenic variant carries the unique serosubtype marker of P1.7,1. Strains of C:2a:P1.7,1 meningococci were not isolated in Canada in large numbers prior to 2001, and the characteristics of these meningococcal strains linked to an outbreak in Quebec, Canada, are described in the present study.

  20. Double blind comparison of T-antigen and ABO(H) cell surface antigens in bladder cancer.

    Science.gov (United States)

    Vafier, J A; Javadpour, N; Worsham, G F; O'Connell, K J

    1984-04-01

    Transitional cell carcinoma of the bladder is a major cause of cancer deaths. Recently, much attention has been focused on ABO(H) antigen deletion in terms of prediction of prognosis. Furthermore, several studies have shown a correlation between T-antigen (a precursor of blood MN glycoprotein) expression in carcinomas of the breast, colon, and stomach. We have studied 56 specimens from 41 patients with transitional cell carcinoma of the bladder for T-antigen expression and ABO(H) antigen deletion. Results were analyzed with respect to tumor grade, tumor stage, and clinical course. The data indicate that T-antigen expression was not completely useful prognostically; it did not correlate with grade, stage, or clinical course. ABO(H) antigen expression or deletion was found to be a better predictor of tumor behavior than tumor grade, despite a false negative rate of 20 to 30 per cent in blood group O patients. We suggest that use of immunoperoxidase techniques will increase the sensitivity in group O patients, thus making ABO(H) deletion a useful predictive parameter of tumor aggressiveness. This is currently being evaluated in our patients.

  1. Human leukocyte antigen DR surface expression on CD14+ monocytes during adverse events after hematopoietic stem cell transplantation.

    Science.gov (United States)

    Döring, Michaela; Rohrer, Karin Melanie; Erbacher, Annika; Gieseke, Friederike; Schwarze, Carl-Philipp; Bader, Peter; Handgretinger, Rupert; Hofbeck, Michael; Kerst, Gunter

    2015-02-01

    The human leukocyte antigen DR surface expression on CD14+ monocytes reflects the degree to which these cells have been activated. Given the central role monocytes and macrophages play in the immune system, a decreased human leukocyte antigen DR expression on CD14+ monocytes results in a hallmark of altered immune status during systemic inflammatory response syndrome. We hypothesize that human leukocyte antigen DR expression might be similarly altered after hematopoietic stem cell transplantation and during post-transplant complications. Using flow cytometry, this study investigates the human leukocyte antigen DR surface expression of CD14+ monocytes in 30 pediatric and young adult patients up to 1 year after hematopoietic stem cell transplantation. Normal values were derived from a control group of healthy children, adolescents, and young adults. Human leukocyte antigen DR expression decreased significantly prior and during bacterial infection or sepsis. By contrast, human leukocyte antigen DR expression levels were elevated before and at the time of viremia. Human leukocyte antigen DR expression was also elevated during acute graft-versus-host disease. In contrast, the expression was reduced when patients had hepatic veno-occlusive disease. A significant decrease of human leukocyte antigen DR expression was associated with a relapse of the underlying disease and before death. Human leukocyte antigen DR expression on CD14+ monocytes appears to be a promising parameter that might allow identification of patients at risk after hematopoietic stem cell transplantation.

  2. A Major Cell Surface Antigen of Coccidioides immitis Which Elicits Both Humoral and Cellular Immune Responses

    Science.gov (United States)

    Hung, Chiung-Yu; Ampel, Neil M.; Christian, Lara; Seshan, Kalpathi R.; Cole, Garry T.

    2000-01-01

    Multinucleate parasitic cells (spherules) of Coccidioides immitis isolates produce a membranous outer wall component (SOW) in vitro which has been reported to be reactive with antibody from patients with coccidioidal infection, elicits a potent proliferative response of murine immune T cells, and has immunoprotective capacity in a murine model of coccidioidomycosis. To identify the antigenic components of SOW, the crude wall material was first subjected to Triton X-114 extraction, and a water-soluble fraction derived from this treatment was examined for protein composition and reactivity in humoral and cellular immunoassays. Protein electrophoresis revealed that the aqueous fraction of three different isolates of C. immitis each contained one or two major glycoproteins (SOWgps), distinguished by their molecular sizes, which ranged from 58 to 82 kDa. The SOWgps, however, showed identical N-terminal amino acid sequences, and each was recognized by sera from patients with C. immitis infection. Antibody raised against the purified 58-kDa glycoprotein (SOWgp58) of the Silveira isolate was used for Western blot and immunolocalization analyses. Expression of SOWgp was shown to be parasitic phase specific, and the antigen was localized to the membranous SOW. The water-soluble fraction of SOW and the purified SOWgp58 were tested for the ability to stimulate proliferation of human peripheral monocytic cells (PBMC). The latter were obtained from healthy volunteers with positive skin test reaction to spherulin, a parasitic-phase antigen of C. immitis, and from volunteers who showed no skin test reaction to the same antigen. The SOW preparations stimulated proliferation of PBMC from skin test-positive but not skin test-negative donors, and the activated cells secreted gamma interferon, which is indicative of a T helper 1 pathway of immune response. Results of this study suggest that SOWgp is a major parasitic cell surface-expressed antigen that elicits both humoral and cellular

  3. Human leukocyte antigen class II variants and adult-onset asthma: does occupational allergen exposure play a role?

    NARCIS (Netherlands)

    Smit, L.A.M.; Strachan, D.P.; Vermeulen, R.; Bakker, P.I.W. de; Demenais, F.; Dumas, O.; Carsin, A.E.; Cullinan, P.; Curjuric, I.; Ghosh, R.E.; Heederik, D.; Imboden, M.; Jarvis, D.; Lathrop, M.; Moual, N. le; Mehta, A.; Miedinger, D.; Sigsgaard, T.; Siroux, V.; Vernez, D.; Zock, J.P.; Kauffmann, F.; Probst-Hensch, N.; Kogevinas, M.; Bouzigon, E.

    2014-01-01

    Recently, a locus centred on rs9273349 in the HLA-DQ region emerged from genome-wide association studies of adult-onset asthma. We aimed to further investigate the role of human leukocyte antigen (HLA) class II in adult-onset asthma and a possible interaction with occupational exposures. We imputed

  4. Human leukocyte antigen class II variants and adult-onset asthma : does occupational allergen exposure play a role?

    NARCIS (Netherlands)

    Smit, Lidwien A M; Strachan, David P; Vermeulen, Roel; de Bakker, Paul I W; Demenais, Florence; Dumas, Orianne; Carsin, Anne-Elie; Cullinan, Paul; Curjuric, Ivan; Ghosh, Rebecca E; Heederik, Dick; Imboden, Medea; Jarvis, Deborah; Lathrop, Mark; Le Moual, Nicole; Mehta, Amar; Miedinger, David; Sigsgaard, Torben; Siroux, Valérie; Vernez, David; Zock, Jan Paul; Kauffmann, Francine; Probst-Hensch, Nicole; Kogevinas, Manolis; Bouzigon, Emmanuelle

    2014-01-01

    Recently, a locus centred on rs9273349 in the HLA-DQ region emerged from genome-wide association studies of adult-onset asthma. We aimed to further investigate the role of human leukocyte antigen (HLA) class II in adult-onset asthma and a possible interaction with occupational exposures. We imputed

  5. Immunogenicity and diagnostic potential of synthetic antigenic cell surface glycans of Leishmania.

    Science.gov (United States)

    Anish, Chakkumkal; Martin, Christopher E; Wahlbrink, Annette; Bogdan, Christian; Ntais, Pantelis; Antoniou, Maria; Seeberger, Peter H

    2013-11-15

    Detection and quantification of pathogen-derived antigenic structures is a key method for the initial diagnosis and follow-up of various infectious diseases. Complex parasitic diseases such as leishmaniasis require highly sensitive and specific tests prior to treatment with potentially toxic drugs. To investigate the diagnostic potential of cell surface glycans found on Leishmania parasites, we identified diagnostically relevant glycan epitopes and used synthetic glycan microarrays to screen sera from infected humans and dogs. On the basis of the screening results, we selected a tetrasaccharide to generate anti-glycan antibodies. The corresponding tetrasaccharide-carrier protein conjugate was immunogenic in mice, and sera obtained from immunized mice specifically detected the Leishmania parasite. These results demonstrate how synthetic glycan arrays, in combination with immunological methods, help to identify promising carbohydrate antigens for pathogen detection.

  6. Surface-Engineering of Red Blood Cells as Artificial Antigen Presenting Cells Promising for Cancer Immunotherapy.

    Science.gov (United States)

    Sun, Xiaoqi; Han, Xiao; Xu, Ligeng; Gao, Min; Xu, Jun; Yang, Rong; Liu, Zhuang

    2017-10-01

    The development of artificial antigen presenting cells (aAPCs) to mimic the functions of APCs such as dendritic cells (DCs) to stimulate T cells and induce antitumor immune responses has attracted substantial interests in cancer immunotherapy. In this work, a unique red blood cell (RBC)-based aAPC system is designed by engineering antigen peptide-loaded major histocompatibility complex-I and CD28 activation antibody on RBC surface, which are further tethered with interleukin-2 (IL2) as a proliferation and differentiation signal. Such RBC-based aAPC-IL2 (R-aAPC-IL2) can not only provide a flexible cell surface with appropriate biophysical parameters, but also mimic the cytokine paracrine delivery. Similar to the functions of matured DCs, the R-aAPC-IL2 cells can facilitate the proliferation of antigen-specific CD8+ T cells and increase the secretion of inflammatory cytokines. As a proof-of-concept, we treated splenocytes from C57 mice with R-aAPC-IL2 and discovered those splenocytes induced significant cancer-cell-specific lysis, implying that the R-aAPC-IL2 were able to re-educate T cells and induce adoptive immune response. This work thus presents a novel RBC-based aAPC system which can mimic the functions of antigen presenting DCs to activate T cells, promising for applications in adoptive T cell transfer or even in direct activation of circulating T cells for cancer immunotherapy. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Bacteroides gingivalis antigens and bone resorbing activity in root surface fractions of periodontally involved teeth

    Energy Technology Data Exchange (ETDEWEB)

    Patters, M.R.; Landsberg, R.L.; Johansson, L.A.; Trummel, C.L.; Robertson, P.R. (Department of Periodontology, University of Connecticut, School of Dental Medicine, Farmington, Connecticut, U.S.A.)

    1982-01-01

    Bone resorbing activity and the presence of antigens of Bacteroides gingivalis were assessed in plaque, calculus, cementum, and dentin obtained from roots of teeth previously exposed to periodontitis. Each fraction was obtained by scaling the root surface. The fraction were extracted by stirring and sonication, and the soluble centrifuged, sterilized, dialyzed, and adjusted to equivalent protein concentrations. Cementum and dentin extracts from impacted teeth were prepared similarly and served as controls. Stimulation of bone resorption by each extract was assessed in organ cultures of fetal rat bones by measurement of release of previously-incorporated /sup 45/Ca from the bone into the medium. In some groups of teeth, calculus and cementum were treated with acid prior to scaling. Citric acid washes were recovered and dialyzed. An enzyme-linked immunosorbent assay (ELISA) was used to assess the extracts for the presence of antigens reactive with an antiserum to B. gingivalis. Significant stimulation of bone resorption was found in all calculus and periodontally-involved cementum preparations. ELISA showed significant levels of B.gingivalis antigens in plaque, calculus, and cementum of periodontally-involved teeth, but not in involved dentin nor in cementum or dentin of impact teeth. Treatment with citric acid removed essentially all B.gingivalis antigens from cementum but not calculus. The results suggest that substances which stimulate bone resorption and substances which react with B. gingivalis antiserum are present in surface plaque, calculus, and cementum or periodontally-involved teeth. These substances are not present in cementum and dentin of impacted teeth nor in dentin of periodontally-involved teeth. Treatment by both scaling and citric demineralization will remove most of these substances from cementum of teeth previously exposed to periodontitis.

  8. Effect of UV radiation on the surface of mammalian immunocompetent cells. 1. The change in expression of some antigens and receptors of murine spleen lymphocyte surface

    Energy Technology Data Exchange (ETDEWEB)

    Krylenkov, V.A.; Malygin, A.M. (AN SSSR, Leningrad. Inst. Tsitologii)

    1982-12-01

    Short-wave (254nm) and long-wave (365 nm) UV rays (ShUS and LUV rays) induce the increase in the expression of surface markers of T lymphocytes-THETA(Thy-1) antigens and B lymphocytes-MBLA-antigens and EAS receptors when affecting mouse spleen cells in nonlethal and small lethal doses. Total cell content with T and B lymphocyte characters in an irradiated suspension exceeds even the total cell quantity in non-irradiated suspension (100%) which points to the possibility of the expression of plasmatic membrane antigens and receptors not manifested on the surface of nonirradiated lymphocytes. In the isolethal dose range (LD/sup 15/-LD/sup 28/) ShUV rays suppress and LUV rays induce further increase of THETA and MBLA antigens expression. Among B lymphocytes surface markers the MBLA antigens are more resistant to ShUV an LUV radiation as compared with the EAC receptors.

  9. Impact of hepatitis B virus (HBV) preS/S genomic variability on HBV surface antigen and HBV DNA serum levels.

    Science.gov (United States)

    Pollicino, Teresa; Amaddeo, Giuliana; Restuccia, Agnese; Raffa, Giuseppina; Alibrandi, Angela; Cutroneo, Giuseppina; Favaloro, Angelo; Maimone, Sergio; Squadrito, Giovanni; Raimondo, Giovanni

    2012-08-01

    To evaluate whether hepatitis B virus (HBV) preS/S gene variability has any impact on serum hepatitis B surface antigen (HBsAg) levels and to analyze the replication capacity of naturally occurring preS/S variants, sera from 40 untreated patients with HBV-related chronic liver disease (hepatitis B e antigen [HBeAg]-positive, n = 11; HBeAg-negative, n = 29) were virologically characterized. Additionally, phenotypic analysis of three different preS/S variant isolates (carrying a 183-nucleotide deletion within the preS1 region, the deletion of preS2 start codon, and a stop signal at codon 182 within the S gene, respectively) was performed. HBV infecting 14 (35%) patients had single or multiple preS/S genomic mutations (i.e., preS1 and/or preS2 deletions, preS2 start codon mutations, C-terminally truncated and/or "a" determinant mutated S protein). Presence of preS/S variants negatively correlated with HBsAg titers (r = -0.431; P = 0.005) and its prevalence did not significantly differ between HBeAg-positive and HBeAg-negative patients. No correlation was found between HBsAg and HBV DNA levels in patients infected with preS/S mutants, whereas a significant correlation was found between HBsAg and viremia levels (r = 0.607; P = 0.001) in patients infected with wild-type HBV strains. HepG2 cells replicating the above-mentioned three preS/S variants showed significant reduction of HBsAg secretion, retention of envelope proteins in the endoplasmic reticulum, less efficient virion secretion and nuclear accumulation of significantly higher amounts of covalently closed circular DNA compared with wild-type HBV replicating cells. In patients infected with preS/S variants, HBV DNA replication and HBsAg synthesis/secretion appear to be dissociated. Therefore, the use of HBsAg titer as diagnostic/prognostic tool has to take into account the frequent emergence of preS/S variants in chronic HBV infection. Copyright © 2012 American Association for the Study of Liver Diseases.

  10. Recombinant hepatitis B surface antigen production in Aspergillus niger: evaluating the strategy of gene fusion to native glucoamylase

    CSIR Research Space (South Africa)

    James, ER

    2012-10-01

    Full Text Available Microbiology and Biotechnology October 2012/ Vol. 96, No.2 Recombinant hepatitis B surface antigen production in Aspergillus niger: evaluating the strategy of gene fusion to native glucoamylase ER James a,c & WH van Zyl b & PJ van Zyl c & JF Görgens..., Pretoria 0001, South Africa Abstract This study demonstrates the potential of Aspergillus niger as a candidate expression system for virus- like particle production using gene fusion. Hepatitis B surface antigen (HBsAg) production, targeted...

  11. Characterization of zinc transporter 8 (ZnT8) antibodies in autoimmune diabetic patients from Argentinian population using monomeric, homodimeric, and heterodimeric ZnT8 antigen variants.

    Science.gov (United States)

    Faccinetti, Natalia I; Guerra, Luciano L; Penas Steinhardt, Alberto; Iacono, Ruben F; Frechtel, Gustavo D; Trifone, Liliana; Poskus, Edgardo; Trabucchi, Aldana; Valdez, Silvina N

    2016-02-01

    In order to gain further knowledge of the structure of zinc transporter 8 (ZnT8) epitopes, we studied the role of the amino acid at position 325 in the antigen and its dimeric conformation for autoantibodies to ZnT8 (ZnT8A) recognition. For this purpose, several ZnT8 C-terminal domain variants were designed: monomer carrying Arg325 or Trp325, homo-dimers ZnT8-Arg-Arg325 and ZnT8-Trp-Trp325, and hetero-dimer ZnT8-Arg-Trp325. Two groups of Argentinian diabetic patients were subjected to analysis using [(35)S]-ZnT8 variants by radioligand binding assay (RBA): i) 100 new-onset, insulin-dependent, type 1 diabetic patients and ii) 282 slowly progressing to insulin requirement, non-obese adult-onset diabetic patients. In addition, 50 type 1 diabetic patients and 100 normal control sera provided by the American Diabetes Association (ADA) were evaluated in order to calculate the sensitivity and specificity of ZnT8A assays for each antigenic variant. Other routine β-cell autoantibodies were also tested by RBA. Of the 100 Argentinian type 1 diabetic patients, 65 were ZnT8A+. Out of them, 8 patients recognized all recombinant forms of ZnT8 and most patients (56) reacted against the heterodimer. Additionally, out of 282 non-obese adult-onset diabetic patients 46 were ZnT8A+, whereas 29 patients recognized only dimers. Besides, exclusive reactivity against ZnT8A was found in 9.0% for type 1 diabetes mellitus and 10.3% for non-obese adult-onset diabetic patients. Significantly higher signal values in RBA were obtained with the heterodimeric variant. An increased detection of humoral autoimmunity was found in both groups when ZnT8A was employed in combination with the other β-cell autoantibodies. The inclusion of homodimeric immunoreactive peptides revealed the existence of quaternary structure-defined epitopes probably resembling the actual state of the autoantigen in vivo. Finally, the differential profiles of ZnT8A exhibited by type 1 and non-obese adult-onset diabetic

  12. Coexistence of sense and anti-sense mRNAs of variant surface protein in Giardia lamblia trophozoites.

    Science.gov (United States)

    Guo, Junli; Zheng, Wenyu; Wang, Yuehua; Li, Yao; Lu, Siqi; Feng, Xianmin

    2014-02-14

    A strategy of the parasitic protozoan Giardia lamblia to evade attack from the host immune system is periodic changes of its surface antigen, a member of the variant surface protein (VSP) family. A post-transcriptional gene silencing mechanism has been proposed to explain the presence of only one among many possible VSPs at any time. To investigate this phenomenon further, we extracted total RNA from cultured trophozoites of the G. lamblia C2 isolate, and cDNA was reverse-transcribed from the RNA. Sense and anti-sense VSPs were amplified from the total cDNA using nested PCR with primers designed from the 3'-conserved region and the known 5' or 3' end of the cDNA library. Sequence analyses of the amplified products revealed more than 34 full-length antisense VSPs and a smear of sense VSPs. Sequence alignments and comparisons revealed that these VSPs contained variable N-termini and conserved C-termini, and could be classified into 5 clades based on the sizes and variations of the N-terminal sequence. All antisense VSPs existed in the sense forms, but no corresponding antisense VSP existed for sense RNA (snsRNA) 16. The coexistence of sense and antisense VSP mRNAs in cultured G. lamblia supports the post-transcriptional regulation of VSP expression. We propose that VSPs transcribed simultaneously in the sense and antisense forms form double-stranded RNAs (dsRNAs) which are degraded by the Dicer endonuclease, while a VSP without an antisense transcription (e.g., snsRNA16) will be expressed on the surface of Giardia. In addition, in the course of this investigation VSPs were identified that were previously not known. PCR-based amplification of specific sense and antisense VSP cDNAs can be used to identify the specific VSP on G. lamblia trophozoites, which is easier than using specific monoclonal antibody approaches. Copyright © 2014. Published by Elsevier Inc.

  13. Squamous cell carcinoma antigen 1 and 2 mRNA and a new variant expressed in hepatocellular carcinoma.

    Science.gov (United States)

    Li, S; Gao, Y; Yang, B; Liang, Z; Wang, Y; Zhai, D; Jing, L; Liu, T; Wang, F; DU, Z; Wang, Y

    2014-01-01

    New tools for diagnostic of HCC remain to further investigate. We have evaluated the expression of SCCA1, 2 mRNA and their prognostic value in hepatocellular carcinoma (HCC). Reverse transcription polymerase chain reaction (RT-PCR) and direct sequencing were performed to evaluate the mRNA expression of SCCA1 and SCCA2 in 93 HCCs, and 93 paired adjacent non-cancerous tissues (PNT), 16 cirrhosis livers and 9 normal livers. The correlation of SCCA variants expression with the clinical parameters and the factors affecting survival were analyzed statistically.Total SCCA was detected in 33.3% of HCCs (31/93), in 9.68% of PNT (9/93) and 22.2% of normal livers (2/9). No expression was found in cirrhosis livers (0/16). The frequencies of total SCCA expression were significantly higher in HCCs than that in PNT and liver cirrhosis (p = 0.000, 0.006). From mRNA sequencing of HCCs, a new SCCA1 variant (presenting a T357A mutation) was identified in 16 specimens, while wild type SCCA1 was identified in 11 specimens and SCCA2 in 27 specimens. Clinicopathological analysis showed that the frequency of SCCA1 was significantly higher in poorly differentiated HCC, compared with moderately and well differentiated tumors (p = 0.021). T357A variant has a significantly higher frequency in nonencapsulated tumors than wild type SCCA1 (p = 0.034).The SCCA1, 2 mRNA is effective for detecting HCC and could be potentially applied in HCC diagnosis.

  14. Hepatitis B Surface Antigen Screening Among Pregnant Women and Care of Infants of Hepatitis B Surface Antigen-Positive Mothers - Guam, 2014.

    Science.gov (United States)

    Abara, Winston E; Cha, Susan; Malik, Tasneem; DeSimone, Mia S; Schumann, Bernadette; Mallada, Esther; Klemme, Michael; Aguon, Vince; Santos, Anne Marie; Collier, Melissa; Kamb, Mary

    2017-05-19

    Hepatitis B virus (HBV) infection is endemic among adults in the U.S. territory of Guam (1,2). Perinatal HBV transmission, which occurs at birth from an infected mother to her newborn infant, is a major mode of HBV transmission and maintains HBV endemicity (3). Approximately 90% of HBV-infected infants will develop chronic HBV infection, and approximately 25% of those will die prematurely from liver failure or hepatocellular carcinoma (4,5). Since 1988, the Advisory Committee on Immunization Practices has recommended that all pregnant women be screened for hepatitis B surface antigen (HBsAg), an indicator of HBV infection, and that infants of women who screen positive (HBsAg-positive women) receive postexposure prophylaxis (PEP) (hepatitis B vaccine and hepatitis B immunoglobulin [HBIG]). When received within 12 hours of birth, PEP is 85%-95% effective in preventing perinatal HBV transmission (5,6). Hepatitis B vaccine provides long-term active immunity to HBV infection and HBIG provides short-term passive immunity to HBV infection until the infant responds to the vaccine (5). Hepatitis B vaccine was introduced into the routine universal infant vaccination schedule in Guam in 1988 (1).

  15. 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

    How immunity to malaria develops remains one of the great unresolved issues in bio-medicine and resolution of its various paradoxes is likely to be the key to developing effective malaria vaccines. The basic epidemiological observations are; under conditions of intense natural transmission, humans...... do become immune to P. falciparum malaria, but this is a slow process requiring multiple disease episodes which many, particularly young children, do not survive. Adult survivors are immune to the symptoms of malaria, and unless pregnant, can control the growth of most or all new inoculations....... 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...

  16. Postvaccination seroconversion against the surface antigen of Hepatitis B virus, in nursing students

    Directory of Open Access Journals (Sweden)

    Gladys Amanda Mera-Urbano

    2013-09-01

    Full Text Available Objective: To determine the status of seroconversion after vaccination against the surface antigen of hepatitis B virus in nursing students, University of Cauca. Methods: Cross sectional study in students of V and VI semester. The sample was taken from 37 students, 15 of V and 22 of VI semester. The instrument used was a survey that included 11 questions of multiple selections. Records for weight, height and laboratory results were collected; blood samples for antibody titers were performed with informed consent. The data were tabulated and analyzed using SPSS, version 17.0. Results: 89.2% of students had levels of antibodies to the surface antigen. This value was greater than 10 mUI/ml, considered by the scientific community as a protector value of Hepatitis B. 10.8% of had lesser values. Regarding vaccination scheme, 24% had a dose, 19% two, 48% three and 8% had a one dose. The population with 3 doses and reinforcement seroconverted by 100%. Conclusion: This study demonstrated failings in the scheme of vaccination of the students of nursing and that 10.8 % presented lower values than 10 mIU/ml. It is necessary to apply the institutional rules with more strength as a preventive measure for hepatitis B.

  17. Blastocystis surface antigen is stable in chemically preserved stool samples for at least 1 year.

    Science.gov (United States)

    Gould, Rick; Boorom, Kenneth

    2013-07-01

    Blastocystis spp. refer to a group of prevalent enteric protists found in humans and animals. Detection of Blastocystis spp. in fecal samples is often performed by clinicians with direct microscopy, which provides low sensitivity, or with culture and polymerase chain reaction testing, a method which is problematic when used with formalin-preserved stool samples. Prior study of Blastocystis and other enteric protists suggests that immunofluorescence antibody (IFA) stain could provide sensitivity and compatibility with formalin, but no information is available on the longevity of Blastocystis sp.'s surface antigens in formalin. We collected fecal samples from animals at a country fair held in the summer of 2009 in Oregon, USA. Samples were tested for the presence of Blastocystis infection using an IFA stain shortly after collection, and again after 1 year, with samples stored refrigerated at 4-8 °C. Most samples collected from steer, pigs, and goats were found to be Blastocystis positive. All fecal samples that were Blastocystis positive initially remained positive after 1 year. Blastocystis-negative samples remained negative. Minimal degradation was observed in stained slides. Blastocystis surface antigens detected by a polyclonal stain remained stable in formalin for a period of at least 1 year.

  18. Probing the effects of surface hydrophobicity and tether orientation on antibody-antigen binding

    Science.gov (United States)

    Bush, Derek B.; Knotts, Thomas A.

    2017-04-01

    Antibody microarrays have the potential to revolutionize molecular detection for many applications, but their current use is limited by poor reliability, and efforts to change this have not yielded fruitful results. One difficulty which limits the rational engineering of next-generation devices is that little is known, at the molecular level, about the antibody-antigen binding process near solid surfaces. Atomic-level structural information is scant because typical experimental techniques (X-ray crystallography and NMR) cannot be used to image proteins bound to surfaces. To overcome this limitation, this study uses molecular simulation and an advanced, experimentally validated, coarse-grain, protein-surface model to compare fab-lysozyme binding in bulk solution and when the fab is tethered to hydrophobic and hydrophilic surfaces. The results show that the tether site in the fab, as well as the surface hydrophobicity, significantly impacts the binding process and suggests that the optimal design involves tethering fabs upright on a hydrophilic surface. The results offer an unprecedented, molecular-level picture of the binding process and give hope that the rational design of protein-microarrays is possible.

  19. 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.

  20. Genetic Diversity of Polymorphic Vaccine Candidate Antigens (Apical Membrane Antigen-1, Merozoite Surface Protein-3, and Erythrocyte Binding Antigen-175) in Plasmodium falciparum Isolates from Western and Central Africa

    Science.gov (United States)

    Soulama, Issiaka; Bigoga, Jude D.; Ndiaye, Magatte; Bougouma, Edith C.; Quagraine, Josephine; Casimiro, Prisca N.; Stedman, Timothy T.; Sirima, Sodiomon B.

    2011-01-01

    The malaria vaccine candidate antigens erythrocyte binding antigen 175 (EBA-175), merozoite surface protein 3 (MSP-3), and apical membrane antigen (AMA-1) from Plasmodium falciparum isolates from countries in central and west Africa were assessed for allelic diversity. Samples were collected on filter paper from 600 P. falciparum-infected symptomatic patients in Cameroon, Republic of Congo, Burkina Faso, Ghana, and Senegal and screened for class-specific amplification fragments. Genetic diversity, assessed by mean heterozygosity, was comparable among countries. We detected a clinical increase in eba 175 F-allele frequency from west to east across the study region. No statistical difference in msp-3 allele distribution between countries was observed. The ama-1 3D7 alleles were present at a lower frequency in central Africa than in West Africa. We also detected little to no genetic differentiation among sampling locations. This finding indicates that, at least at the level of resolution offered by restriction fragment length polymorphism analysis, these antigens showed remarkable genetic homogeneity throughout the region sampled, perhaps caused by balancing selection to maintain a diverse array of antigen haplotyes. PMID:21292899

  1. A monoclonal antibody reactive with Marek's disease tumor-associated surface antigen.

    Science.gov (United States)

    Lee, L F; Liu, X; Sharma, J M; Nazerian, K; Bacon, L D

    1983-02-01

    An antibody-secreting hybridoma, RPH-6, to Marek's disease tumor-associated surface antigen (MATSA) was produced by somatic-cell hybridization between the mouse myeloma SP2/O-Ag/14 and spleen cells from MSB1 immunized mice. The antibody reacted with 95 to 100% of the cells from eight of 10 chicken MD cell lines and one of two turkey MD cell lines. It did not react with chicken MD cell lines RP1 and SK3, turkey MD cell line RP19, lymphoid leukosis (LL) cell lines, RP9 and RP12, and reticuloendotheliosis virus (REV) cell line RP14. A weak reaction was observed with REV cell line RP13 (10 to 20%), and a very slight reaction with normal chicken spleen cells (1 to 5%). RPH-6 produces immunoglobulin of the IgM class. Cell culture and mouse ascitic fluids have titers of 10(2) and 10(6), respectively, by fluorescent antibody (FA) test against MD tumor cell lines. The antibody did not react with Marek's disease virus (MDV) internal antigen or membrane antigen as detected by indirect FA test on chicken embryo fibroblast cultures grown on coverslips. The 51Cr release assay showed RPH-6 is highly cytotoxic (60% release) only against MD lymphoblastoid cell lines, with antibody prepared from mouse ascites having a cytotoxic titer approaching 10(6). Results from using RPH-6 for differential diagnosis of chicken lymphoid tumors of unknown origin were in complete agreement with those obtained with rabbit anti-MATSA reference serum and pathologic diagnosis.

  2. Antibody production by injection of living cells expressing non self antigens as cell surface type II transmembrane fusion protein.

    Science.gov (United States)

    Nizet, Yannick; Gillet, Laurent; Schroeder, Hélène; Lecuivre, Corinne; Louahed, J; Renauld, J-C; Gianello, Pierre; Vanderplasschen, Alain

    2011-03-31

    Antigen expression and purification are laborious, time consuming and frequently difficult steps in the process of antibody production. In the present study, we developed a method avoiding these two steps. This method relies on the injection of histocompatible living cells stably expressing the antigen as a cell surface type II transmembrane fusion protein. A vector, nicknamed pCD1-CD134L, was constructed to express the antigen fused at the carboxyterminal end of the human CD134 ligand (CD134L) type II transmembrane protein on the surface of eucaryotic cells. This vector was shown to induce cell surface expression of epitopes from human c-Myc (soluble protein), uterogloblin-related protein 1 (secreted protein) and CD94 (type II transmembrane protein). Using this vector, we developed a method to produce antibodies without antigen production. The flowchart of this method is as follows: (i) cloning of the antigen in the pCD1-CD134L vector; (ii) production of a histocompatible cell line stably expressing the CD134L-antigen fusion protein; (iii) testing for cell surface expression of the fusion protein by targeting the CD134L carrier; and (iv) prime-boost immunisation with living cells expressing the fusion protein. This method was successfully used for production of polyclonal antibodies raised against Ixodes ricinus calreticulin (secreted protein) in mice and for production of monoclonal antibodies raised against an epitope of Vaccinia virus A56 (type I transmembrane protein) protein in rat. The present study is the first to demonstrate the use of a type II transmembrane protein as a carrier for cell surface display of antigens. Copyright © 2011 Elsevier B.V. All rights reserved.

  3. Longitudinal microarray analysis of cell surface antigens on peripheral blood mononuclear cells from HIV+ individuals on highly active antiretroviral therapy

    Directory of Open Access Journals (Sweden)

    Wang Bin

    2008-03-01

    Full Text Available Abstract Background The efficacy of highly active antiretroviral therapy (HAART determined by simultaneous monitoring over 100 cell-surface antigens overtime has not been attempted. We used an antibody microarray to analyze changes in the expression of 135 different cell-surface antigens overtime on PBMC from HIV+ patients on HAART. Two groups were chosen, one (n = 6 achieved sustainable response by maintaining below detectable plasma viremia and the other (n = 6 responded intermittently. Blood samples were collected over an average of 3 years and 5–8 time points were selected for microarray assay and statistical analysis. Results Significant trends over time were observed for the expression of 7 cell surface antigens (CD2, CD3epsilon, CD5, CD95, CD36, CD27 and CD28 for combined patient groups. Between groups, expression levels of 10 cell surface antigens (CD11a, CD29, CD38, CD45RO, CD52, CD56, CD57, CD62E, CD64 and CD33 were found to be differential. Expression levels of CD9, CD11a, CD27, CD28 and CD52, CD44, CD49d, CD49e, CD11c strongly correlated with CD4+ and CD8+ T cell counts, respectively. Conclusion Our findings not only detected markers that may have potential prognostic/diagnostic values in evaluating HAART efficacy, but also showed how density of cell surface antigens could be efficiently exploited in an array-like manner in relation to HAART and HIV-infection. The antigens identified in this study should be further investigated by other methods such as flow cytometry for confirmation as biological analysis of these antigens may help further clarify their role during HAART and HIV infection.

  4. Hepatitis B surface antigen prevalence in pregnant women: A cross-sectional survey in Iran.

    Science.gov (United States)

    Shoghli, Alireza; Nabavi, Seyed Mahmood; Alavian, Seyed Moayed; Kolifarhood, Goodarz; Goya, Mohammad Mehdi; Namazi, Roshanak; Fallahnezhad, Mojtaba; Mohajeri, Mansor; Mousavinasab, Nouraldin; Zanjani, Rahim Sorouri; Saeini, Mohammad Reza; Jalilvand, Ahmad

    2014-12-01

    Vertical transmission of hepatitis B virus (HBV) from infected mothers to their neonates is one of the most important routes of infection. The exact prevalence rate of HBV in Iranian pregnant mothers is not well known but based on different studies it is estimated between 0.35% and 6.5%. The aim of this study was to determine the seroprevalence of hepatitis B surface antigen (HBsAg) in pregnant women of selected provinces in Iran. At this cross-sectional study, seven provinces supposed to be of high and low prevalence of hepatitis B in the general population selected. Multistage sampling was used to enroll 5261 parturient women who attended the target provinces birth facilities, during January to March of 2011, were recruited to study. To determine the statistically significant mean and proportion differences, t-test and χ (2) test were used, respectively. Overall 1.2% was positive HBsAg of which 11% of them were hepatitis B e-antigen positive as well. The eastern and north eastern provinces were considerably higher in HBsAg seropositivity than the west and northwest of the country. In view of the considerable prevalence of hepatitis B in pregnant women, screening all pregnant women prioritizing the eastern and north-eastern provinces is strongly recommended.

  5. HCV core protein modulates the immune response against the HBV surface antigen in mice.

    Science.gov (United States)

    Aguilar, Julio C; Acosta-Rivero, Nelson; Dueñas-Carrera, Santiago; Morales Grillo, Juan; Pichardo, Dagmara; Urquiza, Dioslaida; Guillen, Gerardo; Muzio, Verena

    2003-10-10

    Mucosal vaccination is currently arousing a great deal of interest, since mucosally induced immunity is able to protect not only against microorganisms using mucosa as a door of entry, but also against those parenterally transmitted. Hepatitis C virus (HCV) is considered a worldwide health problem and a current vaccine is not available. In the present work, immunogenicity of particulate HCcAg was evaluated, administered alone and also in formulations with the main protective antigen of HBV, the surface antigen (HBsAg), both by mucosal (i.n.) and parenteral (i.m) routes. HCcAg was able to induce strong immune responses after nasal as well as parenteral administration, developing a strong Th1-like antibody response in serum. Preliminary data also suggested the ability of HCcAg to efficiently enhance and modulate the host immune response against HBsAg. These results support the use of the particulate HCcAg in the rational design of candidates for HCV therapeutic or preventive vaccine strategies or inclusively in the development of future combined vaccines.

  6. Involvement of TSSA (trypomastigote small surface antigen) in Trypanosoma cruzi invasion of mammalian cells.

    Science.gov (United States)

    Cánepa, Gaspar E; Degese, Maria Sol; Budu, Alexandre; Garcia, Celia R S; Buscaglia, Carlos A

    2012-06-01

    TSSA (trypomastigote small surface antigen) is a polymorphic mucin-like molecule displayed on the surface of Trypanosoma cruzi trypomastigote forms. To evaluate its functional properties, we undertook comparative biochemical and genetic approaches on isoforms present in parasite stocks from extant evolutionary lineages (CL Brener and Sylvio X-10). We show that CL Brener TSSA, but not the Sylvio X-10 counterpart, exhibits dose-dependent and saturable binding towards non-macrophagic cell lines. This binding triggers Ca(2+)-based signalling responses in the target cell while providing an anchor for the invading parasite. Accordingly, exogenous addition of either TSSA-derived peptides or specific antibodies significantly inhibits invasion of CL Brener, but not Sylvio X-10, trypomastigotes. Non-infective epimastigote forms, which do not express detectable levels of TSSA, were stably transfected with TSSA cDNA from either parasite stock. Although both transfectants produced a surface-associated mucin-like TSSA product, epimastigotes expressing CL Brener TSSA showed a ~2-fold increase in their attachment to mammalian cells. Overall, these findings indicate that CL Brener TSSA functions as a parasite adhesin, engaging surface receptor(s) and inducing signalling pathways on the host cell as a prerequisite for parasite internalization. More importantly, the contrasting functional features of TSSA isoforms provide one appealing mechanism underlying the differential infectivity of T. cruzi stocks.

  7. Antigenicity and Immunogenicity of Rotavirus VP6 Protein Expressed on the Surface of Lactococcus lactis

    Directory of Open Access Journals (Sweden)

    L. E. Esteban

    2013-01-01

    Full Text Available Group A rotaviruses are the major etiologic agents of acute gastroenteritis worldwide in children and young animals. Among its structural proteins, VP6 is the most immunogenic and is highly conserved within this group. Lactococcus lactis is a food-grade, Gram-positive, and nonpathogenic lactic acid bacteria that has already been explored as a mucosal delivery system of heterologous antigens. In this work, the nisin-controlled expression system was used to display the VP6 protein at the cell surface of L. lactis. Conditions for optimal gene expression were established by testing different nisin concentrations, cell density at induction, and incubation times after induction. Cytoplasmic and cell wall protein extracts were analyzed by Western blot and surface expression was confirmed by flow cytometry. Both analysis provided evidence that VP6 was efficiently expressed and displayed on the cell surface of L. lactis. Furthermore, the humoral response of mice immunized with recombinant L. lactis was evaluated and the displayed recombinant VP6 protein proved to be immunogenic. In conclusion, this is the first report of displaying VP6 protein on the surface of L. lactis to induce a specific immune response against rotavirus. These results provide the basis for further evaluation of this VP6-displaying L. lactis as a mucosal delivery vector in a mouse model of rotavirus infection.

  8. Ac-SAA-1, an immunodominant 16 kDa surface-associated antigen of infective larvae and adults of Ancylostoma caninum.

    Science.gov (United States)

    Zhan, Bin; Wang, Yan; Liu, Yueyuan; Williamson, Angela; Loukas, Alex; Hawdon, John M; Xue, Hae-Chou; Xiao, Shu-Hua; Hotez, Peter J

    2004-08-01

    A cDNA encoding a surface-associated antigen was cloned from an Ancylostoma caninum infective larvae (L(3)) cDNA library by immunoscreening with pooled human immune sera. The sera were obtained from individuals living in an Ancylostoma duodenale hookworm-endemic region of China, who had light intensity infections and high antibody titers against A. caninum L(3). Ancylostoma caninum surface-associated antigen-1 is encoded by an 843 bp mRNA with a predicted open reading frame of 162 amino acids. Recombinant Ancylostoma caninum surface-associated antigen-1 was expressed in Escherichia coli and used to prepare a specific antiserum. A Western blot with anti-Ancylostoma caninum surface-associated antigen-1 specific antiserum showed that native Ancylostoma caninum surface-associated antigen-1 protein is expressed by both L(3) and adult hookworms; RT-PCR confirmed that the mRNA is transcribed in both stages. In adult hookworms, the protein localised to the basal layer of the cuticle and hypodermis of adult worms. Serological analysis determined that recombinant Ancylostoma caninum surface-associated antigen-1 protein is recognised by 61% of human sera from a Necator americanus hookworm endemic area in China, indicating the antigen is immunodominant. Anti-Ancylostoma caninum surface-associated antigen-1 antiserum partially inhibited (46.7%) invasion of hookworm L(3) into dog skin in vitro. Together these results suggest that Ancylostoma caninum surface-associated antigen-1 offers promise as a protective vaccine antigen.

  9. Genetic variants in human leukocyte antigen-DP influence both hepatitis C virus persistence and hepatitis C virus F protein generation in the Chinese Han population.

    Science.gov (United States)

    Xu, Xiaodong; Yue, Ming; Jiang, Longfeng; Deng, Xiaozhao; Zhang, Yongxiang; Zhang, Yun; Zhu, Danyan; Xiao, Wen; Zhou, Zhenxian; Yao, Wenjuan; Kong, Jing; Yu, Xiaojie; Wei, Juan

    2014-06-03

    Chronic hepatitis C is a serious liver disease that often results in cirrhosis or hepatocellular carcinoma. The aim of this study was to assess the association of human leukocyte antigen-DP (HLA-DP) variants with risk of chronic hepatitis C virus (HCV) or anti-F antibody generation. We selected two single nucleotide polymorphisms (SNPs) in a region including HLA-DPA1 (rs3077) and HLA-DPB1 (rs9277534) and genotyped SNPs in 702 cases and 342 healthy controls from the Chinese population using TaqMan SNP genotyping assay. Moreover, the exon 2 of the HLA-DPA1 and HLA-DPB1 genes were amplified and determined by sequencing-based typing (SBT). The results showed that rs3077 significantly increased the risk of chronic HCV infection in additive models and dominant models (odds ratio (OR) = 1.32 and 1.53). The rs3077 also contributed to decrease the risk of anti-F antibody generation in additive models and dominant models (OR = 0.46 and 0.56). Subsequent analyses revealed the risk haplotypes (DPA1*0103-DPB1*0501 and DPA1*0103-DPB1*0201) and protective haplotypes (DPA1*0202-DPB1*0501 and DPA1*0202-DPB1*0202) to chronic HCV infection. Moreover, we also found that the haplotype of DPA1*0103-DPB1*0201 and DPA1*0202-DPB1*0202 were associated with the anti-F antibody generation. Our findings show that genetic variants in HLA-DP gene are associated with chronic HCV infection and anti-F antibody generation.

  10. Association of human leukocyte antigen variants and allopurinol-induced Stevens-Johnson syndrome and toxic epidermal necrolysis: A meta-analysis.

    Science.gov (United States)

    Li, Xingang; Zhao, Zhigang; Sun, Shu-Sen

    2017-05-01

    The association between human leukocyte antigen (HLA) variants and allopurinol-induced Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) was evaluated through a pooled analysis of published studies. A comprehensive search was performed in multiple databases, including PubMed, MEDLINE, ISI Web of Knowledge, EMBASE, Cochrane Register of Controlled Trials, and Science Direct. Studies investigating the association between HLA alleles with allopurinol-induced SJS or TEN were retrieved, and the data were independently extracted. The overall odds ratios (ORs) with corresponding 95% confidence intervals were calculated to determine the association between the presence of HLA variant in at least one allele and allopurinol-induced SJS or TEN. To test the robustness of the meta-analysis results, a sensitivity analysis was performed by removing each study one at a time and calculating the pooled ORs of the remaining studies. The fixed-effects and random-effects models were used to pool the collected data. A total of 4 studies with 81 allopurinol-induced SJS or TEN cases and matched controls (allopurinol-tolerant patients) or population controls (general population) were identified. SJS and TEN were found to be significantly associated with HLA-A*33:03 and HLA-C*03:02 alleles in both groups of studies with matched controls and population controls. All of the pooled ORs were not significantly affected by the remaining studies and different modeling methods, indicating robust results. A strong association was found between HLA-A*33:03 and HLA-C*03:02 alleles and allopurinol-induced SJS or TEN, especially in an Asian population. Copyright © 2017 by the American Society of Health-System Pharmacists, Inc. All rights reserved.

  11. DNA secondary structures are associated with recombination in major Plasmodium falciparum variable surface antigen gene families

    DEFF Research Database (Denmark)

    Sander, Adam F.; Lavstsen, Thomas; Rask, Thomas Salhøj

    2014-01-01

    -wide recombination hotspots in var genes, we show that during the parasite’s sexual stages, ectopic recombination between isogenous var paralogs occurs near low folding free energy DNA 50-mers and that these sequences are heavily concentrated at the boundaries of regions encoding individual Plasmodium falciparum...... falciparum-erythrocyte membrane protein 1 class on the infected erythrocyte surface. Recombination clearly generates var diversity, but the nature and control of the genetic exchanges involved remain unclear. By experimental and bioinformatic identification of recombination events and genome...... of recombination during DNA replication in P. falciparum sexual stages, and that these DSS-regulated genetic exchanges generate functional and diverse P. falciparum adhesion antigens. DSS-induced recombination may represent a common mechanism for optimizing the evolvability of virulence gene families in pathogens....

  12. Identification of the Streptococcus pyogenes surface antigens recognised by pooled human immunoglobulin

    Science.gov (United States)

    Reglinski, Mark; Gierula, Magdalena; Lynskey, Nicola N.; Edwards, Robert J.; Sriskandan, Shiranee

    2015-01-01

    Immunity to common bacteria requires the generation of antibodies that promote opsonophagocytosis and neutralise toxins. Pooled human immunoglobulin is widely advocated as an adjunctive treatment for clinical Streptococcus pyogenes infection however, the protein targets of the reagent remain ill defined. Affinity purification of the anti-streptococcal antibodies present within pooled immunoglobulin resulted in the generation of an IgG preparation that promoted opsonophagocytic killing of S. pyogenes in vitro and provided passive immunity in vivo. Isolation of the streptococcal surface proteins recognised by pooled human immunoglobulin permitted identification and ranking of 94 protein antigens, ten of which were reproducibly identified across four contemporary invasive S. pyogenes serotypes (M1, M3, M12 and M89). The data provide novel insight into the action of pooled human immunoglobulin during invasive S. pyogenes infection, and demonstrate a potential route to enhance the efficacy of antibody based therapies. PMID:26508447

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

    DEFF Research Database (Denmark)

    Bengtsson, Dominique; Sowa, Kordai M; Salanti, Ali

    2008-01-01

    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...... 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...

  14. Association of Cytotoxic T-Lymphocyte Antigen 4 (CTLA4) and Thyroglobulin (TG) Genetic Variants with Autoimmune Hypothyroidism

    Science.gov (United States)

    Patel, Hinal; Mansuri, Mohmmad Shoab; Singh, Mala; Begum, Rasheedunnisa; Shastri, Minal; Misra, Ambikanandan

    2016-01-01

    Autoimmune hypothyroidism is known to be caused by immune responses related to the thyroid gland and its immunological feature includes presence of autoimmune antibodies. Therefore the aim was to analyze presence of anti-TPO antibodies in hypothyroidism patients in Gujarat. Cytotoxic T-Lymphocyte Antigen 4 (CTLA4) is one of the susceptibility genes for various autoimmune diseases. Hence, exon1 +49A/G and 3’UTR CT60A/G single nucleotide polymorphisms (SNPs) in CTLA4 and its mRNA expression levels were investigated in autoimmune hypothyroidism patients. Thyroglobulin (TG) is known to be associated with autoimmune thyroid disorders and thus exon 33 (E33) SNP in TG was investigated. We analyzed the presence of anti-TPO antibodies in the plasma samples of 84 hypothyroidism patients and 62 controls by ELISA. PCR-RFLP technique was used for genotyping of polymorphisms. sCTLA4 and flCTLA4 mRNA expression levels were assessed by real time PCR. 59.52% of hypothyroid patients had anti-TPO antibodies in their circulation. The genotype and allele frequencies differed significantly for +49A/G (p = 0.0004 for +49AG, p = 0.0019 for +49GG & p = 0.0004 for allele), CT60 (p = 0.0110 for CT60AG, p = 0.0005 for CT60GG & phypothyroidism when adjusted for age and gender. Our results suggest +49A/G and CT60 polymorphism of CTLA4 and E33 polymorphism of TG may be genetic risk factors for autoimmune hypothyroidism susceptibility and down regulation of both forms of CTLA4 advocates the crucial role of CTLA4 in pathogenesis of autoimmune hypothyroidism. PMID:26963610

  15. AntigenMap 3D: an online antigenic cartography resource.

    Science.gov (United States)

    Barnett, J Lamar; Yang, Jialiang; Cai, Zhipeng; Zhang, Tong; Wan, Xiu-Feng

    2012-05-01

    Antigenic cartography is a useful technique to visualize and minimize errors in immunological data by projecting antigens to 2D or 3D cartography. However, a 2D cartography may not be sufficient to capture the antigenic relationship from high-dimensional immunological data. AntigenMap 3D presents an online, interactive, and robust 3D antigenic cartography construction and visualization resource. AntigenMap 3D can be applied to identify antigenic variants and vaccine strain candidates for pathogens with rapid antigenic variations, such as influenza A virus. http://sysbio.cvm.msstate.edu/AntigenMap3D

  16. Identification of conserved surface proteins as novel antigenic vaccine candidates of Actinobacillus pleuropneumoniae.

    Science.gov (United States)

    Chen, Xiabing; Xu, Zhuofei; Li, Lu; Chen, Huanchun; Zhou, Rui

    2012-12-01

    Actinobacillus pleuropneumoniae is an important swine respiratory pathogen causing great economic losses worldwide. Identification of conserved surface antigenic proteins is helpful for developing effective vaccines. In this study, a genome-wide strategy combined with bioinformatic and experimental approaches, was applied to discover and characterize surface-associated immunogenic proteins of A. pleuropneumoniae. Thirty nine genes encoding outer membrane proteins (OMPs) and lipoproteins were identified by comparative genomics and gene expression profiling as being-highly conserved and stably transcribed in the different serotypes of A. pleuropneumoniae reference strains. Twelve of these conserved proteins were successfully expressed in Escherichia coli and their immunogenicity was estimated by homologous challenge in the mouse model, and then three of these proteins (APJL_0126, HbpA and OmpW) were further tested in the natural host (swine) by homologous and heterologous challenges. The results showed that these proteins could induce high titers of antibodies, but vaccination with each protein individually elicited low protective immunity against A. pleuropneumoniae. This study gives novel insights into immunogenicity of the conserved OMPs and lipoproteins of A. pleuropneumoniae. Although none of the surface proteins characterized in this study could individually induce effective protective immunity against A. pleuropneumoniae, they are potential candidates for subunit vaccines in combination with Apx toxins.

  17. Successful antiviral therapy determines a significant decrease in squamous cell carcinoma antigen-associated (SCCA) variants' serum levels in anti-HCV positive cirrhotic patients.

    Science.gov (United States)

    Giannini, E G; Basso, M; Bazzica, M; Contini, P; Marenco, S; Savarino, V; Picciotto, A

    2010-08-01

    Aberrant squamous cell carcinoma antigen (SCCA) expression is an early event in hepatocarcinogenesis, and increasing serum levels of SCCA variants IgM immune complexes (SCCA-IgM IC) have been found in cirrhotic patients developing hepatocellular carcinoma (HCC). We longitudinally evaluated a cohort of cirrhotic patients with hepatitis C virus infection (HCV) who underwent pegylated interferon (PEG-IFN) and ribavirin treatment. SCCA-IgM IC levels were assessed in the sera of 33 cirrhotic patients with HCV (21 males, median age 57 years) before, at the end and at 6-month and 1-year follow-up after treatment with PEG-IFN and ribavirin. SCCA-IgM IC serum levels (arbitrary units/mL, AU/mL) were evaluated according to treatment outcome: sustained virological response (SVR) vs nonresponse (NR). Overall, 15 patients obtained a SVR to antiviral therapy (45%). There was no significant difference in baseline SCCA-IgM IC serum levels between SVR and NR patients. When compared to baseline (451.2 AU/mL), SVR patients showed a significant decrease in median SCCA-IgM IC serum levels at the end of treatment (186.8 AU/mL, P = 0.013) and at both 6-month (96.8 AU/mL, P SCCA-IC serum levels. Because of the pathophysiological correlation between SCCA and liver carcinogenesis, it is hypothesized that in patients with liver cirrhosis, SVR may be accompanied by a decreased proliferative stimulation.

  18. Cloning and characterization of a variant surface glycoprotein expression site from Trypanosoma equiperdum.

    Science.gov (United States)

    Raibaud, A; Buck, G; Baltz, T; Eisen, H

    1986-08-01

    Variant surface glycoprotein (VSG) genes of African trypanosomes are expressed when they are inserted into one of several telomere-linked expression sites. We cloned and characterized an 11-kilobase (kb) DNA fragment located upstream of an expressed VSG gene. A DNA sequence of 1.8 kb that is located immediately upstream of the inserted VSG gene contains sequences homologous to the 76-base-pair repeats described as being upstream of VSG genes in Trypanosoma brucei (D. A. Campbell, M. P. Van Bree, and J. C. Boothroyd, Nucleic Acids Res. 12:2759-2774). There are no such sequences elsewhere in the 11-kb cloned region. Southern blot analysis using probes from the cloned region revealed multiple unlinked copies of the same or very similar regions. At least three of these are located near telomeres, and two have been shown to be used for the expression of known Trypanosoma equiperdum VSG genes. Like VSG genes, the upstream sequences themselves can be duplicated and deleted. The choice of expression site to be used by a duplicated VSG gene is nonrandom; the site used for expression of the parental VSG gene is strongly favored for use in the daughter variant. Furthermore, even when the parental expression site is not used, the VSG gene occupying it is replaced. Thus, an active expression site is a preferential target for gene conversion in the next variation event.

  19. Conformation of human leucocyte antigen-C molecules at the surface of human trophoblast cells.

    Science.gov (United States)

    Apps, Richard; Gardner, Lucy; Hiby, Sue E; Sharkey, Andrew M; Moffett, Ashley

    2008-07-01

    Human leucocyte antigen (HLA)-C is expressed at lower levels than other classical HLA-I molecules on somatic cells. Surface HLA-C proteins can occur as conventionally beta(2)-microglobulin (beta2m)-associated complexes or as open conformers dissociated from peptide and/or beta(2)m. We investigated the conformation of HLA-C molecules on normal human trophoblast cells, which invade the maternal decidua during placentation. A panel of monoclonal antibodies to different conformations of HLA-I molecules was used in flow cytometry and surface immunoprecipitation experiments. On the surface of trophoblast cells only beta(2)m-associated complexes of HLA-C molecules were detected. In contrast, both open conformers and beta(2)m-associated HLA-C could be detected on other cells from the decidua, HLA-C-transfectants and cell lines. The levels of HLA-C expressed on primary trophoblast cells could be detected by antibodies specific to non-beta(2)m-associated conformations because binding was seen after acid-induced denaturation of surface proteins. In contrast to HLA-G molecules on trophoblasts, we found no evidence for the presence of disulphide-linked multimers of HLA-C complexes. These results show that most HLA-C molecules present at the trophoblast cell surface are in the conventional beta(2)m-associated conformation. These findings have implications regarding the stability of trophoblast HLA-C molecules and how they interact with receptors on decidual leucocytes during placentation.

  20. Mechanisms of Surface Antigenic Variation in the Human Pathogenic Fungus Pneumocystis jirovecii

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    Emanuel Schmid-Siegert

    2017-11-01

    Full Text Available Microbial pathogens commonly escape the human immune system by varying surface proteins. We investigated the mechanisms used for that purpose by Pneumocystis jirovecii. This uncultivable fungus is an obligate pulmonary pathogen that in immunocompromised individuals causes pneumonia, a major life-threatening infection. Long-read PacBio sequencing was used to assemble a core of subtelomeres of a single P. jirovecii strain from a bronchoalveolar lavage fluid specimen from a single patient. A total of 113 genes encoding surface proteins were identified, including 28 pseudogenes. These genes formed a subtelomeric gene superfamily, which included five families encoding adhesive glycosylphosphatidylinositol (GPI-anchored glycoproteins and one family encoding excreted glycoproteins. Numerical analyses suggested that diversification of the glycoproteins relies on mosaic genes created by ectopic recombination and occurs only within each family. DNA motifs suggested that all genes are expressed independently, except those of the family encoding the most abundant surface glycoproteins, which are subject to mutually exclusive expression. PCR analyses showed that exchange of the expressed gene of the latter family occurs frequently, possibly favored by the location of the genes proximal to the telomere because this allows concomitant telomere exchange. Our observations suggest that (i the P. jirovecii cell surface is made of a complex mixture of different surface proteins, with a majority of a single isoform of the most abundant glycoprotein, (ii genetic mosaicism within each family ensures variation of the glycoproteins, and (iii the strategy of the fungus consists of the continuous production of new subpopulations composed of cells that are antigenically different.

  1. Amino acid substitutions at positions 122 and 145 of hepatitis B virus surface antigen (HBsAg) determine the antigenicity and immunogenicity of HBsAg and influence in vivo HBsAg clearance.

    Science.gov (United States)

    Wu, Chunchen; Deng, Wanyu; Deng, Liu; Cao, Liang; Qin, Bo; Li, Songxia; Wang, Yun; Pei, Rongjuan; Yang, Dongliang; Lu, Mengji; Chen, Xinwen

    2012-04-01

    A variety of amino acid substitutions, such as K122I and G145R, have been identified around or within the a determinant of hepatitis B surface antigen (HBsAg), impair HBsAg secretion and antibody binding, and may be responsible for immune escape in patients. In this study, we examined how different substitutions at amino acid positions 122 and 145 of HBsAg influence HBsAg expression, secretion, and recognition by anti-HBs antibodies. The results showed that the hydrophobicity, the presence of the phenyl group, and the charges in the side chain of the amino acid residues at position 145 reduced HBsAg secretion and impaired reactivity with anti-HBs antibodies. Only the substitution K122I at position 122 affected HBsAg secretion and recognition by anti-HBs antibodies. Genetic immunization in mice demonstrated that the priming of anti-HBs antibody response was strongly impaired by the substitutions K122I, G145R, and others, like G145I, G145W, and G145E. Mice preimmunized with wild-type HBsAg (wtHBsAg) or variant HBsAg (vtHBsAg) were challenged by hydrodynamic injection (HI) with a replication-competent hepatitis B virus (HBV) clone. HBsAg persisted in peripheral blood for at least 3 days after HI in mice preimmunized with vtHBsAg but was undetectable in mice preimmunized with wtHBsAg, indicating that vtHBsAgs fail to induce proper immune responses for efficient HBsAg clearance. In conclusion, the biochemical properties of amino acid residues at positions 122 and 145 of HBsAg have a major effect on antigenicity and immunogenicity. In addition, the presence of proper anti-HBs antibodies is indispensable for the neutralization and clearance of HBsAg during HBV infection.

  2. cDNA sequence analysis of a 29-kDa cysteine-rich surface antigen of pathogenic Entamoeba histolytica

    Energy Technology Data Exchange (ETDEWEB)

    Torian, B.E.; Stroeher, V.L.; Stamm, W.E. (Univ. of Washington, Seattle (USA)); Flores, B.M. (Louisiana State Univ. Medical Center, New Orleans (USA)); Hagen, F.S. (Zymogenetics Incorporated, Seattle, WA (USA))

    1990-08-01

    A {lambda}gt11 cDNA library was constructed from poly(U)-Spharose-selected Entamoeba histolytica trophozoite RNA in order to clone and identify surface antigens. The library was screened with rabbit polyclonal anti-E. histolytica serum. A 700-base-pair cDNA insert was isolated and the nucleotide sequence was determined. The deduced amino acid sequence of the cDNA revealed a cysteine-rich protein. DNA hybridizations showed that the gene was specific to E. histolytica since the cDNA probe reacted with DNA from four axenic strains of E. histolytica but did not react with DNA from Entamoeba invadens, Acanthamoeba castellanii, or Trichomonas vaginalis. The insert was subcloned into the expression vector pGEX-1 and the protein was expressed as a fusion with the C terminus of glutathione S-transferase. Purified fusion protein was used to generate 22 monoclonal antibodies (mAbs) and a mouse polyclonal antiserum specific for the E. histolytica portion of the fusion protein. A 29-kDa protein was identified as a surface antigen when mAbs were used to immunoprecipitate the antigen from metabolically {sup 35}S-labeled live trophozoites. The surface location of the antigen was corroborated by mAb immunoprecipitation of a 29-kDa protein from surface-{sup 125}I-labeled whole trophozoites as well as by the reaction of mAbs with live trophozoites in an indirect immunofluorescence assay performed at 4{degree}C. Immunoblotting with mAbs demonstrated that the antigen was present on four axenic isolates tested. mAbs recognized epitopes on the 29-kDa native antigen on some but not all clinical isolates tested.

  3. Circulating hepatitis B surface antigen particles carry hepatocellular microRNAs.

    Directory of Open Access Journals (Sweden)

    Luisa Novellino

    Full Text Available Hepatitis B virus (HBV produces high quantities of subviral surface antigen particles (HBsAg which circulate in the blood outnumbering virions of about 1\\10(3-6 times. In individuals coinfected with the defective hepatitis Delta virus (HDV the small HDV-RNA-genome and Delta antigen circulate as ribonucleoprotein complexes within HBsAg subviral particles. We addressed the question whether subviral HBsAg particles may carry in the same way cellular microRNAs (miRNAs which are released into the bloodstream within different subcellular forms such as exosomes and microvescicles. Circulating HBsAg particles were isolated from sera of 11 HBsAg carriers by selective immunoprecipitation with monoclonal anti-HBs-IgG, total RNA was extracted and human miRNAs were screened by TaqMan real-time quantitative PCR Arrays. Thirty-nine human miRNAs were found to be significantly associated with the immunoprecipitated HBsAg, as determined by both comparative DDCT analysis and non-parametric tests (Mann-Whitney, p<0.05 with respect to controls. Moreover immunoprecipitated HBsAg particles contained Ago2 protein that could be revealed in ELISA only after 0.5% NP40. HBsAg associated miRNAs were liver-specific (most frequent = miR-27a, miR-30b, miR-122, miR-126 and miR-145 as well as immune regulatory (most frequent = miR-106b and miR-223. Computationally predicted target genes of HBsAg-associated miRNAs highlighted molecular pathways dealing with host-pathogen. The finding that HBsAg particles carry selective pools of hepatocellular miRNAs opens new avenues of research to disentangle the complex interactions between host and HBV and provides a non invasive tool to study the physiopathology of liver epigenetics.

  4. Identification of new meningococcal serogroup B surface antigens through a systematic analysis of neisserial genomes.

    Science.gov (United States)

    Pajon, Rolando; Yero, Daniel; Niebla, Olivia; Climent, Yanet; Sardiñas, Gretel; García, Darién; Perera, Yasser; Llanes, Alejandro; Delgado, Maité; Cobas, Karem; Caballero, Evelin; Taylor, Stephen; Brookes, Charlotte; Gorringe, Andrew

    2009-12-11

    The difficulty of inducing an effective immune response against the Neisseria meningitidis serogroup B capsular polysaccharide has lead to the search for vaccines for this serogroup based on outer membrane proteins. The availability of the first meningococcal genome (MC58 strain) allowed the expansion of high-throughput methods to explore the protein profile displayed by N. meningitidis. By combining a pan-genome analysis with an extensive experimental validation to identify new potential vaccine candidates, genes coding for antigens likely to be exposed on the surface of the meningococcus were selected after a multistep comparative analysis of entire Neisseria genomes. Eleven novel putative ORF annotations were reported for serogroup B strain MC58. Furthermore, a total of 20 new predicted potential pan-neisserial vaccine candidates were produced as recombinant proteins and evaluated using immunological assays. Potential vaccine candidate coding genes were PCR-amplified from a panel of representative strains and their variability analyzed using maximum likelihood approaches for detecting positive selection. Finally, five proteins all capable of inducing a functional antibody response vs N. meningitidis strain CU385 were identified as new attractive vaccine candidates: NMB0606 a potential YajC orthologue, NMB0928 the neisserial NlpB (BamC), NMB0873 a LolB orthologue, NMB1163 a protein belonging to a curli-like assembly machinery, and NMB0938 (a neisserial specific antigen) with evidence of positive selection appreciated for NMB0928. The new set of vaccine candidates and the novel proposed functions will open a new wave of research in the search for the elusive neisserial vaccine.

  5. Experimental verification of the identity of variant-specific surface proteins in Giardia lamblia trophozoites.

    Science.gov (United States)

    Li, Wei; Saraiya, Ashesh A; Wang, Ching C

    2013-05-21

    The cell membrane of a Giardia lamblia trophozoite is covered with a single species of variant-specific surface protein (VSP) that is replaced by another VSP every 6 to 13 generations of cell growth, possibly for an evasion of host immunity. Experimentally, only six VSP species have been verified to localize to the cell membrane thus far. By assuming that VSP contains multiple CXXC motifs, 219 vsp genes were annotated in GiardiaDB of the WB isolate. By further assuming that VSP possesses both CXXC motifs and a CRGKA tail at the C terminus, Adam et al. (BMC Genomics 11:424, 2010) identified a total of 303 potential vsp genes in Giardia WB. The discrepancies between these two assumed VSP identities have caused some confusion. Here, we used experimental approaches to further verify what is required of the structures of a VSP to localize to the surface of cell membrane. The data led to the following conclusions. (i) The C-terminal CRGKA sequence is not essential for localizing VSPs to the cell membrane. (ii) A "motif 1" of 45 residues, consisting of two CXXCs separated by 12 to 15 amino acid residues, located close to the C terminus and a hydrophobic "motif 2" of 38 residues at the C terminus are both essential and sufficient for localizing the protein to the cell membrane. (ii) An N-terminal sequence upstream from motif 1 is not required for targeting VSPs to the cell membrane. By these criteria, we are able to identify 73 open reading frames as the putative vsp genes in Giardia. IMPORTANCE The intestinal pathogen Giardia lamblia expresses only one variant-specific surface protein (VSP) on the cell membrane surface at a given time, but it changes spontaneously every 6 to 13 generations of growth, presumably for evading the host immunity. Only 6 VSPs have been empirically shown to localize to the cell membrane surface thus far. Here, we used mutations of VSPs and methods of identifying their locations in Giardia cells and found that a "motif 1" of 45 residues

  6. Measurement of Hepatitis B Surface Antigen Concentrations Using a Piezoelectric Microcantilever as a Mass Sensor

    Directory of Open Access Journals (Sweden)

    Sangkyu Lee

    2012-01-01

    Full Text Available Hepatitis B surface antigen (HBsAg concentrations were measured using a piezoelectric microcantilever sensor (PEMS developed by the authors. The developed PEMS is label-free and detects the sensing signal electrically. It was designed to measure the mass of biomolecules attached to it using an accurate mass-microbalancing technique; its probe area is confined to the end of the cantilever, and its equivalent spring constant is relatively high to minimize the effect of changes in the surface stress when the biomolecules are attached to it. The “dip- and-dry” technique was used to enable the probe area of the sensor to react with reagents in controlled environmental conditions. HBsAg was detected by an immunoreaction whereas the reaction time, antibody density, and its area on the probe were kept at a constant level. The mass of the detected HBsAg was measured in the range of 0.1–100 ng/mL.

  7. Association between CTL precursor frequency to HLA-C mismatches and HLA-C antigen cell surface expression

    Directory of Open Access Journals (Sweden)

    Moshe eIsraeli

    2014-10-01

    Full Text Available Previous studies showed the relevance of the cytotoxic T cell precursor frequency assay (CTLp for prediction of the outcome of HLA mismatched hematopoietic cell transplantation (HCT. Recently it has been shown that HLA-C cell surface expression is correlated with virus specific cytotoxic T cell responses and viremia control in HIV patients.The aim of the current study was to investigate the association between HLA-C antigen expression and the CTLp frequency to the mismatched HLA-C antigen.In total 115 recipient–donor pairs, for whom a successful CTLp assay was performed, were evaluated for this pilot study. All donor-recipient pairs were matched at 9/10 alleles with a single mismatch at the HLA-C locus. Antigen expression level of the mismatched HLA-C allele for each recipient and donor was based on the MFI values as described by Apps et al (Science, 2013.The cell surface expression of recipient’s mismatched HLA-C antigen was significantly lower among CTLp negative (n=59 compared to CTLp positive (n=56 pairs (154 and 193 MFI units, respectively; p=0.0031. This difference was more pronounced in donor-recipient pairs that were mismatched for amino-acid residue-116 located in the groove of the HLA-C antigen, suggesting the importance of peptide binding in the allo-recognition. Furthermore, in the particular case of low expression of the recipient mismatched HLA-C antigen (MFI<115, CTLp reactivity depended on HLA-C expression level in the donor; the median MFI of donor’s mismatched HLA-C antigen was 114 in CTLp negative cases (n=26, while in CTLp positive cases (n=15 the median MFI of donor’s HLA-C antigen was 193. (P=0.0093.We conclude that the expression level of the donor and recipient mismatched HLA-C antigens affect CTLp outcome. HLA-C antigen expression levels in combination with the CTLp assay may prove useful for the prediction of the clinical outcome of HLA-C mismatched HCT.

  8. Maternal serum alpha fetoprotein concentrations in mid trimester in hepatitis B surface antigen positive and negative subjects.

    Science.gov (United States)

    Kitau, M J; Grint, P; Heath, R; Chard, T

    1987-11-01

    Maternal serum alpha fetoprotein (AFP) was measured as part of a routine antenatal screening programme in 48 patients positive for hepatitis B surface antigen (HBsAg). After exclusion of two cases with obvious obstetric abnormalities there was no difference in AFP concentrations between subjects who were positive or negative for HBsAg.

  9. Hepatitis B surface antigen fusions delivered by DNA vaccination elicit CTL responses to human papillomavirus oncoproteins associated with tumor protection.

    Science.gov (United States)

    Haigh, O; Kattenbelt, J; Cochrane, M; Thomson, S; Gould, A; Tindle, R

    2010-10-01

    We describe the construction and evaluation of a recombinant hepatitis B surface antigen (HBsAg)-vectored DNA vaccine encoding the E7 and E6 tumor-associated oncoproteins of human papillomavirus (HPV) type 16. We show the induction of effector and memory cytotoxic T lymphocyte responses to E7 and E6 class I-restricted epitopes after a single immunization, which were associated with tumor prevention and therapy. The findings vindicate the use of a HBsAg-based DNA vaccine as a vehicle to elicit responses to co-encoded tumor antigens, and have specific implications for the development of a therapeutic vaccine for HPV-associated squamous carcinomas.

  10. Fractionation of T cell subsets on Ig anti-Ig columns: isolation of helper T cells from nonresponder mice, demonstration of antigen-specific T suppressor cells, and selection of CD-3 negative variants of Jurkat T cells

    DEFF Research Database (Denmark)

    Rubin, B; Geisler, C; Kuhlmann, J

    1989-01-01

    In the present experiments we have explored the possibilities of a modified immunoadsorbent technique to select for (1) mutagenized T cell receptor (Tcr) negative variants of Jurkat T lymphoma cells and (2) purified CD-4+ or CD-8+ T lymphocytes. The basic principle was to make large numbers...... of immunoglobulin (Ig) negative T cells Ig+ by T cell subset-specific monoclonal antibodies (mAb), and to select such cells on Ig anti-Ig columns. Our results demonstrated that Thy-1+, Fc receptor positive, antigen-specific T cells regulate the immune response in mice nonresponders to pork insulin......." The most important finding is the demonstration of antigen-specific Thy-1+, CD-8+, and Fc receptor+ T suppressor cell that apparently react with antigen in a non-major histocompatibility complex-restricted manner....

  11. Immunogenic Eimeria tenella glycosylphosphatidylinositol-anchored surface antigens (SAGs induce inflammatory responses in avian macrophages.

    Directory of Open Access Journals (Sweden)

    Yock-Ping Chow

    Full Text Available At least 19 glycosylphosphatidylinositol (GPI-anchored surface antigens (SAGs are expressed specifically by second-generation merozoites of Eimeria tenella, but the ability of these proteins to stimulate immune responses in the chicken is unknown.Ten SAGs, belonging to two previously defined multigene families (A and B, were expressed as soluble recombinant (r fusion proteins in E. coli. Chicken macrophages were treated with purified rSAGs and changes in macrophage nitrite production, and in mRNA expression profiles of inducible nitric oxide synthase (iNOS and of a panel of cytokines were measured. Treatment with rSAGs 4, 5, and 12 induced high levels of macrophage nitric oxide production and IL-1β mRNA transcription that may contribute to the inflammatory response observed during E. tenella infection. Concomitantly, treatment with rSAGs 4, 5 and 12 suppressed the expression of IL-12 and IFN-γ and elevated that of IL-10, suggesting that during infection these molecules may specifically impair the development of cellular mediated immunity.In summary, some E. tenella SAGs appear to differentially modulate chicken innate and humoral immune responses and those derived from multigene family A (especially rSAG 12 may be more strongly linked with E. tenella pathogenicity associated with the endogenous second generation stages.

  12. Quantitative Measurement of Serum Hepatitis B Surface Antigen Using an Immunoradiometric Assay in Chronic Hepatitis B

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, Hyun Woo; Lee, Ho Young; Kim, Seog Gyun; Kim, Won; Jung, Wong Jin; Kang, Keon Wook; Chung, June Key; Lee, Myung Chul; Lee, Dong Soo [Seoul National Univ. Seoul (Korea, Republic of)

    2011-03-15

    Measurement of serum hepatitis B virus surface antigen (HBsAg) levels is important for the management of chronic hepatitis D patients in terms of monitoring response to antiviral therapy. This study aimed to evaluate the diagnostic performance of a new diagnostic kit, which quantitatively measures serum HBsAg level using an immunoradiometric assay (IRMA) based method. Measurements were compared with those obtained using a chemiluminescent microparticle immunoassay (CMIA) based method. The blood samples of 96 patients with chronic hepatitis B were used in this study. Copy numbers of serum hepatitis B virus (HBV) DNA were determined in 23 of these samples. The correlation between and the concordance of IRMA and CMIA results were determined using Pearson's correlation coefficients. P values of 0.05 were considered to be statistically significant throughout. Laboratory diagnoses based on CMIA. Furthermors, serum HBsAg levels by IRMA were found to be highly correlated with those determined by CMIA (correlation coefficient R{sup 2=}0.838, P<0.001). Serum HBsAg level and serum HBV DNA copies were found to be linearly related by both methods (R{sup 2=}0.067, P=0.316 by IRMA, and R{sup 2=}0.101, P=0.215 by CMIA). The diagnostic performance of the investigated IRMA method of determining HBsAg levels was found to be comparable with that of a CMIA based method in chronic hepatitis B patients.

  13. The Rickettsia Surface Cell Antigen 4 Applies Mimicry to Bind to and Activate Vinculin*

    Science.gov (United States)

    Park, HaJeung; Lee, Jun Hyuck; Gouin, Edith; Cossart, Pascale; Izard, Tina

    2011-01-01

    Pathogenic Rickettsia species cause high morbidity and mortality, especially R. prowazekii, the causative agent of typhus. Like many intracellular pathogens, Rickettsia exploit the cytoskeleton to enter and spread within the host cell. Here we report that the cell surface antigen sca4 of Rickettsia co-localizes with vinculin in cells at sites of focal adhesions in sca4-transfected cells and that sca4 binds to and activates vinculin through two vinculin binding sites (VBSs) that are conserved across all Rickettsia. Remarkably, this occurs through molecular mimicry of the vinculin-talin interaction that is also seen with the IpaA invasin of the intracellular pathogen Shigella, where binding of these VBSs to the vinculin seven-helix bundle head domain (Vh1) displaces intramolecular interactions with the vinculin tail domain that normally clamp vinculin in an inactive state. Finally, the vinculin·sca4-VBS crystal structures reveal that vinculin adopts a new conformation when bound to the C-terminal VBS of sca4. Collectively, our data define the mechanism by which sca4 activates vinculin and interacts with the actin cytoskeleton, and they suggest important roles for vinculin in Rickettsia pathogenesis. PMID:21841197

  14. In vivo gene transfer targeting in pancreatic adenocarcinoma with cell surface antigens

    Directory of Open Access Journals (Sweden)

    Lafitte Marie

    2012-10-01

    Full Text Available Abstract Background Pancreatic ductal adenocarcinoma is a deadly malignancy resistant to current therapies. It is critical to test new strategies, including tumor-targeted delivery of therapeutic agents. This study tested the possibility to target the transfer of a suicide gene in tumor cells using an oncotropic lentiviral vector. Results Three cell surface markers were evaluated to target the transduction of cells by lentiviruses pseudotyped with a modified glycoprotein from Sindbis virus. Only Mucin-4 and the Claudin-18 proteins were found efficient for targeted lentivirus transductions in vitro. In subcutaneous xenografts of human pancreatic cancer cells models, Claudin-18 failed to achieve efficient gene transfer but Mucin-4 was found very potent. Human pancreatic tumor cells were modified to express a fluorescent protein detectable in live animals by bioimaging, to perform a direct non invasive and costless follow up of the tumor growth. Targeted gene transfer of a bicistronic transgene bearing a luciferase gene and the herpes simplex virus thymidine kinase gene into orthotopic grafts was carried out with Mucin-4 oncotropic lentiviruses. By contrast to the broad tropism VSV-G carrying lentivirus, this oncotropic lentivirus was found to transduce specifically tumor cells, sparing normal pancreatic cells in vivo. Transduced cells disappeared after ganciclovir treatment while the orthotopic tumor growth was slowed down. Conclusion This work considered for the first time three aspect of pancreatic adenocarcinoma targeted therapy. First, lentiviral transduction of human pancreatic tumor cells was possible when cells were grafted orthotopically. Second, we used a system targeting the tumor cells with cell surface antigens and sparing the normal cells. Finally, the TK/GCV anticancer system showed promising results in vivo. Importantly, the approach presented here appeared to be a safer, much more specific and an as efficient way to perform gene

  15. Serum hepatitis B core-related antigen is more accurate than hepatitis B surface antigen to identify inactive carriers, regardless of hepatitis B virus genotype.

    Science.gov (United States)

    Riveiro-Barciela, M; Bes, M; Rodríguez-Frías, F; Tabernero, D; Ruiz, A; Casillas, R; Vidal-González, J; Homs, M; Nieto, L; Sauleda, S; Esteban, R; Buti, M

    2017-11-01

    To investigate whether hepatitis B surface antigen (HBsAg) and hepatitis B core-related antigen (HBcrAg) levels are useful to identify inactive carriers among HBeAg-negative patients infected by different hepatitis B virus (HBV) genotypes. In all, 202 consecutive HBeAg-negative patients with chronic hepatitis B, 135 inactive carriers and 67 with HBV activity, were prospectively followed for 1 year. In HBeAg-negative patients, HBsAg levels differed across the different genotypes (p 85% in all HBV genotypes, except genotype H or F, with values of 62.5% and 72.7%, respectively, for the two parameters. HBsAg levels varied across genotypes in HBeAg-negative patients. HBsAg levels <3 logIU/mL were only useful for identifying genotype D inactive carriers. A single HBcrAg measurement ≤3 logU/mL plus HBV DNA ≤2000 IU/mL was highly accurate for identifying inactive carriers, regardless of their HBV genotype. Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  16. Immunoelectron microscopic identification and localization of Streptococcus sanguis with peroxidase-labeled antibody: localization of surface antigens in pure cultures.

    Science.gov (United States)

    Lai, C H; Listgarten, M A; Rosan, B

    1975-01-01

    An indirect method of localizing antigens with horseradish peroxidase-labeled antibody was used to identify and localize surface antigens of Streptococcus sanguis at the ultrastructural level. An electron dense layer surrounding the cell wall could be distinguished without any additional electron microscope staining. This labeled layer represents an immune complex consisting of bacterial surface antigens, specific rabbit antisera, and peroxidase-labeled goat anti-rabbit globulins. Although with undiluted antisera slight cross-reactions occurred with S. salivarius and S. miteor (mitis), these could be readily distinguished from the more intense homologous reaction by their patchiness and the difference in distribution of the label. These cross-reactions were eliminated by appropriate dilutions of the antiserum. No cross-reactions occurred with S. mutans, S. faecalis, Actinomyces species, or Bacterionema, microorganisms wents indicated that horseradish peroxidase can become non-specifically adsorbed to the membrane of certain bacterial cells. Appropriate controls must, therefore, be included for localization of membrane associated antigens with horseradish peroxidase. Images PMID:1090524

  17. The use of incomplete genes for the construction of a Trypanosoma equiperdum variant surface glycoprotein gene.

    Science.gov (United States)

    Roth, C W; Longacre, S; Raibaud, A; Baltz, T; Eisen, H

    1986-05-01

    The expression of Trypanosoma equiperdum variant surface protein (VSG) 78 is accomplished by the duplicative transposition of silent basic copy (BC) genes into a telomer-linked expression site to form an expression-linked copy (ELC). In two independent isolates expressing VSG 78, the ELC is a composite gene. The analysis of VSG 78 cDNA clones from these two Bo Tat 78 isolates and the respective BC genes revealed that both ELCs were constructed from the same three BC genes, a 3' BC which donated the last 255 bp of each ELC and two closely related 5' BCs. Although sequences of both 5' BC genes were found in each ELC, the junction with the 3' BC was provided by the same 5' BC in both cases. This 5' BC is an incomplete gene with insufficient open reading frame to code for a complete VSG and thus can only be used when joined to a competent 3' end. Furthermore, both 5' BC genes lack a conserved 14 nucleotide sequence found on all VSG mRNAs. These results support a model in which composite gene formation plays a role in the determination of the order of VSG expression. They also illustrate similarities between immunoglobulin gene and VSG gene construction.

  18. Vaccine Containing the Three Allelic Variants of the Plasmodium vivax Circumsporozoite Antigen Induces Protection in Mice after Challenge with a Transgenic Rodent Malaria Parasite

    Directory of Open Access Journals (Sweden)

    Alba Marina Gimenez

    2017-10-01

    Full Text Available Plasmodium vivax is the most common species that cause malaria outside of the African continent. The development of an efficacious vaccine would contribute greatly to control malaria. Recently, using bacterial and adenoviral recombinant proteins based on the P. vivax circumsporozoite protein (CSP, we demonstrated the possibility of eliciting strong antibody-mediated immune responses to each of the three allelic forms of P. vivax CSP (PvCSP. In the present study, recombinant proteins representing the PvCSP alleles (VK210, VK247, and P. vivax-like, as well as a hybrid polypeptide, named PvCSP-All epitopes, were generated. This hybrid containing the conserved C-terminal of the PvCSP and the three variant repeat domains in tandem were successfully produced in the yeast Pichia pastoris. After purification and biochemical characterization, they were used for the experimental immunization of C57BL/6 mice in a vaccine formulation containing the adjuvant Poly(I:C. Immunization with a recombinant protein expressing all three different allelic forms in fusion elicited high IgG antibody titers reacting with all three different allelic variants of PvCSP. The antibodies targeted both the C-terminal and repeat domains of PvCSP and recognized the native protein on the surface of P. vivax sporozoites. More importantly, mice that received the vaccine formulation were protected after challenge with chimeric Plasmodium berghei sporozoites expressing CSP repeats of P. vivax sporozoites (Pb/PvVK210. Our results suggest that it is possible to elicit protective immunity against one of the most common PvCSP alleles using soluble recombinant proteins expressed by P. pastoris. These recombinant proteins are promising candidates for clinical trials aiming to develop a multiallele vaccine against P. vivax malaria.

  19. Experimental Verification of the Identity of Variant-Specific Surface Proteins in Giardia lamblia Trophozoites

    Science.gov (United States)

    Li, Wei; Saraiya, Ashesh A.; Wang, Ching C.

    2013-01-01

    ABSTRACT The cell membrane of a Giardia lamblia trophozoite is covered with a single species of variant-specific surface protein (VSP) that is replaced by another VSP every 6 to 13 generations of cell growth, possibly for an evasion of host immunity. Experimentally, only six VSP species have been verified to localize to the cell membrane thus far. By assuming that VSP contains multiple CXXC motifs, 219 vsp genes were annotated in GiardiaDB of the WB isolate. By further assuming that VSP possesses both CXXC motifs and a CRGKA tail at the C terminus, Adam et al. (BMC Genomics 11:424, 2010) identified a total of 303 potential vsp genes in Giardia WB. The discrepancies between these two assumed VSP identities have caused some confusion. Here, we used experimental approaches to further verify what is required of the structures of a VSP to localize to the surface of cell membrane. The data led to the following conclusions. (i) The C-terminal CRGKA sequence is not essential for localizing VSPs to the cell membrane. (ii) A “motif 1” of 45 residues, consisting of two CXXCs separated by 12 to 15 amino acid residues, located close to the C terminus and a hydrophobic “motif 2” of 38 residues at the C terminus are both essential and sufficient for localizing the protein to the cell membrane. (ii) An N-terminal sequence upstream from motif 1 is not required for targeting VSPs to the cell membrane. By these criteria, we are able to identify 73 open reading frames as the putative vsp genes in Giardia. PMID:23695837

  20. Clinical performance of a new hepatitis B surface antigen quantitative assay with automatic dilution

    Directory of Open Access Journals (Sweden)

    Ta-Wei Liu

    2015-01-01

    Full Text Available Hepatitis B virus surface antigen (HBsAg levels reflect disease status and can predict the clinical response to antiviral treatment; however, the emergence of HBsAg mutant strains has become a challenge. The Abbott HBsAg quantification assay provides enhanced detection of HBsAg and HBsAg mutants. We aimed to evaluate the performance of the Abbott HBsAg quantification assay with automatic sample dilutions (shortened as automatic Architect assay, compared with the Abbott HBsAg quantification assay with manual sample dilutions (shortened as manual Architect assay and the Roche HBsAg quantification assay with automatic sample dilutions (shortened as Elecsys. A total of 130 sera samples obtained from 87 hepatitis B virus (HBV-infected patients were collected to assess the correlation between the automatic and manual Architect assays. Among the 87 patients, 41 provided 42 sera samples to confirm the linearity and reproducibility of the automatic Architect assay, and find out the correlation among the Elecsys and two Architect assays. The coefficients of variation (0.44–9.53% and R2 = 0.996–1, which were both determined using values obtained from the automatic Architect assay, showed good reproducibility and linearity. Results of the two Architect assays demonstrated a feasible correlation (n = 130 samples; R = 0.898, p  0.93 in all cases. In conclusion, the correlation between the automatic and manual dilution Architect assays was feasible, particularly in the HBeAg-negative and low DNA groups. With lower labor costs and less human error than the manual version, the Abbott automatic dilution Architect assay provided a good clinical performance with regard to the HBsAg levels.

  1. Prevalence of Hepatitis B Surface Antigen in Pregnant Women in Beheshti Hospital of Kashan, Isfahan.

    Science.gov (United States)

    Afzali, Hasan; Momen Heravi, Mansooreh; Moravveji, Seyyed Alireza; Poorrahnama, Maryam

    2015-07-01

    The transmission of hepatitis B virus (HBV) is parenteral, sexual and prenatal. Prevention of vertical transmission of HBV is extremely important, because HBV infection in early life usually results in a chronic carrier state. There has been so much debate about hepatitis B surface antigen (HBsAg) screening in pregnant women. The aim of this study was to determine the prevalence of HBsAg+ among pregnant women referred to Beheshti hospital in Kashan in 2012. This descriptive study was carried out on 768 pregnant women, hospitalized in Beheshti Hospital of Kashan in 2012. After obtaining consent forms, the questionnaires including demographic and HBV infection-associated risk factors were filled through interview and then 5 mL blood was taken from each patient and HBsAg was examined by the enzyme-linked immunosorbent assay (ELISA) method. These data were analyzed by statistical package for the social science (SPSS) software. A total of 12 (1.56%) out of 768 pregnant women were HBsAg+. The mean age of HBsAg+ cases was 24.5 ± 4 years. Most of the HBsAg+ cases (66.6%) were uneducated; 17.7% of the pregnant women were not Iranian, of which 7.4% were HBsAg+. There was no high-risk job, recent dentistry interruption or skin tattoo among the HBsAg+ cases. In this study, 1.56% of pregnant women were HBsAg+, which was higher than the previous studies. This increasing prevalence may be due to the increase of non-Iranians' migrations to Iran. Control of migration and screening and vaccination of these groups should be considered by health policy makers.

  2. Hepatitis B genotypes and surface antigen mutants present in Pakistani blood donors.

    Directory of Open Access Journals (Sweden)

    Barbara J Harris

    Full Text Available The prevalence of chronic Hepatitis B Virus (HBV infection is 2-4% in the Pakistani population, defining Pakistan as an intermediate prevalence country. In this study, hepatitis B surface antigen (HBsAg reactive blood donations were screened using a combination of serological and molecular methods to identify immune escape HBV mutant strains and to determine the HBV genotypes and subtypes present in Pakistan.Blood donations were collected at the Armed Forces Institute of Transfusion (AFIT located in northern Pakistan and the Hussaini Blood Bank (HBB located in the south. From 2009 to 2013 a total of 706,575 donations were screened with 2.04% (14,409 HBsAg reactive. A total of 2055 HBsAg reactive specimens, were collected and screened using a monoclonal antibody based research assay to identify immune escape mutants followed by PCR amplification and DNA sequencing to identify the mutation present. DNA sequences obtained from 192 specimens, including mutant candidates and wild type strains, were analyzed for escape mutations, genotype, and HBsAg subtype.Mutations were identified in approximately 14% of HBsAg reactive donations. Mutations at HBsAg amino acid positions 143-145 are the most common (46% with the mutation serine 143 to leucine the most frequently occurring change (28%. While regional differences were observed, the most prevalent HBV strains are subgenotypes of D with subgenotype D1/subtype ayw2 accounting for the majority of infections; 90.2% at AFIT and 52.5% at HBB.The high frequency of immune escape HBV mutants in HBV infected Pakistani blood donors highlights the need for more studies into the prevalence of escape mutants. Differences between vaccinated and unvaccinated populations, the correlation of escape mutant frequency with genotype, and impact of escape mutations in different genotype backgrounds on the performance of commercially available HBsAg assays represent avenues for further investigation.

  3. Prevalence of Hepatitis B surface antigen in children with sickle cell anemia

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    Baba Jibrin

    2014-01-01

    Full Text Available Background: Hepatitis B virus is known to be endemic in Africa. The seroepidemiological studies of HBV have shown that infection commonly occurs in childhood in Africa resulting in an increased tendency to chronicity. This cross-sectional study was undertaken to determine the seroprevalence of hepatitis B surface antigen among pediatric patients with homozygous hemoglobin S. Materials and Methods: Three hundred sickle cell anemia children aged 6 months-15 years (both in steady state and in crises attending the SCA clinic and on admission in emergency pediatrics unit and pediatrics medical ward, Usmanu Danfodiyo University Teaching Hospital, Sokoto, Nigeria, were screened for hepatitis B infection using HBsAg as marker of infection. The sensitive enzyme linked immunosorbent assay method was used for detection of the marker. Three hundred children with minor illness attending pediatrics outpatient department and on admission in EPU/PMW for various treatment in the same hospital served as gender- and age-marched controls cohorts. Results: The sero-prevalence of HBsAg seropositivity for hepatitis B virus infection among SCA children was 17.3% (52/300 compared to 10.7% (32/300 of the control (P = 0.0875. The peak prevalence age group for HBV infection among SCA children was in the age group 1.1-5.0 years (6% compared to 10.1-15.0 years (4.7% in the control. Risk factors for HBV infection such as blood transfusion, traditional scarification/circumcision/uvulectomy, and tattooing did not significantly affect the prevalence of HBV infection in both SCA children and controls. Conclusion: Hepatitis B infection is common in Sokoto. The need for strict adherence to HBV immunization and further community-based studies on the risk factors are recommended.

  4. Variation of expression defects in cell surface 190-kDa protein antigen of Streptococcus mutans.

    Science.gov (United States)

    Lapirattanakul, Jinthana; Nomura, Ryota; Matsumoto-Nakano, Michiyo; Srisatjaluk, Ratchapin; Ooshima, Takashi; Nakano, Kazuhiko

    2015-05-01

    Streptococcus mutans, which consists of four serotypes, c, e, f, and k, possesses a 190-kDa cell surface protein antigen (PA) for initial tooth adhesion. We used Western blot analysis to determine PA expression in 750 S. mutans isolates from 150 subjects and found a significantly higher prevalence of the isolates with PA expression defects in serotypes f and k compared to serotypes c and e. Moreover, the defect patterns could be classified into three types; no PA expression on whole bacterial cells and in their supernatant samples (Type N1), PA expression mainly seen in supernatant samples (Type N2), and only low expression of PA in the samples of whole bacterial cells (Type W). The underlying reasons for the defects were mutations in the gene encoding PA as well as in the transcriptional processing of this gene for Type N1, defects in the sortase gene for Type N2, and low mRNA expression of PA for Type W. Since cellular hydrophobicity and phagocytosis susceptibility of the PA-defective isolates were significantly lower than those of the normal expression isolates, the potential implication of such defective isolates in systemic diseases involving bacteremia other than dental caries was suggested. Additionally, multilocus sequence typing was utilized to characterize S. mutans clones that represented a proportion of isolates with PA defects of 65-100%. Therefore, we described the molecular basis for variation defects in PA expression of S. mutans. Furthermore, we also emphasized the strong association between PA expression defects and serotypes f and k as well as the clonal relationships among these isolates. Copyright © 2015 Elsevier GmbH. All rights reserved.

  5. Prevalence of hepatitis B surface antigen among refugees entering the United States between 2006 and 2008.

    Science.gov (United States)

    Rein, David B; Lesesne, Sarah B; O'Fallon, Ann; Weinbaum, Cindy M

    2010-02-01

    The Centers for Disease Control and Prevention recommends hepatitis B surface antigen (HBsAg) testing to identify chronic hepatitis B virus infection for foreign-born persons from countries or regions with HBsAg prevalence of >or=2%. However, limited data exist to indicate which countries meet this definition. To address this data gap, we estimated the HBsAg prevalence among refugees entering the United States between 2006 and 2008. We contacted state refugee health coordinators and asked them to report the number of refugees, country of origin, and HBsAg prevalence among refugees screened in their jurisdiction during the most recently available 12-month period prior to August 2008. We pooled data across jurisdictions and calculated the prevalence for any country with more than 30 refugees entering the United States, and where this level of data was not available by country, continents were considered. Of the 47 jurisdictions contacted, we received basic information from 31, with nine jurisdictions reporting HBsAg prevalence by country of origin applicable to 31,980 refugees (approximately 42% of refugees entering the United States during the observation period). We estimated an HBsAg prevalence of 2.8% (95% confidence interval 2.6%-3.0%) for refugees overall. Of the 37 countries with 30 or more refugees entering the United States, 25 had a prevalence of >or=2%. Prevalence was highest among refugees from Africa and Southeast Asia, and lowest among refugees from the Middle East and South/Central America. In the eight countries for which we had comparison data, six had lower HBsAg prevalence than in 1991.

  6. Serological responses to Cryptosporidium antigens in inhabitants of Hungary using conventionally filtered surface water and riverbank filtered drinking water.

    Science.gov (United States)

    Farkas, K; Plutzer, J; Moltchanova, E; Török, A; Varró, M J; Domokos, K; Frost, F; Hunter, P R

    2015-10-01

    In this study the putative protective seroprevalence (PPS) of IgG antibodies to the 27-kDa and 15/17-kDa Cryptosporidium antigens in sera of healthy participants who were and were not exposed to Cryptosporidium oocysts via surface water-derived drinking water was compared. The participants completed a questionnaire regarding risk factors that have been shown to be associated with infection. The PPS was significantly greater (49-61%) in settlements where the drinking water originated from surface water, than in the control city where riverbank filtration was used (21% and 23%). Logistic regression analysis on the risk factors showed an association between bathing/swimming in outdoor pools and antibody responses to the 15/17-kDa antigen complex. Hence the elevated responses were most likely due to the use of contaminated water. Results indicate that waterborne Cryptosporidium infections occur more frequently than reported but may derive from multiple sources.

  7. Study of Admission Rate of Hepatitis B Surface Antigen Positive Patients in 50 Dentistry Centers in Tehran (Spring 2003)

    OpenAIRE

    Seyed Moayed Alavian; Seyed-Hamed Moosavi; Seyed-Hadi Mousavi; Behzad Azizi; Hossein Akbari

    2008-01-01

    Background and Aims: Hepatitis B is one of the common diseases in the world and in Iran, caused by hepatitis B virus (HBV). This virus is transmitted by various ways including dentistry procedures and this may discourage dentists to admit hepatitis B surface antigen (HBsAg) positive patients leading to hide the disease by the patients and increased transmission risk. Methods: During a cross-sectional descriptive study, 50 dentistry centers were chosen among dentistry centers of Tehran in var...

  8. Prevalence of Hepatitis B Surface Antigen in US-Born and Foreign-Born Asian/Pacific Islander College Students

    Science.gov (United States)

    Quang, Yen N.; Vu, Joanne; Yuk, Jihey; Li, Chin-Shang; Chen, Moon; Bowlus, Christopher L.

    2010-01-01

    The prevalence of chronic hepatitis B (HBV) among college-age US-born Asian and Pacific Islanders (A/PI) is not well known. Objectives: To compare the prevalence of hepatitis B surface antigen (HBsAg) seropositivity in US-born to A/PI-born students at a public university. Participants: Undergraduate who self-identified themselves as A/PI. Results:…

  9. Freeze-Drying of Plant Tissue Containing HBV Surface Antigen for the Oral Vaccine against Hepatitis B

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    Marcin Czyż

    2014-01-01

    Full Text Available The aim of this study was to develop a freeze-drying protocol facilitating successful processing of plant material containing the small surface antigen of hepatitis B virus (S-HBsAg while preserving its VLP structure and immunogenicity. Freeze-drying of the antigen in lettuce leaf tissue, without any isolation or purification step, was investigated. Each process step was consecutively evaluated and the best parameters were applied. Several drying profiles and excipients were tested. The profile of 20°C for 20 h for primary and 22°C for 2 h for secondary drying as well as sucrose expressed efficient stabilisation of S-HBsAg during freeze-drying. Freezing rate and postprocess residual moisture were also analysed as important factors affecting S-HBsAg preservation. The process was reproducible and provided a product with VLP content up to 200 µg/g DW. Assays for VLPs and total antigen together with animal immunisation trials confirmed preservation of antigenicity and immunogenicity of S-HBsAg in freeze-dried powder. Long-term stability tests revealed that the stored freeze-dried product was stable at 4°C for one year, but degraded at elevated temperatures. As a result, a basis for an efficient freeze-drying process has been established and a suitable semiproduct for oral plant-derived vaccine against HBV was obtained.

  10. [Hepatitis B virus surface antigen reactivity in the absence of antibodies to core antigen: an atypical serological pattern having diverse significance].

    Science.gov (United States)

    Echevarría, José María; León, Pilar; Pozo, Francisco

    2004-01-01

    Reactivity for hepatitis B virus (HBV) surface antigen (HBsAg) in samples lacking antibody to the HBV core antigen (anti-HBc) may be due to either non-specific reactions or sample contamination, but it can also reflect other situations that may present clinical relevance. In Spain, the frequency described for this atypical pattern of HBV markers ranged from 0.05 to 1.3% in different series, so that it can be considered as moderately frequent. Confirmatory assays for HBsAg and tests for detecting total and IgM anti-HBc, HBV "e" antigen and viral DNA were performed on serum samples taken from 96 patients who showed reactivity for HBsAg in absence of total anti-HBc on routine studies. These samples were collected between January, 2001 and May, 2002 and were sent for study from different Spanish laboratories. Follow-up samples were also studied from selected cases. Presence of HBsAg was excluded in 70 cases (72.9%) and total anti-HBc was detected in two additional patients, so that the original results were confirmed in 24 patients (25.0%). After further investigations, two early phases of the window period of the acute HBV infection were identified, as well as a further case of chronic HBV carriage in absence of antibody response. A single case could be confirmed as due to a contamination of the aliquot of sample studied. The pattern of HBV markers and its evolution on the follow-up were the characteristic of the phenomenon known as "hepatitis B virus type 2 infection" in eight patients. No conclusions could be drawn from the remainder 12 cases, since no follow-up samples were available for study. Besides most cases respond to non-specific reactions or reflect situations of low clinical relevance, the reactivity for HBsAg in samples lacking anti-HBc should be taken into consideration and routinely investigated, since this atypical pattern can also reflect unusual, but clinically relevant facts of the HBV infection that must be noticed.

  11. Rapid Isolation of Monoclonal Antibodies Specific for Cell Surface Differentiation Antigens

    Science.gov (United States)

    Barclay, Stephen L.; Smith, Alan M.

    1986-06-01

    Two immunization procedures were compared for their ability to yield monoclonal antibodies that react with plasma membrane-bound differentiation antigens of Dictyostelium. In the first method, hybridomas prepared from BALB/c mice immunized with aggregating amoebae produced monoclonal antibodies that recognized antigens present on both growing and aggregating Dictyostelium amoebae. None of the monoclonal antibodies reacted with only the injected aggregation-stage cell type. In contrast, monoclonal antibodies that reacted with differentiation antigens were easily obtained by primary immunization of BALB/c mice with living aggregation-stage cells, followed by secondary immunization with a preparation of plasma membrane from aggregating cells or intact aggregating cells mixed with polyclonal BALB/c antiserum raised against undifferentiated cells. By this method, approximately 20% of all anti-Dictyostelium monoclonal antibodies obtained in a fusion are specific for differentiation antigens. The properties and developmental regulation of several of these antigens are described. The possible uses of this immunological method to detect unique determinants on other kinds of cells and the likely immune mechanisms that make it successful are discussed.

  12. The Severity of Chorioamnionitis in Pregnant Sheep Is Associated with In Vivo Variation of the Surface-Exposed Multiple-Banded Antigen/Gene of Ureaplasma parvum1

    Science.gov (United States)

    Knox, Christine L.; Dando, Samantha J.; Nitsos, Ilias; Kallapur, Suhas G.; Jobe, Alan H.; Payton, Diane; Moss, Timothy J.M.; Newnham, John P.

    2010-01-01

    Ureaplasma species are the bacteria most frequently isolated from human amniotic fluid in asymptomatic pregnancies and placental infections. Ureaplasma parvum serovars 3 and 6 are the most prevalent serovars isolated from men and women. We hypothesized that the effects on the fetus and chorioamnion of chronic ureaplasma infection in amniotic fluid are dependent on the serovar, dose, and variation of the ureaplasma multiple-banded antigen (MBA) and mba gene. We injected high- or low-dose U. parvum serovar 3, serovar 6, or vehicle intra-amniotically into pregnant ewes at 55 days of gestation (term = 150 days) and examined the chorioamnion, amniotic fluid, and fetal lung tissue of animals delivered by cesarean section at 125 days of gestation. Variation of the multiple banded antigen/mba generated by serovar 3 and serovar 6 ureaplasmas in vivo were compared by PCR assay and Western blot. Ureaplasma inoculums demonstrated only one (serovar 3) or two (serovar 6) MBA variants in vitro, but numerous antigenic variants were generated in vivo: serovar 6 passage 1 amniotic fluid cultures contained more MBA size variants than serovar 3 (P = 0.005), and ureaplasma titers were inversely related to the number of variants (P = 0.025). The severity of chorioamnionitis varied between animals. Low numbers of mba size variants (five or fewer) within amniotic fluid were associated with severe inflammation, whereas the chorioamnion from animals with nine or more mba variants showed little or no inflammation. These differences in chorioamnion inflammation may explain why not all women with in utero Ureaplasma spp. experience adverse pregnancy outcomes. PMID:20519696

  13. Trichinella britovi human infection in Spain : antibody response to surface, excretory/secretory and somatic antigens

    Directory of Open Access Journals (Sweden)

    Rodríguez-Osorio M.

    2003-06-01

    Full Text Available A third outbreak of Trichinella britovi with 140 people involved, occurred in Granada Spain (December 1998. The source of infection was sausage made from uninspected wild boar meat. Fifty-two patients agreed to participated in this study. An elevated eosinophil level (> 5 % was detected in 59.6 % of patients, and persisted in most of these cases for two months. A moderate IgG response was observed. At the onset of symptoms, Western blot (WB test detected more positive cases than Enzyme linked immunosorbent assay (ELISA and indirect immunofluorescence (IIF. Six months from infection, ELISA revealed fewer positive cases than the other two tests. It would appear that the response to somatic antigens starts earlier than those to cuticular and excretory/secretory (ES antigens and that the response to ES antigens is the first to decrease.

  14. Characterisation of a GII-4 norovirus variant-specific surface-exposed site involved in antibody binding

    Directory of Open Access Journals (Sweden)

    Gray Jim J

    2009-09-01

    Full Text Available Abstract Background The human noroviruses are a highly diverse group of viruses with a single-stranded RNA genome encoding a single major structural protein (VP1, which has a hypervariable domain (P2 domain as the most exposed part of the virion. The noroviruses are classified on the basis of nucleotide sequence diversity in the VP1-encoding ORF2 gene, which divides the majority of human noroviruses into two genogroups (GI and GII. GII-4 noroviruses are the major aetiological agent of outbreaks of gastroenteritis around the world. During a winter season the diversity among the GII-4 noroviruses has been shown to fluctuate, driving the appearance of new virus variants in the population. We have previously shown that sequence data and in silico modelling experiments suggest there are two surface-exposed sites (site A and site B in the hypervariable P2 domain. We predict these sites may form a functional variant-specific epitope that evolves under selective pressure from the host immune response and gives rise to antibody escape mutants. Results In this paper, we describe the construction of recombinant baculoviruses to express VLPs representing one pre-epidemic and one epidemic variant of GII-4 noroviruses, and the production of monoclonal antibodies against them. We use these novel reagents to provide evidence that site A and site B form a conformational, variant-specific, surface-exposed site on the GII-4 norovirus capsid that is involved in antibody binding. Conclusion As predicted by our earlier study, significant amino acid changes at site A and site B give rise to GII-4 norovirus epidemic variants that are antibody escape mutants.

  15. Middle-aged Subjects With Habitual Low-speed Cycling Exercise Have Greater Mononuclear Cell Responsiveness Against Human Hepatitis B Virus Surface Antigen

    Directory of Open Access Journals (Sweden)

    Kuen-Chang Hsieh

    2010-06-01

    Conclusion: The results reveal that the immune response of MNC, which are stimulated by PHA to suppress hepatitis B surface antigen expression, is greater in middle-aged subjects with low-speed HCE than in sedentary subjects.

  16. 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

  17. Escherichia coli Nissle 1917 bacterial ghosts retain crucial surface properties and express chlamydial antigen: an imaging study of a delivery system for the ocular surface

    Directory of Open Access Journals (Sweden)

    Montanaro J

    2015-07-01

    Full Text Available Jacqueline Montanaro,1 Aleksandra Inic-Kanada,1 Angela Ladurner,1 Elisabeth Stein,1 Sandra Belij,1 Nora Bintner,1 Simone Schlacher,1 Nadine Schuerer,1 Ulrike Beate Mayr,2 Werner Lubitz,2 Nikolaus Leisch,3 Talin Barisani-Asenbauer11Laura Bassi Centres of Expertise, OCUVAC – Centre of Ocular Inflammation and Infection, Centre for Pathophysiology, Infectiology, and Immunology, Medical University of Vienna, Vienna, Austria; 2BIRD-C GmbH & Co KG, Kritzendorf, Austria; 3Department of Ecogenomics and Systems Biology, University of Vienna, Vienna, AustriaAbstract: To target chronic inflammatory ocular surface diseases, a drug delivery platform is needed that is safe, possesses immunomodulatory properties, and can be used either for drug delivery, or as a foreign antigen carrier. A new therapeutic approach that we have previously proposed uses nonliving bacterial ghosts (BGs as a carrier-delivery system which can be engineered to carry foreign antigens and/or be loaded with therapeutic drugs. The parent strain chosen for development of our BG delivery system is the probiotic Escherichia coli strain Nissle 1917 (EcN, whose intrinsic properties trigger the innate immune system with the flagella and fimbriae used to attach and stimulate epithelial cells. In previous studies, we have shown that EcN BGs are safe for the ocular surface route, but evidence that EcN BGs retain flagella and fimbriae after transformation, has never been visually confirmed. In this study, we used different visualization techniques to determine whether flagella and fimbriae are retained on EcN BGs engineered either for drug delivery or as a foreign antigen carrier. We have also shown by immunoelectron microscopy that EcN retains two foreign antigens after processing to become EcN BGs. Furthermore, we demonstrated that BGs derived from EcN and expressing a foreign antigen attachment to conjunctival epithelial cells in vitro without causing reduced cell viability. These results

  18. Cell proliferation-inducing protein 52/mitofilin is a surface antigen on undifferentiated human dental pulp stem cells.

    Science.gov (United States)

    Hwang, Hyo-In; Lee, Tae-Hyong; Jang, Young-Joo

    2015-06-01

    Dental pulp is a soft tissue located inside the hard part of a tooth and it contains a stem cell population that can regenerate damaged dentin and/or pulp itself. Human dental pulp stem cells (hDPSCs) are multipotent adult stem cells that have the potential to be differentiated into a variety of cell types. Although cells cultured primarily from pulp tissue show heterogeneous phenotypes and variable efficiency in their dentinogenic differentiation, proper selection markers, which are specific to hDPSCs, are essential for the osteo/dentinogenic study of human dental pulp cells. We had previously screened a set of undifferentiation-specific cell surface antibodies of hDPSCs through decoy immunization. In this study, we show that one of these surface monoclonal antibodies, 3C4, is bound to intact pulp cells in a highly undifferentiation-specific manner. The surface antigen protein bound specifically to 3C4 antibody was identified through direct immunoprecipitation and liquid chromatography-tandem mass spectrometry as the cell proliferation-inducing protein 52/Mitofilin, which is a protein of the inner mitochondrial membrane and is a possible antagonist to maintaining mitochondrial activation during differentiation. The expression of mitofilin/3C4 antigen dramatically decreased during differentiation, and the depletion of mitofilin/3C4 antigen induced the expression of osteogenic/dentinogenic markers earlier than during normal differentiation. The 3C4-positive cells isolated by a magnetic-activated cell sorting system were differentiated with a higher efficiency than 3C4-negative cells. These results indicate that finding mitochondria-related stem cell markers is valuable to be able to identify and isolate primitive stem cells.

  19. Chimeric foot-and-mouth disease virus serotype O displaying a serotype Asia1 antigenic epitope at the surface.

    Science.gov (United States)

    Biswal, Jitendra K; Ranjan, Rajeev; Pattnaik, Bramhadev

    2016-09-01

    To determine whether the G-H loop of foot-and-mouth disease virus (FMDV) serotype O can function as a target structure to harbour and display serotype Asia1 antigenic epitope at the surface. Using reverse genetics, FMDV serotype O IND R2/1975 displaying a FMDV serotype Asia1 B cell epitope at the capsid surface was constructed. The epitope-inserted recombinant chimeric virus was genetically stable up to ten serial passages in cell culture and exhibited growth properties similar to the parental serotype O virus. Furthermore, the surface-displayed Asia1 epitope able to react with serotype Asia1 specific antibodies in a competitive ELISA. Importantly, the recombinant chimeric virus showed neutralizing activity to both serotype O and Asia1 polyclonal antibodies. The capsid protein of FMDV serotype O can effectively display potent epitope of other serotypes, making this an attractive approach for the design of new generation bi-valent FMD vaccines.

  20. Development of Lateral Flow Assay Based on Size-Controlled Gold Nanoparticles for Detection of Hepatitis B Surface Antigen

    OpenAIRE

    Kim, Dong Seok; Kim, Yong Tae; Hong, Seok Bok; Kim, Jinwoon; Heo, Nam Su; Lee, Moon-Keun; Lee, Seok Jae; Kim, Byeong Il; Kim, In Soo; Huh, Yun Suk; Choi, Bong Gill

    2016-01-01

    In this study, we developed lateral flow assay (LFA) biosensors for the detection of hepatitis B surface antigens using well-controlled gold nanoparticles (AuNPs). To enhance colorimetric signals, a seeded growth method was used for the preparation of size-controlled AuNPs with a narrow size distribution. Different sizes of AuNPs in the range of 342–137.8 nm were conjugated with antibodies and then optimized for the efficient detection of LFA biosensors. The conjugation stability was investig...

  1. Evaluation of the fully automated Cobas Core enzyme immunoassay for the quantitation of antibodies against hepatitis B virus surface antigen.

    Science.gov (United States)

    Doche, C; Thomé, M; Dimet, I; Bienvenu, J

    1996-04-01

    The Roche Cobas Core automated enzyme immunoassay of antibodies against hepatitis B virus surface antigen (anti-HBs) was evaluated using a protocol complying with the Société Française de Biologie Clinique guidelines. Results showed good precision (CV below 6 and 9% for intra- and inter-assay precision) and accuracy (according to the Paul-Ehrlich-Institute standard); the detection limit of the test was 2 IU/l. The manufacturer's specifications were confirmed and a satisfactory correlation (r = 0.901) was found with an automated method (IMx, Abbott) used for comparison.

  2. Differential induction of immunoglobulin G to Plasmodium falciparum variant surface antigens during the transmission season in Daraweesh, Sudan

    DEFF Research Database (Denmark)

    Nielsen, Morten A; Grevstad, Berit; A-Elgadir, Thoraya M E

    2005-01-01

    with severe malaria (VSA(SM)) are frequently recognized by IgG. METHODS: We analyzed levels of anti-VSA IgG in 57 individuals in Daraweesh, Sudan, before and after the transmission season. IgG responses to 79 Plasmodium falciparum isolates from children with defined malaria syndromes and exposed to high...

  3. Effects of pregnancy and intensity of Plasmodium falciparum transmission on immunoglobulin G subclass responses to variant surface antigens

    DEFF Research Database (Denmark)

    Megnekou, Rosette; Staalsoe, Trine; Taylor, Diane W

    2005-01-01

    , the levels of VSA(PAM)-specific antibodies either were very low or negative or were very high, whereas the levels of the antibodies specific for non-PAM type VSA were uniformly high. Interestingly, the levels of VSA(PAM)-specific IgG1 increased with increasing gestational age, while the levels...... status, parity, gestational age, and parasite transmission intensity, whereas only the parasite transmission intensity influenced the levels of IgG specific for non-PAM type VSA. For both types of VSA, the responses were dominated by the cytophilic subclass IgG1, followed by IgG3. In pregnant women...

  4. Correlation Between Hepatitis B Surface Antigen Titers and HBV DNA Levels

    OpenAIRE

    Amal Alghamdi; Nagwa Aref; Malak El-Hazmi; Waleed Al-Hamoudi; Khalid Alswat; Ahmed Helmy; Sanai, Faisal M; Abdo, Ayman A.

    2013-01-01

    Background/Aim: To assess the correlation between serum HBsAg titers and hepatitis B virus (HBV) DNA levels in patients with hepatitis B envelop antigen-negative (HBeAg −ve) HBV genotype-D (HBV/D) infection. Patients and Methods: A total of 106 treatment- naïve, HBeAg −ve HBV/D patients were included; 78 in the inactive carrier (IC) state and 28 in the active hepatitis (AH) stage. HBV DNA load and HBsAg titers were tested using TaqMan real-time polymerase chain reaction (PCR) and automated ch...

  5. 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.

  6. Biological, immunological and functional properties of two novel multi-variant chimeric recombinant proteins of CSP antigens for vaccine development against Plasmodium vivax infection.

    Science.gov (United States)

    Shabani, Samaneh H; Zakeri, Sedigheh; Salmanian, Ali H; Amani, Jafar; Mehrizi, Akram A; Snounou, Georges; Nosten, François; Andolina, Chiara; Mourtazavi, Yousef; Djadid, Navid D

    2017-10-01

    The circumsporozoite protein (CSP) of the malaria parasite Plasmodium vivax is a major pre-erythrocyte vaccine candidate. The protein has a central repeat region that belongs to one of repeat families (VK210, VK247, and the P. vivax-like). In the present study, computer modelling was employed to select chimeric proteins, comprising the conserved regions and different arrangements of the repeat elements (VK210 and VK247), whose structure is similar to that of the native counterparts. DNA encoding the selected chimeras (named CS127 and CS712) were synthetically constructed based on E. coli codons, then cloned and expressed. Mouse monoclonal antibodies (mAbs; anti-Pv-210-CDC and -Pv-247-CDC), recognized the chimeric antigens in ELISA, indicating correct conformation and accessibility of the B-cell epitopes. ELISA using IgG from plasma samples collected from 221 Iranian patients with acute P. vivax showed that only 49.32% of the samples reacted to both CS127 and CS712 proteins. The dominant subclass for the two chimeras was IgG1 (48% of the positive responders, OD492=0.777±0.420 for CS127; 48.41% of the positive responders, OD492=0.862±0.423 for CS712, with no statistically significant difference P>0.05; Wilcoxon signed ranks test). Binding assays showed that both chimeric proteins bound to immobilized heparan sulphate and HepG2 hepatocyte cells in a concentration-dependent manner, saturable at 80μg/mL. Additionally, anti-CS127 and -CS712 antibodies raised in mice recognized the native protein on the surface of P. vivax sporozoite with high intensity, confirming the presence of common epitopes between the recombinant forms and the native proteins. In summary, despite structural differences at the molecular level, the expression levels of both chimeras were satisfactory, and their conformational structure retained biological function, thus supporting their potential for use in the development of vivax-based vaccine. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Surface antigens and potential virulence factors from parasites detected by comparative genomics of perfect amino acid repeats

    Directory of Open Access Journals (Sweden)

    Adler Joël

    2007-12-01

    Full Text Available Abstract Background Many parasitic organisms, eukaryotes as well as bacteria, possess surface antigens with amino acid repeats. Making up the interface between host and pathogen such repetitive proteins may be virulence factors involved in immune evasion or cytoadherence. They find immunological applications in serodiagnostics and vaccine development. Here we use proteins which contain perfect repeats as a basis for comparative genomics between parasitic and free-living organisms. Results We have developed Reptile http://reptile.unibe.ch, a program for proteome-wide probabilistic description of perfect repeats in proteins. Parasite proteomes exhibited a large variance regarding the proportion of repeat-containing proteins. Interestingly, there was a good correlation between the percentage of highly repetitive proteins and mean protein length in parasite proteomes, but not at all in the proteomes of free-living eukaryotes. Reptile combined with programs for the prediction of transmembrane domains and GPI-anchoring resulted in an effective tool for in silico identification of potential surface antigens and virulence factors from parasites. Conclusion Systemic surveys for perfect amino acid repeats allowed basic comparisons between free-living and parasitic organisms that were directly applicable to predict proteins of serological and parasitological importance. An on-line tool is available at http://genomics.unibe.ch/dora.

  8. Specific Detection of Carcinoembryonic Antigen Based on Fluorescence Quenching of Hollow Porous Gold Nanoshells with Roughened Surface.

    Science.gov (United States)

    Xing, Ting-Yang; Zhao, Jing; Weng, Guo-Jun; Zhu, Jian; Li, Jian-Jun; Zhao, Jun-Wu

    2017-10-25

    The detection of tumor biomarkers in the early stage is highly desirable for the therapy of cancer. However, rapid, low-cost, sensitive, and selective detection of biomarkers remains a challenge owing to the sequence homology, short length, and low abundance. This Research Article describes the synthesis of a novel carcinoembryonic antigen (CEA) probe using hollow porous gold nanoparticles (HPGNPs) with roughened surface based on fluorescence quenching. For specific detection of CEA, the surface of HPGNP is modified by carboxyl modification, carboxyl activation, and antibody conjugation. Furthermore, to enhance the detection performance, we have systematically optimized the parameters, such as particle size, surfactants, surface roughness, surface hole size, and the molecule-particle distance (MPD). The results demonstrate that the fluorescence quenching efficiency would be enhanced with a larger particle size and surface hole size, roughened surface and a greater MPD. Also, with careful inspection of different surfactants of CTAB and PVP, we find that PVP has the optimal performance on fluorescence quenching. Under these optimized conditions, CEA could be detected with an ultralow detection limit of 1.5 pg/mL, and the probe shows a linear range from 2 to 100 pg/mL. The limit of detection is an order of intensity lower than related methods. Interference experiment results have shown that the influence of the interfering proteins could be neglected in the detection procedure.

  9. A new technique to detect antibody-antigen reaction (biological interactions) on a localized surface plasmon resonance (LSPR) based nano ripple gold chip

    Science.gov (United States)

    Saleem, Iram; Widger, William; Chu, Wei-Kan

    2017-07-01

    We demonstrate that the gold nano-ripple localized surface plasmon resonance (LSPR) chip is a low cost and a label-free method for detecting the presence of an antigen. A uniform stable layer of an antibody was coated on the surface of a nano-ripple gold pattern chip followed by the addition of different concentrations of the antigen. A red shift was observed in the LSPR spectral peak caused by the change in the local refractive index in the vicinity of the nanostructure. The LSPR chip was fabricated using oblique gas cluster ion beam (GCIB) irradiation. The plasmon-resonance intensity of the scattered light was measured by a simple optical spectroscope. The gold nano ripple chip shows monolayer scale sensitivity and high selectivity. The LSPR substrate was used to detect antibody-antigen reaction of rabbit X-DENTT antibody and DENTT blocking peptide (antigen).

  10. A DNA vaccine encoding a cell-surface protein antigen of Streptococcus mutans protects gnotobiotic rats from caries.

    Science.gov (United States)

    Fan, M W; Bian, Z; Peng, Z X; Zhong, Y; Chen, Z; Peng, B; Jia, R

    2002-11-01

    A cell-surface protein antigen (PAc) of Streptococcus mutans is considered a virulence factor because it may mediate initial attachment of Streptococcus mutans to tooth surfaces. Thus, inhibiting PAc is predicted to provide protection against caries. To develop vaccines against dental caries, we constructed a DNA vaccine, pCIA-P, which encodes two high-conservative regions of PAc. Expression of the recombinant protein was obtained in eukaryotic cells in vitro and in vivo. In this report, we provide evidence that fewer caries lesions, and high levels of PAc-specific salivary IgA antibody and serum IgG antibody, were observed in gnotobiotic rats following targeted salivary gland (TSG) administration of pCIA-P. This study shows that the recombinant DNA vaccine pCIA-P could induce protective anti-caries immune responses and that TSG immunization is a promising strategy for the inhibition of dental caries.

  11. The O-Antigen Capsule of Salmonella enterica Serovar Typhimurium Facilitates Serum Resistance and Surface Expression of FliC.

    Science.gov (United States)

    Marshall, Joanna M; Gunn, John S

    2015-10-01

    Group IV polysaccharide capsules are common in enteric bacteria and have more recently been described in nontyphoidal Salmonella species. Such capsules are known as O-antigen (O-Ag) capsules, due to their high degree of similarity to the O-Ag of the lipopolysaccharide (LPSO-Ag). Capsular polysaccharides are known virulence factors of many bacterial pathogens, facilitating evasion of immune recognition and systemic dissemination within the host. Previous studies on the O-Ag capsule of salmonellae have focused primarily on its role in bacterial surface attachment and chronic infection; however, the potential effects of the O-Ag capsule on acute pathogenesis have yet to be investigated. While much of the in vivo innate immune resistance of Salmonella enterica serovar Typhimurium is attributed to the high-molecular-weight LPS, we hypothesized that the O-Ag capsule may enhance this resistance by diminishing surface expression of pathogen-associated molecular patterns, such as flagella, and increasing resistance to host immune molecules. To test this hypothesis, O-Ag capsule-deficient mutants were constructed, and the loss of O-Ag capsular surface expression was confirmed through microscopy and immunoblotting. Loss of O-Ag capsule production did not alter bacterial growth or production of LPS. Western blot analysis and confocal microscopy revealed that O-Ag capsule-deficient mutants demonstrate reduced resistance to killing by human serum. Furthermore, O-Ag capsule-deficient mutants produced exclusively phase I flagellin (FliC). Although O-Ag capsule-deficient mutants did not exhibit reduced virulence in a murine model of acute infection, in vitro results indicate that the O-Ag capsule may function to modify the antigenic nature of the bacterial surface, warranting additional investigation of a potential role of the structure in pathogenesis. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  12. Clinical significance of surface antigen expression in children with acute myeloid leukemia: results of study AML-BFM-87.

    Science.gov (United States)

    Creutzig, U; Harbott, J; Sperling, C; Ritter, J; Zimmermann, M; Löffler, H; Riehm, H; Schellong, G; Ludwig, W D

    1995-10-15

    Immunophenotyping using a panel of 15 antibodies was performed in 267 (87%) and cytogenetic analysis in 196 (64%) of 307 children under 17 years of age enrolled in the AML-BFM-87 study. Treatment consisted of cytosine arabinoside, daunorubicin, etoposide induction and a 6-week seven-drug consolidation chemotherapy, followed by two blocks of high-dose cytosine arabinoside with or without cranial irradiation and maintenance therapy for 1 year. Five-year event-free survival for patients with immunophenotypic data was .43 +/- .03 SE. The diagnostic value of the pan-myeloid reagents CD13, CD33, and CDw65 for the recognition of childhood acute myeloid leukemia (AML) was high with a sensitivity of 98% (positivity of at least one of these antigens), whereas, with the exception of CD41 for French American British (FAB) subtype M7, the expression of single cell-surface antigens showed no correlation with morphologic or cytogenetic subgroups. On the other hand, characteristic subgroups of AML defined by morphologic features and karyotypes could be described by low or high rates of surface antigen expression compared with those of other patients. These immunophenotypic features most probably associated with specific entities include expression of CD34 or CD13 and absence of CD14 or CD4 in M2 with Auer rods/t(8;21); absence of HLA-DR, CD34, and CD14, but expression of CD33 in M3/t(15;17); positivity of either CD34 or CD13 and either CD14 or CD2 for M4Eo/inv(16); and absence of either CD34 or CD13 and expression of either CD33 or CDw65 and either CD15 or CD4 for M5/t(9;11). In FAB M0, negativity of one or two of the three panmyeloid-associated markers (CD13/33/w65) was common; and cytogenetic results frequently showed random abnormalities. Expression of lymphoid-, progenitor- and most myeloid-associated antigens had no influence on the prognosis, whereas the outcome was significantly better for children with M2 with Auer rods, M3, or M4Eo or for those with the associated

  13. Major role for carbohydrate epitopes preferentially recognized by chronically infected mice in the determination of Schistosoma mansoni schistosomulum surface antigenicity

    Energy Technology Data Exchange (ETDEWEB)

    Omer-ali, P.; Magee, A.I.; Kelly, C.; Simpson, A.J.G.

    1986-12-01

    A radioimmunoassay that makes use of whole Schistosomula and /sup 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.

  14. Multiple Hepatitis B Virus (HBV) Quasispecies and Immune-Escape Mutations Are Present in HBV Surface Antigen and Reverse Transcriptase of Patients With Acute Hepatitis B.

    Science.gov (United States)

    Aragri, Marianna; Alteri, Claudia; Battisti, Arianna; Di Carlo, Domenico; Minichini, Carmine; Sagnelli, Caterina; Bellocchi, Maria Concetta; Pisaturo, Maria Antonietta; Starace, Mario; Armenia, Daniele; Carioti, Luca; Pollicita, Michela; Salpini, Romina; Sagnelli, Evangelista; Perno, Carlo Federico; Coppola, Nicola; Svicher, Valentina

    2016-06-15

    This study characterizes and defines the clinical value of hepatitis B virus (HBV) quasispecies with reverse transcriptase and HBV surface antigen (HBsAg) heterogeneity in patients with acute HBV infection. Sixty-two patients with acute HBV infection (44 with genotype D infection and 18 with genotype A infection) were enrolled from 2000 to 2010. Plasma samples obtained at the time of the first examination were analyzed by ultradeep pyrosequencing. The extent of HBsAg amino acid variability was measured by Shannon entropy. Median alanine aminotransferase and serum HBV DNA levels were 2544 U/L (interquartile range, 1938-3078 U/L) and 5.88 log10 IU/mL (interquartile range, 4.47-7.37 log10 IU/mL), respectively. Although most patients serologically resolved acute HBV infection, only 54.1% developed antibody to HBsAg (anti-HBs). A viral population with ≥1 immune-escape mutation was found in 53.2% of patients (intrapatient prevalence range, 0.16%-100%). Notably, by Shannon entropy, higher genetic variability at HBsAg amino acid positions 130, 133, and 157 significantly correlated with no production of anti-HBs in individuals infected with genotype D (P < .05). Stop codons were detected in 19.3% of patients (intrapatient prevalence range, 1.6%-47.5%) and occurred at 11 HBsAg amino acid positions, including 172 and 182, which are known to increase the oncogenic potential of HBV.Finally, ≥1 drug resistance mutation was detected in 8.1% of patients (intrapatient prevalence range, 0.11%-47.5% for primary mutations and 10.5%-99.9% for compensatory mutations). Acute HBV infection is characterized by complex array of viral quasispecies with reduced antigenicity/immunogenicity and enhanced oncogenic potential. These viral variants may induce difficult-to-treat HBV forms; favor HBV reactivation upon iatrogenic immunosuppression, even years after infection; and potentially affect the efficacy of the current HBV vaccination strategy. © The Author 2016. Published by Oxford

  15. Recombinant Forms of Leishmania amazonensis Excreted/Secreted Promastigote Surface Antigen (PSA Induce Protective Immune Responses in Dogs.

    Directory of Open Access Journals (Sweden)

    Elodie Petitdidier

    2016-05-01

    Full Text Available Preventive vaccination is a highly promising strategy for interrupting leishmaniasis transmission that can, additionally, contribute to elimination. A vaccine formulation based on naturally excreted secreted (ES antigens was prepared from L. infantum promastigote culture supernatant. This vaccine achieved successful results in Phase III trials and was licensed and marketed as CaniLeish. We recently showed that newly identified ES promastigote surface antigen (PSA, from both viable promastigotes and axenically-grown amastigotes, represented the major constituent and the highly immunogenic antigen of L. infantum and L. amazonensis ES products. We report here that three immunizations with either the recombinant ES LaPSA-38S (rPSA or its carboxy terminal part LaPSA-12S (Cter-rPSA, combined with QA-21 as adjuvant, confer high levels of protection in naive L. infantum-infected Beagle dogs, as checked by bone marrow parasite absence in respectively 78.8% and 80% of vaccinated dogs at 6 months post-challenge. The parasite burden in infected vaccinated dogs was significantly reduced compared to placebo group, as measured by q-PCR. Moreover, our results reveal humoral and cellular immune response clear-cut differences between vaccinated and control dogs. An early increase in specific IgG2 antibodies was observed in rPSA/QA-21- and Cter-rPSA/QA-21-immunized dogs only. They were found functionally active in vitro and were highly correlated with vaccine protection. In vaccinated protected dogs, IFN-γ and NO productions, as well as anti-leishmanial macrophage activity, were increased. These data strongly suggest that ES PSA or its carboxy-terminal part, in recombinant forms, induce protection in a canine model of zoonotic visceral leishmaniasis by inducing a Th1-dominant immune response and an appropriate specific antibody response. These data suggest that they could be considered as important active components in vaccine candidates.

  16. Recombinant Forms of Leishmania amazonensis Excreted/Secreted Promastigote Surface Antigen (PSA) Induce Protective Immune Responses in Dogs.

    Science.gov (United States)

    Petitdidier, Elodie; Pagniez, Julie; Papierok, Gérard; Vincendeau, Philippe; Lemesre, Jean-Loup; Bras-Gonçalves, Rachel

    2016-05-01

    Preventive vaccination is a highly promising strategy for interrupting leishmaniasis transmission that can, additionally, contribute to elimination. A vaccine formulation based on naturally excreted secreted (ES) antigens was prepared from L. infantum promastigote culture supernatant. This vaccine achieved successful results in Phase III trials and was licensed and marketed as CaniLeish. We recently showed that newly identified ES promastigote surface antigen (PSA), from both viable promastigotes and axenically-grown amastigotes, represented the major constituent and the highly immunogenic antigen of L. infantum and L. amazonensis ES products. We report here that three immunizations with either the recombinant ES LaPSA-38S (rPSA) or its carboxy terminal part LaPSA-12S (Cter-rPSA), combined with QA-21 as adjuvant, confer high levels of protection in naive L. infantum-infected Beagle dogs, as checked by bone marrow parasite absence in respectively 78.8% and 80% of vaccinated dogs at 6 months post-challenge. The parasite burden in infected vaccinated dogs was significantly reduced compared to placebo group, as measured by q-PCR. Moreover, our results reveal humoral and cellular immune response clear-cut differences between vaccinated and control dogs. An early increase in specific IgG2 antibodies was observed in rPSA/QA-21- and Cter-rPSA/QA-21-immunized dogs only. They were found functionally active in vitro and were highly correlated with vaccine protection. In vaccinated protected dogs, IFN-γ and NO productions, as well as anti-leishmanial macrophage activity, were increased. These data strongly suggest that ES PSA or its carboxy-terminal part, in recombinant forms, induce protection in a canine model of zoonotic visceral leishmaniasis by inducing a Th1-dominant immune response and an appropriate specific antibody response. These data suggest that they could be considered as important active components in vaccine candidates.

  17. An improved Abbott ARCHITECT assay for the detection of hepatitis B virus surface antigen (HBsAg).

    Science.gov (United States)

    Lou, Sheng C; Pearce, Sandra K; Lukaszewska, Teresa X; Taylor, Russell E; Williams, Gregg T; Leary, Thomas P

    2011-05-01

    The sensitive and accurate detection of hepatitis B virus surface antigen (HBsAg) is critical to the identification of infection and the prevention of transfusion transmitted disease. Improvement in HBsAg assay sensitivity is essential to reduce the window to detect an acute HBV infection. Additionally, the sensitive detection of HBsAg mutants that continue to evolve due to vaccine escape, immune selection and an error prone reverse transcriptase is a necessity. A fully automated HBsAg prototype assay on the Abbott ARCHITECT instrument was developed to improve sensitivity and mutant detection. This magnetic microparticle-based assay utilizes anti-HBsAg monoclonal antibodies to capture antigen present in serum or plasma. Captured antigen is then detected using anti-HBsAg antibody conjugated with the chemiluminescent compound, acridinium. The sensitivity of the ARCHITECT HBsAg prototype assay was improved as compared to the current ARCHITECT, PRISM, and competitor HBsAg assays. The enhancement in assay sensitivity was demonstrated by the use of commercially available HBV seroconversion panels. The prototype assay detected more panel members (185 of 383) vs. the current ARCHITECT (171), PRISM (181), or competitor HBsAg assays (73/140 vs. 62/140, respectively). The ARCHITECT prototype assay also efficiently detected all mutants evaluated. Finally, the sensitivity improvement did not compromise the specificity of the assay (99.94%). An improved Abbott ARCHITECT HBsAg prototype assay with enhanced detection of HBsAg and HBsAg mutants, as well as equivalent specificity was developed for the detection, diagnosis, and management of HBV infection. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. Antigen-presenting particle technology using inactivated surface-engineered viruses: induction of immune responses against infectious agents

    Directory of Open Access Journals (Sweden)

    Chang Yung-Nien

    2007-05-01

    particles can be surface-engineered to produce antigen-presenting particles that mimic antigen-presenting cells to induce immune responses in human peripheral blood lymphocytes. The viral particles behave as "biological carriers" for recombinant proteins, thereby establishing a new therapeutic paradigm for molecular medicine.

  19. MetQ of Neisseria gonorrhoeae Is a Surface-Expressed Antigen That Elicits Bactericidal and Functional Blocking Antibodies.

    Science.gov (United States)

    Semchenko, Evgeny A; Day, Christopher J; Seib, Kate L

    2017-02-01

    Neisseria gonorrhoeae, the causative agent of the sexually transmitted infection (STI) gonorrhea, is a growing public health threat for which a vaccine is urgently needed. We characterized the functional role of the gonococcal MetQ protein, which is the methionine binding component of an ABC transporter system, and assessed its potential as a candidate antigen for inclusion in a gonococcal vaccine. MetQ has been found to be highly conserved in all strains investigated to date, it is localized on the bacterial surface, and it binds l-methionine with a high affinity. MetQ is also involved in gonococcal adherence to cervical epithelial cells. Mutants lacking MetQ have impaired survival in human monocytes, macrophages, and serum. Furthermore, antibodies raised against MetQ are bactericidal and are able to block gonococcal adherence to epithelial cells. These data suggest that MetQ elicits both bactericidal and functional blocking antibodies and is a valid candidate antigen for additional investigation and possible inclusion in a vaccine for prevention of gonorrhea. Copyright © 2017 Semchenko et al.

  20. A self-amplified transistor immunosensor under dual gate operation: highly sensitive detection of hepatitis B surface antigen

    Science.gov (United States)

    Lee, I.-K.; Jeun, M.; Jang, H.-J.; Cho, W.-J.; Lee, K. H.

    2015-10-01

    Ion-sensitive field-effect transistors (ISFETs), although they have attracted considerable attention as effective immunosensors, have still not been adopted for practical applications owing to several problems: (1) the poor sensitivity caused by the short Debye screening length in media with high ion concentration, (2) time-consuming preconditioning processes for achieving the highly-diluted media, and (3) the low durability caused by undesirable ions such as sodium chloride in the media. Here, we propose a highly sensitive immunosensor based on a self-amplified transistor under dual gate operation (immuno-DG ISFET) for the detection of hepatitis B surface antigen. To address the challenges in current ISFET-based immunosensors, we have enhanced the sensitivity of an immunosensor by precisely tailoring the nanostructure of the transistor. In the pH sensing test, the immuno-DG ISFET showed superior sensitivity (2085.53 mV per pH) to both standard ISFET under single gate operation (58.88 mV per pH) and DG ISFET with a non-tailored transistor (381.14 mV per pH). Moreover, concerning the detection of hepatitis B surface antigens (HBsAg) using the immuno-DG ISFET, we have successfully detected trace amounts of HBsAg (22.5 fg mL-1) in a non-diluted 1× PBS medium with a high sensitivity of 690 mV. Our results demonstrate that the proposed immuno-DG ISFET can be a biosensor platform for practical use in the diagnosis of various diseases.Ion-sensitive field-effect transistors (ISFETs), although they have attracted considerable attention as effective immunosensors, have still not been adopted for practical applications owing to several problems: (1) the poor sensitivity caused by the short Debye screening length in media with high ion concentration, (2) time-consuming preconditioning processes for achieving the highly-diluted media, and (3) the low durability caused by undesirable ions such as sodium chloride in the media. Here, we propose a highly sensitive immunosensor

  1. Aeromonas surface glucan attached through the O-antigen ligase represents a new way to obtain UDP-glucose.

    Directory of Open Access Journals (Sweden)

    Susana Merino

    Full Text Available We previously reported that A. hydrophila GalU mutants were still able to produce UDP-glucose introduced as a glucose residue in their lipopolysaccharide core. In this study, we found the unique origin of this UDP-glucose from a branched α-glucan surface polysaccharide. This glucan, surface attached through the O-antigen ligase (WaaL, is common to the mesophilic Aeromonas strains tested. The Aeromonas glucan is produced by the action of the glycogen synthase (GlgA and the UDP-Glc pyrophosphorylase (GlgC, the latter wrongly indicated as an ADP-Glc pyrophosphorylase in the Aeromonas genomes available. The Aeromonas glycogen synthase is able to react with UDP or ADP-glucose, which is not the case of E. coli glycogen synthase only reacting with ADP-glucose. The Aeromonas surface glucan has a role enhancing biofilm formation. Finally, for the first time to our knowledge, a clear preference on behalf of bacterial survival and pathogenesis is observed when choosing to produce one or other surface saccharide molecules to produce (lipopolysaccharide core or glucan.

  2. Localization of distinct surface antigens on Chlamydia trachomatis HAR-13 by immune electron microscopy with monoclonal antibodies.

    Science.gov (United States)

    Clark, R B; Nachamkin, I; Schatzki, P F; Dalton, H P

    1982-01-01

    Monoclonal antibodies were produced against the HAR-13 strain (A) of Chlamydia trachomatis. By an indirect immunofluorescence technique, three clones (1-7, 2-8, 9F) were found to produce antibody that reacted only with serotype A strains. Clone 3-5 produced antibody that cross-reacted with serotypes A, C, H, I, and J. Using an indirect immunoferritin technique, we examined the binding of these antibodies to strain HAR-13 by electron microscopy. Antibodies from all four clones were shown to bind to the outer membrane surface of both reticulate and elementary bodies. Two distinct binding patterns were demonstrated. Antibodies from clones 1-7 and 9F bound to the outer membrane surface in a homogeneous pattern, whereas antibodies 2-8 and 3-5 bound to the outer membrane surface in an irregular, patchy distribution. There was a direct correlation between the distribution of antigen on the outer membrane surface and neutralization of C. trachomatis HAR-13 infectivity in vitro. Neutralization of HAR-13 infectivity by antibodies 1-7 and 9F was complement dependent, whereas antibodies 2-8 and 3-5 did not neutralize under any conditions. Images PMID:7152670

  3. Immunogenicity and protective immunity against bubonic plague and pneumonic plague by immunization of mice with the recombinant V10 antigen, a variant of LcrV.

    Science.gov (United States)

    DeBord, Kristin L; Anderson, Deborah M; Marketon, Melanie M; Overheim, Katie A; DePaolo, R William; Ciletti, Nancy A; Jabri, Bana; Schneewind, Olaf

    2006-08-01

    In contrast to Yersinia pestis LcrV, the recombinant V10 (rV10) variant (lacking residues 271 to 300) does not suppress the release of proinflammatory cytokines by immune cells. Immunization with rV10 generates robust antibody responses that protect mice against bubonic plague and pneumonic plague, suggesting that rV10 may serve as an improved plague vaccine.

  4. Comparative evaluation of hepatitis B surface antigen-loaded elastic liposomes and ethosomes for human dendritic cell uptake and immune response.

    Science.gov (United States)

    Mishra, Dinesh; Mishra, Pradyumna Kumar; Dabadghao, Sunil; Dubey, Vaibhav; Nahar, Manoj; Jain, Narendra K

    2010-02-01

    The aim of the present study was to evaluate two vesicular carrier systems, ethosomes and elastic liposomes loaded with hepatitis B surface antigen, for in vitro qualitative and quantitative uptake by human dendritic cells (DCs) and ability to stimulate T lymphocytes. Quantitative uptake of antigen-loaded carriers was documented by flow cytometry, and internalization of the systems by the DCs was studied using spectral bioimaging. Ability of antigen-pulsed DCs to stimulate autologous peripheral blood lymphocytes and levels of TH1/TH2 cytokines were also examined using flow cytometry. Both vesicular carrier systems as antigen delivery modules and DCs as antigen-presenting cells were able to generate a protective immune response. However, ethosomes were found to have higher internalizing ability and immunogenicity in comparison with elastic liposomes. These properties of ethosomes coupled with their skin-navigating potential, make it an attractive vehicle for development of a transcutaneous vaccine against hepatitis B in preference to elastic liposomes. Two carrier systems for more potent vaccine administration - ethosomes and elastic liposomes loaded with hepatitis B surface antigen - are compared. Ethosomes demonstrated higher internalizing ability and immunogenicity. Due to their known skin-navigating potential, ethosomes may represent an attractive vehicle for development of a transcutaneous vaccine against hepatitis B. Copyright 2010 Elsevier Inc. All rights reserved.

  5. Correlation between hepatitis B surface antigen titers and HBV DNA levels.

    Science.gov (United States)

    Alghamdi, Amal; Aref, Nagwa; El-Hazmi, Malak; Al-Hamoudi, Waleed; Alswat, Khalid; Helmy, Ahmed; Sanai, Faisal M; Abdo, Ayman A

    2013-01-01

    To assess the correlation between serum HBsAg titers and hepatitis B virus (HBV) DNA levels in patients with hepatitis B envelop antigen-negative (HBeAg -ve) HBV genotype-D (HBV/D) infection. A total of 106 treatment- naïve, HBeAg -ve HBV/D patients were included; 78 in the inactive carrier (IC) state and 28 in the active hepatitis (AH) stage. HBV DNA load and HBsAg titers were tested using TaqMan real-time polymerase chain reaction (PCR) and automated chemiluminescent microparticle immunoassay, respectively. The median (range) log10 HBsAg titer was significantly lower in the IC group compared with AH group, 3.09 (-1 to -4.4) versus 3.68 (-0.77 to 5.09) IU/mL, respectively; P correlation between HBsAg and HBV DNA levels in the whole cohort, AH, and IC groups (r = 0.6, P correlate with HBV DNA in treatment-naïve HBeAg -ve HBV/D patients, and supports the use of HBsAg levels in clinical practice as a predictor of serum HBV DNA levels.

  6. Development of Lateral Flow Assay Based on Size-Controlled Gold Nanoparticles for Detection of Hepatitis B Surface Antigen.

    Science.gov (United States)

    Kim, Dong Seok; Kim, Yong Tae; Hong, Seok Bok; Kim, Jinwoon; Huh, Nam Su; Lee, Moon-Keun; Lee, Seok Jae; Kim, Byeong Il; Kim, In Soo; Huh, Yun Suk; Choi, Bong Gill

    2016-12-16

    In this study, we developed lateral flow assay (LFA) biosensors for the detection of hepatitis B surface antigens using well-controlled gold nanoparticles (AuNPs). To enhance colorimetric signals, a seeded growth method was used for the preparation of size-controlled AuNPs with a narrow size distribution. Different sizes of AuNPs in the range of 342-137.8 nm were conjugated with antibodies and then optimized for the efficient detection of LFA biosensors. The conjugation stability was investigated by UV-vis spectroscopy of AuNP dispersion at various pH values and concentrations of antibody. Based on optimized conjugation conditions, the use of 42.7 ± 0.8 nm AuNPs exhibited superior performance for the detection of LFAs relative to other sizes of AuNPs.

  7. Development of Lateral Flow Assay Based on Size-Controlled Gold Nanoparticles for Detection of Hepatitis B Surface Antigen

    Directory of Open Access Journals (Sweden)

    Dong Seok Kim

    2016-12-01

    Full Text Available In this study, we developed lateral flow assay (LFA biosensors for the detection of hepatitis B surface antigens using well-controlled gold nanoparticles (AuNPs. To enhance colorimetric signals, a seeded growth method was used for the preparation of size-controlled AuNPs with a narrow size distribution. Different sizes of AuNPs in the range of 342–137.8 nm were conjugated with antibodies and then optimized for the efficient detection of LFA biosensors. The conjugation stability was investigated by UV-vis spectroscopy of AuNP dispersion at various pH values and concentrations of antibody. Based on optimized conjugation conditions, the use of 42.7 ± 0.8 nm AuNPs exhibited superior performance for the detection of LFAs relative to other sizes of AuNPs.

  8. Multiplex flow cytometry barcoding and antibody arrays identify surface antigen profiles of primary and metastatic colon cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Kumar Sukhdeo

    Full Text Available Colon cancer is a deadly disease affecting millions of people worldwide. Current treatment challenges include management of disease burden as well as improvements in detection and targeting of tumor cells. To identify disease state-specific surface antigen signatures, we combined fluorescent cell barcoding with high-throughput flow cytometric profiling of primary and metastatic colon cancer lines (SW480, SW620, and HCT116. Our multiplexed technique offers improvements over conventional methods by permitting the simultaneous and rapid screening of cancer cells with reduced effort and cost. The method uses a protein-level analysis with commercially available antibodies on live cells with intact epitopes to detect potential tumor-specific targets that can be further investigated for their clinical utility. Multiplexed antibody arrays can easily be applied to other tumor types or pathologies for discovery-based approaches to target identification.

  9. Impaired cell surface expression of HLA-B antigens on mesenchymal stem cells and muscle cell progenitors

    DEFF Research Database (Denmark)

    Isa, Adiba; Nehlin, Jan; Sabir, Hardee Jawad

    2010-01-01

    HLA class-I expression is weak in embryonic stem cells but increases rapidly during lineage progression. It is unknown whether all three classical HLA class-I antigens follow the same developmental program. In the present study, we investigated allele-specific expression of HLA-A, -B, and -C...... at the mRNA and protein levels on human mesenchymal stem cells from bone marrow and adipose tissue as well as striated muscle satellite cells and lymphocytes. Using multicolour flow cytometry, we found high cell surface expression of HLA-A on all stem cells and PBMC examined. Surprisingly, HLA-B was either...... undetectable or very weakly expressed on all stem cells protecting them from complement-dependent cytotoxicity (CDC) using relevant human anti-B and anti-Cw sera. IFNgamma stimulation for 48-72 h was required to induce full HLA-B protein expression. Quantitative real-time RT-PCR showed that IFNgamma induced...

  10. Mentally retarded hepatitis-B surface antigen carriers in NYC public school classes: a public health dilemma.

    Science.gov (United States)

    Bakal, C W; Novick, L F; Marr, J S; Millner, E S; Goldman, W D; Pitkin, O

    1980-07-01

    The placement of retarded children who have been institutionalized and are asymptomatic hepatitis-B surface antigen carriers into public school classes for the retarded has caused controversy and presented the New York City Health Department with an unusual medical-ethical dilemma. In this situation, the cost of interfering with deinstitutionalization, an important social advance, must be balanced against the benefit of controlling the unquantified but real risk of transmitting a potentially serious disease. The Health Department guidelines for managing this problem recommended serological surveillance, promotion of classroom hygiene where possible, and teaching of carriers in classes separate from their susceptible peers. A federal court disallowed the cohorting provisions of these guidelines. Changing policies and practices towards the mentally retarded, such as deinstitutionalization, raise important public health issues which will have to be faced by the involved communities.

  11. Evaluation of hepatitis B surface antigen and hepatitis B virus-DNA results in postmortem plasma specimens

    Directory of Open Access Journals (Sweden)

    Nihan Ziyade

    2015-03-01

    Full Text Available Objective: To assess the presence of hepatitis B surface antigen, one of the serologic markers of hepatitis B virus (HBV infection, in postmortem blood samples from autopsy cases using ELISA, and to compare the results with those obtained by PCR, which is the gold standard method in assessing HBV infection. Methods: The HBV test results of the blood samples from 880 autopsy cases determined in our laboratory, were retrospectively studied. Results: When compared with the gold standard method PCR, the sensitivity and specificity of postmortem ELISA were 100% and 84.1%, respectively. Conclusions: The increasingly used molecular diagnostic methods, such as PCR, should be used in cases where serological tests remain insufficient.We think that prospective studies on the comparison of ELISA and PCR assessment of postmortem blood samples with larger material should be carried out.

  12. Genotypic Characterization of Egypt Enterotoxigenic Escherichia coli Isolates Expressing Coli Surface Antigen 6

    Science.gov (United States)

    2013-02-01

    USA Abstract Introduction: One approach to control enterotoxigenic Escherichia coli (ETEC) infections has been to develop vaccines focused on...results show a lack of clonality among Egypt CS6 E. coli isolates and supports the use and the further research on vaccines targeting this cell surface...organisms must colonize the mucosal epithelium; this process utilizes fimbrial and non-fimbrial colonization factors, also referred to as coli surface

  13. Avaliação da vacina anti-rábica ERA, frente a variantes antigênicas do vírus da raiva, em diferentes períodos pós-imunização Evaluation of the ERA antirabies vaccine against antigenic variants of rabies virus in different post-immunization periods

    Directory of Open Access Journals (Sweden)

    Cláudia C. Cordeiro

    1990-12-01

    Full Text Available Avaliou-se, em camudongos, os níveis de proteção conferidos por uma vacina anti-rábica atenuada, preparada em cultura de tecido renal a partir da amostra ERA, frente a diferentes variantes antigênicas dos vírus da raiva. Utilizaram-se dois esquemas de vacinação, representados por uma única dose e por 6 doses aplicadas em dias alternados. Os diferentes camundogos dos grupos experimentais foram desafiados, em diversos períodos pós-imunização (15, 30, 60 e 120 dias, com variantes procedentes de cães (São Paulo e Nigéria, de morcego (DR-19 e Pernambuco, de raposa (Rio Grande do Norte e CVS "Challenge Virus Standard". Os resultados obtidos permitiram constatar que a vacina ERA foi eficaz contra todas as variantes de rua e silváticas do vírus rábico, independentemente do esquema vacinal. Contra as cepas adaptadas no laboratório DR-19 e CVS, quando aplicada em uma única dose, sua eficácia foi menor do que quando aplicada em 6 doses.Protection levels of an attenuated antirabies vaccine, of ERA origin, prepared in Kidney tissue culture, were evaluated in mice. Two schemes of vaccination were utilized: a single dose and 6 doses on alternate days. Animals of different experimental groups were challenged at 15, 30, 60 and 120 days post-immunization with six antigenic variants of rabies virus: strains of dog origin (S. Paulo, Brazil and Nigéria, vampire origin (DR-19 and Pernambuco, Brazil, fox origin (Rio Grande do Norte,Brazil, and CVS (Challenge Virus Standard. The results showed the effectiveness of the ERA vaccine against all antigenic variants of street and sylvatic rabies virus, independently of the vaccination scheme. Against the laboratory strains (DR-19 and CVS the ERA vaccine was less effective, mainly in the groups vaccinated with a single dose.

  14. Structure of hepatitis B surface antigen from subviral tubes determined by electron cryomicroscopy.

    Science.gov (United States)

    Short, Judith M; Chen, Shaoxia; Roseman, Alan M; Butler, P Jonathan G; Crowther, R Anthony

    2009-07-03

    Hepatitis B virus consists of an icosahedral core containing the double-stranded DNA genome, enveloped by a membrane with embedded surface proteins. The crystal structure of the core protein has been solved but little information about the structure of the surface proteins has so far been available. There are three sizes of surface protein, small (S), medium (M) and large (L), which form disulfide-bonded homo- and heterodimers. The three proteins, expressed from different start sites in the coding sequence, share the common C-terminal S region; the M protein contains an additional preS2 sequence N-terminal to S, and the L protein a further preS1 sequence N-terminal to M. In infected individuals, the surface proteins are produced in huge excess over the amount needed for viral envelopment and are secreted as a heterogeneous mixture of isometric and tubular subviral particles. We have used electron cryomicroscopy to study tubular particles extracted from human serum. Helical Fourier-Bessel analysis was used to calculate a low-resolution map, although it showed that the tubes were quite disordered. From the symmetry derived from this analysis, we used single-particle methods to improve the resolution. We found that the tubes had a diameter of approximately 250 A, with spike-like features projecting from the membrane. In the plane of the membrane the proteins appear to be close packed. We propose a model for the packing arrangement of surface protein dimers in the tubes.

  15. Cloning of a cDNA encoding a surface antigen of Schistosoma mansoni schistosomula recognized by sera of vassinated mice

    Energy Technology Data Exchange (ETDEWEB)

    Dalton, J.P.; Tom, T.D.; Strand, M.

    1987-06-01

    Spleen cells of mice vaccinated with radiation-attenuated Schistosoma mansoni cercariae were used to produce monoclonal antibodies directed against newly transformed schistosomular surface antigens. One of these monoclonal antibodies recognized a polypeptide of 18 kDa. Binding was measured by radioimmunoassay. This glycoprotein was purified by monoclonal antibody immunoaffinity chromatography and a polyclonal antiserum was prepared against it. Immunofluorescence assays showed that the polyclonal antiserum bound to the surface of newly transformed schistosomula and lung-stage organisms but not to the surface of liver-stage and adult worms. Using this polyclonal antiserum we isolated recombinant clones from an adult worm cDNA expression library constructed in lambdagt11. Clone 654.2 contained an insert of 0.52 kilobase and hybridized to a 1.2-kilobase mRNA species from adult worms. Most importantly, clone 654.2 produced a fusion protein of 125 kDa that was reactive with sera of vaccinated mice that are capable of transferring resistance. This result encourages future vaccination trials with the fusion protein.

  16. A new technique to detect antibody-antigen reaction (biological interactions) on a localized surface plasmon resonance (LSPR) based nano ripple gold chip

    Energy Technology Data Exchange (ETDEWEB)

    Saleem, Iram, E-mail: iiram.qau@gmail.com [Department of Physics and Texas Center for Superconductivity, University of Houston, Houston, TX 77204 (United States); Widger, William, E-mail: widger@uh.edu [Department of Biology and Biochemistry and Texas Center for Superconductivity, University of Houston, Houston, TX 77204 (United States); Chu, Wei-Kan, E-mail: wkchu@uh.edu [Department of Physics and Texas Center for Superconductivity, University of Houston, Houston, TX 77204 (United States)

    2017-07-31

    Highlights: • The nano ripple LSPR chip has monolayer molecule-coating sensitivity and specific selectivity. • Gold nano-ripple sensing chip is a low cost, and a label-free method for detecting the antibody-antigen reaction. • The plasmonic resonance shift depends upon the concentration of the biomolecules attached on the surface of the nano ripple pattern. - Abstract: We demonstrate that the gold nano-ripple localized surface plasmon resonance (LSPR) chip is a low cost and a label-free method for detecting the presence of an antigen. A uniform stable layer of an antibody was coated on the surface of a nano-ripple gold pattern chip followed by the addition of different concentrations of the antigen. A red shift was observed in the LSPR spectral peak caused by the change in the local refractive index in the vicinity of the nanostructure. The LSPR chip was fabricated using oblique gas cluster ion beam (GCIB) irradiation. The plasmon-resonance intensity of the scattered light was measured by a simple optical spectroscope. The gold nano ripple chip shows monolayer scale sensitivity and high selectivity. The LSPR substrate was used to detect antibody-antigen reaction of rabbit X-DENTT antibody and DENTT blocking peptide (antigen).

  17. Loss of the waaL O-antigen ligase prevents surface activation of the flagellar gene cascade in Proteus mirabilis.

    Science.gov (United States)

    Morgenstein, Randy M; Clemmer, Katy M; Rather, Philip N

    2010-06-01

    Proteus mirabilis is a Gram-negative bacterium that undergoes a physical and biochemical change from a vegetative swimmer cell (a typical Gram-negative rod) to an elongated swarmer cell when grown on a solid surface. In this study, we report that a transposon insertion in the waaL gene, encoding O-antigen ligase, blocked swarming motility on solid surfaces but had little effect on swimming motility in soft agar. The waaL mutant was unable to differentiate into a swarmer cell. Differentiation was also prevented by a mutation in wzz, encoding a chain length determinant for O antigen, but not by a mutation in wzyE, encoding an enzyme that polymerizes enterobacterial common antigen, a surface polysaccharide different from the lipid A::core. In wild-type P. mirabilis, increased expression of the flhDC operon occurs after growth on solid surfaces and is required for the high-level expression of flagellin that is characteristic of swarmer cells. However, in both the waaL and the wzz mutants, the flhDC operon was not activated during growth on agar. A loss-of-function mutation in the rcsB response regulator or overexpression of flhDC restored swarming to the waaL mutant, despite the absence of O antigen. Therefore, although O antigen may serve a role in swarming by promoting wettability, the loss of O antigen blocks a regulatory pathway that links surface contact with the upregulation of flhDC expression.

  18. Molecular Cloning, Carbohydrate Specificity and the Crystal Structure of Two Sclerotium rolfsii Lectin Variants.

    Science.gov (United States)

    Peppa, Vassiliki I; Venkat, Hemalatha; Kantsadi, Anastassia L; Inamdar, Shashikala R; Bhat, Ganapati G; Eligar, Sachin; Shivanand, Anupama; Chachadi, Vishwanath B; Satisha, Gonchigar J; Swamy, Bale M; Skamnaki, Vassiliki T; Zographos, Spyridon E; Leonidas, Demetres D

    2015-06-12

    SRL is a cell wall associated developmental-stage specific lectin secreted by Sclerotium rolfsii, a soil-born pathogenic fungus. SRL displays specificity for TF antigen (Galβ1→3GalNAc-α-Ser//Thr) expressed in all cancer types and has tumour suppressing effects in vivo. Considering the immense potential of SRL in cancer research, we have generated two variant gene constructs of SRL and expressed in E. coli to refine the sugar specificity and solubility by altering the surface charge. SSR1 and SSR2 are two different recombinant variants of SRL, both of which recognize TF antigen but only SSR1 binds to Tn antigen (GalNAcα-Ser/Thr). The glycan array analysis of the variants demonstrated that SSR1 recognizes TF antigen and their derivative with high affinity similar to SRL but showed highest affinity towards the sialylated Tn antigen, unlike SRL. The carbohydrate binding property of SSR2 remains unaltered compared to SRL. The crystal structures of the two variants were determined in free form and in complex with N-acetylglucosamine at 1.7 Å and 1.6 Å resolution, respectively. Structural analysis highlighted the structural basis of the fine carbohydrate specificity of the two SRL variants and results are in agreement with glycan array analysis.

  19. Molecular Cloning, Carbohydrate Specificity and the Crystal Structure of Two Sclerotium rolfsii Lectin Variants

    Directory of Open Access Journals (Sweden)

    Vassiliki I. Peppa

    2015-06-01

    Full Text Available SRL is a cell wall associated developmental-stage specific lectin secreted by Sclerotium rolfsii, a soil-born pathogenic fungus. SRL displays specificity for TF antigen (Galβ1→3GalNAc-α-Ser//Thr expressed in all cancer types and has tumour suppressing effects in vivo. Considering the immense potential of SRL in cancer research, we have generated two variant gene constructs of SRL and expressed in E. coli to refine the sugar specificity and solubility by altering the surface charge. SSR1 and SSR2 are two different recombinant variants of SRL, both of which recognize TF antigen but only SSR1 binds to Tn antigen (GalNAcα-Ser/Thr. The glycan array analysis of the variants demonstrated that SSR1 recognizes TF antigen and their derivative with high affinity similar to SRL but showed highest affinity towards the sialylated Tn antigen, unlike SRL. The carbohydrate binding property of SSR2 remains unaltered compared to SRL. The crystal structures of the two variants were determined in free form and in complex with N-acetylglucosamine at 1.7 Å and 1.6 Å resolution, respectively. Structural analysis highlighted the structural basis of the fine carbohydrate specificity of the two SRL variants and results are in agreement with glycan array analysis.

  20. Reduced neutrophil count in people of African descent is due to a regulatory variant in the Duffy antigen receptor for chemokines gene.

    Directory of Open Access Journals (Sweden)

    David Reich

    2009-01-01

    Full Text Available Persistently low white blood cell count (WBC and neutrophil count is a well-described phenomenon in persons of African ancestry, whose etiology remains unknown. We recently used admixture mapping to identify an approximately 1-megabase region on chromosome 1, where ancestry status (African or European almost entirely accounted for the difference in WBC between African Americans and European Americans. To identify the specific genetic change responsible for this association, we analyzed genotype and phenotype data from 6,005 African Americans from the Jackson Heart Study (JHS, the Health, Aging and Body Composition (Health ABC Study, and the Atherosclerosis Risk in Communities (ARIC Study. We demonstrate that the causal variant must be at least 91% different in frequency between West Africans and European Americans. An excellent candidate is the Duffy Null polymorphism (SNP rs2814778 at chromosome 1q23.2, which is the only polymorphism in the region known to be so differentiated in frequency and is already known to protect against Plasmodium vivax malaria. We confirm that rs2814778 is predictive of WBC and neutrophil count in African Americans above beyond the previously described admixture association (P = 3.8 x 10(-5, establishing a novel phenotype for this genetic variant.

  1. Antigen 43-mediated autotransporter display, a versatile bacterial cell surface presentation system

    DEFF Research Database (Denmark)

    Kjærgaard, Kristian; Hasman, Henrik; Schembri, Mark

    2002-01-01

    to the outer membrane and secretion through the cell envelope is contained within the protein itself. Ag43 consists of two subunits (alpha and beta), where the beta-subunit forms an integral outer membrane translocator to which the alpha-subunit is noncovalently attached. The simplicity of the Ag43 system...... makes it ideally suited as a surface display scaffold. Here we demonstrate that the Ag43 alpha-module can accommodate and display correctly folded inserts and has the ability to display entire functional protein domains, exemplified by the FimH lectin domain. The presence of heterologous cysteine...... bridges does not interfere with surface display, and Ag43 chimeras are correctly processed into alpha- and beta-modules, offering optional and easy release of the chimeric alpha-subunits. Furthermore, Ag43 can be displayed in many gram-negative bacteria. This feature is exploited for display of our...

  2. Seroprevalence of hepatitis B surface antigen in pregnant women attending antenatal clinic in Honiara Solomon Islands, 2015.

    Science.gov (United States)

    Getahun, Aneley; Baekalia, Margaret; Panda, Nixon; Lee, Alice; Puiahi, Elliot; Khan, Sabiha; Tahani, Donald; Manongi, Doris

    2016-12-08

    To determine the seroprevalence of hepatitis B surface antigen (HBsAg) among pregnant women attending antenatal clinic in Honiara, Solomon Islands. This descriptive cross-sectional study was carried out in seven area health centers in Honiara. From March to June 2015, identification of eligible pregnant women in each site was conducted using systematic random sampling technique. A total of 243 pregnant women who gave written informed consent were enrolled. Standardized tool was used to record demographics, obstetric history and serology results. HBsAg and hepatitis B e antigen (HBeAg) were tested using point-of-care rapid diagnostic test. All HBsAg positive samples were verified using enzyme-linked immunosorbent assay. The mean age of participants was 26 ± 6 years. The overall hepatitis HBsAg prevalence was 13.8% with higher rate (22%) reported in women between 30-34 years of age. Majority of HBsAg positive participants were Melanesians (29 out for 33). None of the pregnant women in the 15-19 years and ≥ 40 years tested positive for HBsAg. There was no statistically significant difference in HBsAg prevalence by age, ethnicity, education and residential location. The overall HBeAg seroprevalence was 36.7%. Women between 20-24 years of age had the highest rate of 54.5%. Low level of knowledge about hepatitis B vaccination was reputed. Overall, 54.6% of participants were not aware of their hepatitis B vaccination status and only 65.2% of mothers reported their child had been vaccinated. Hepatitis B is a disease of public health importance in Solomon Islands and emphasize the need for integrated preventative interventions for its control.

  3. Characterization of hepatitis B virus genotypes and quantitative hepatitis B surface antigen titres in North American tertiary referral liver centres.

    Science.gov (United States)

    Congly, Stephen E; Wong, Philip; Al-Busafi, Said A; Doucette, Karen; Fung, Scott K; Ghali, Peter; Fonseca, Kevin; Myers, Robert P; Osiowy, Carla; Coffin, Carla S

    2013-10-01

    Hepatitis B virus (HBV) genotype and quantitative hepatitis B surface antigen (qHBsAg) have been related to clinical outcome. In this nationwide cross-sectional study, we aimed to investigate the epidemiology and clinical significance of HBV genotype and qHBsAg in patients with chronic hepatitis B (CHB). Six hundred and thirty patients with CHB were seen in four urban tertiary referral centres in Canada. HBV genotype was determined by line probe assay (INNO-LIPA) and HBV DNA quantified by commercial PCR (Roche TaqMan, sensitivity <55 IU/ml or AMPLICOR, sensitivity <60 IU/ml). Titres of qHBsAg were determined by an in-house assay based on the WHO standard (calibration range 0.24-62.5 IU/ml). In 630 patients (57% male, 69% Asian, median age 42 years), 21% were hepatitis B e antigen positive and the median alanine aminotransferase was 29 U/L. The HBV genotype distribution was A (16%), B (29%), C (31%), D (16%), E (6%). HBV genotype was strongly associated with ethnicity, but neither genotype nor qHBsAg correlated with the degree of fibrosis. In the treatment-naïve patients, the baseline qHBsAg levels correlated with HBV DNA (r = 0.2517, P < 0.0008). The median qHBsAg levels were lowest in patients with genotype B (P < 0.0001), but no significant correlation was noted with all other HBV genotypes. In this large North American HBV epidemiological study, genotypes B and C were the most common; however, all genotypes (A-E) were observed with varied distribution nationwide. Baseline qHBsAg significantly correlated with HBV DNA and with HBV genotype B, but not with liver fibrosis. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. Hepatitis B surface antigen genetic elements critical for immune escape correlate with hepatitis B virus reactivation upon immunosuppression.

    Science.gov (United States)

    Salpini, Romina; Colagrossi, Luna; Bellocchi, Maria Concetta; Surdo, Matteo; Becker, Christina; Alteri, Claudia; Aragri, Marianna; Ricciardi, Alessandra; Armenia, Daniele; Pollicita, Michela; Di Santo, Fabiola; Carioti, Luca; Louzoun, Yoram; Mastroianni, Claudio Maria; Lichtner, Miriam; Paoloni, Maurizio; Esposito, Mariarosaria; D'Amore, Chiara; Marrone, Aldo; Marignani, Massimo; Sarrecchia, Cesare; Sarmati, Loredana; Andreoni, Massimo; Angelico, Mario; Verheyen, Jens; Perno, Carlo-Federico; Svicher, Valentina

    2015-03-01

    Hepatitis B virus (HBV) reactivation during immunosuppression can lead to severe acute hepatitis, fulminant liver failure, and death. Here, we investigated hepatitis B surface antigen (HBsAg) genetic features underlying this phenomenon by analyzing 93 patients: 29 developing HBV reactivation and 64 consecutive patients with chronic HBV infection (as control). HBsAg genetic diversity was analyzed by population-based and ultradeep sequencing (UDS). Before HBV reactivation, 51.7% of patients were isolated hepatitis B core antibody (anti-HBc) positive, 31.0% inactive carriers, 6.9% anti-HBc/anti-HBs (hepatitis B surface antibody) positive, 6.9% isolated anti-HBs positive, and 3.4% had an overt HBV infection. Of HBV-reactivated patients, 51.7% were treated with rituximab, 34.5% with different chemotherapeutics, and 13.8% with corticosteroids only for inflammatory diseases. In total, 75.9% of HBV-reactivated patients (vs. 3.1% of control patients; Pviral population as a predominant species. In control patients carrying such mutations, their median intrapatient prevalence was 4.6% (range, 2.5%-11.3%; Pimmunosuppressive settings at reactivation risk and of establishing a prompt therapy to prevent HBV-related clinical complications. © 2014 by the American Association for the Study of Liver Diseases.

  5. Characterization of the disassembly and reassembly of the HBV glycoprotein surface antigen, a pliable nanoparticle vaccine platform.

    Science.gov (United States)

    Gallagher, John R; Torian, Udana; McCraw, Dustin M; Harris, Audray K

    2017-02-01

    While nanoparticle vaccine technology is gaining interest due to the success of vaccines like those for the human papillomavirus that is based on viral capsid nanoparticles, little information is available on the disassembly and reassembly of viral surface glycoprotein-based nanoparticles. One such particle is the hepatitis B virus surface antigen (sAg) that exists as nanoparticles. Here we show, using biochemical analysis coupled with electron microscopy, that sAg nanoparticle disassembly requires both reducing agent to disrupt intermolecular disulfide bonds, and detergent to disrupt hydrophobic interactions that stabilize the nanoparticle. Particles were otherwise resistant to salt and urea, suggesting the driving mechanism of particle formation involves hydrophobic interactions. We reassembled isolated sAg protein into nanoparticles by detergent removal and reassembly resulted in a wider distribution of particle diameters. Knowledge of these driving forces of nanoparticle assembly and stability should facilitate construction of epitope-displaying nanoparticles that can be used as immunogens in vaccines. Published by Elsevier Inc.

  6. Consensus HIV-1 FSU-A integrase gene variants electroporated into mice induce polyfunctional antigen-specific CD4+ and CD8+ T cells.

    Directory of Open Access Journals (Sweden)

    Olga Krotova

    Full Text Available Our objective is to create gene immunogens targeted against drug-resistant HIV-1, focusing on HIV-1 enzymes as critical components in viral replication and drug resistance. Consensus-based gene vaccines are specifically fit for variable pathogens such as HIV-1 and have many advantages over viral genes and their expression-optimized variants. With this in mind, we designed the consensus integrase (IN of the HIV-1 clade A strain predominant in the territory of the former Soviet Union and its inactivated derivative with and without mutations conferring resistance to elvitegravir. Humanized IN gene was synthesized; and inactivated derivatives (with 64D in the active site mutated to V with and without elvitegravir-resistance mutations were generated by site-mutagenesis. Activity tests of IN variants expressed in E coli showed the consensus IN to be active, while both D64V-variants were devoid of specific activities. IN genes cloned in the DNA-immunization vector pVax1 (pVaxIN plasmids were highly expressed in human and murine cell lines (>0.7 ng/cell. Injection of BALB/c mice with pVaxIN plasmids followed by electroporation generated potent IFN-γ and IL-2 responses registered in PBMC by day 15 and in splenocytes by day 23 after immunization. Multiparametric FACS demonstrated that CD8+ and CD4+ T cells of gene-immunized mice stimulated with IN-derived peptides secreted IFN-γ, IL-2, and TNF-α. The multi-cytokine responses of CD8+ and CD4+ T-cells correlated with the loss of in vivo activity of the luciferase reporter gene co-delivered with pVaxIN plasmids. This indicated the capacity of IN-specific CD4+ and CD8+ T-cells to clear IN/reporter co-expressing cells from the injection sites. Thus, the synthetic HIV-1 clade A integrase genes acted as potent immunogens generating polyfunctional Th1-type CD4+ and CD8+ T cells. Generation of such response is highly desirable for an effective HIV-1 vaccine as it offers a possibility to attack virus

  7. Hepatitis B surface antigen quantity positively correlates with plasma levels of microRNAs differentially expressed in immunological phases of chronic hepatitis B in children

    DEFF Research Database (Denmark)

    Winther, Thilde Nordmann; Heiberg, Ida Louise; Bang-Berthelsen, Claus Heiner

    2013-01-01

    Children with chronic hepatitis B (CHB) are at high risk of progressive liver disease. It is suggested that a newly-identified panel of 16 microRNAs is important in the pathogenesis of CHB in children. Subviral hepatitis B surface antigen (HBsAg) particles are produced in large excess over...

  8. Kupffer cells interact with hepatitis B surface antigen in vivo and in vitro, leading to proinflammatory cytokine production and natural killer cell function

    NARCIS (Netherlands)

    Boltjes, Arjan; van Montfoort, Nadine; Biesta, Paula J; Op den Brouw, Marjoleine L; Kwekkeboom, Jaap; van der Laan, Luc J W; Janssen, Harry L A; Boonstra, André; Woltman, Andrea M

    2015-01-01

    BACKGROUND: Based on their localization, Kupffer cells (KCs) likely interact with hepatitis B virus (HBV). However, the role of KCs in inducing immunity toward HBV is poorly understood. Therefore, the interaction of hepatitis B surface antigen (HBsAg) and KCs, and possible functional consequences,

  9. P48 Major Surface Antigen of Mycoplasma agalactiae Is Homologous to a malp Product of Mycoplasma fermentans and Belongs to a Selected Family of Bacterial Lipoproteins

    OpenAIRE

    Rosati, Sergio; Pozzi, Sarah; Robino, Patrizia; Montinaro, Barbara; Conti, Amedeo; Fadda, Manlio; Pittau, Marco

    1999-01-01

    A major surface antigenic lipoprotein of Mycoplasma agalactiae, promptly recognized by the host's immune system, was characterized. The mature product, P48, showed significant similarity and shared conserved amino acid motifs with lipoproteins or predicted lipoproteins from Mycoplasma fermentans, Mycoplasma hyorhinis, relapsing fever Borrelia spp., Bacillus subtilis, and Treponema pallidum.

  10. P48 major surface antigen of Mycoplasma agalactiae is homologous to a malp product of Mycoplasma fermentans and belongs to a selected family of bacterial lipoproteins.

    Science.gov (United States)

    Rosati, S; Pozzi, S; Robino, P; Montinaro, B; Conti, A; Fadda, M; Pittau, M

    1999-11-01

    A major surface antigenic lipoprotein of Mycoplasma agalactiae, promptly recognized by the host's immune system, was characterized. The mature product, P48, showed significant similarity and shared conserved amino acid motifs with lipoproteins or predicted lipoproteins from Mycoplasma fermentans, Mycoplasma hyorhinis, relapsing fever Borrelia spp., Bacillus subtilis, and Treponema pallidum.

  11. 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...

  12. A 135-kilodalton surface antigen of Mycoplasma hominis PG21 contains multiple directly repeated sequences

    DEFF Research Database (Denmark)

    Ladefoged, Søren; Birkelund, Svend; Hauge, S

    1995-01-01

    A monoclonal antibody was used to characterize a 135-kDa surface-located membrane protein (Lmp1) generally present in Mycoplasma hominis strains. The monoclonal antibody, 552, was applied to identify the corresponding gene in an expression library of M. hominis PG21 DNA. The M. hominis PG21 lmp1......,000, was identified. Analysis of the deduced amino acid sequence predicted a hydrophilic protein with a basic pI (10.0). The N-terminal 24 amino acids were a typical leader sequence. Downstream from the first 726 nucleotides, six similar direct repeats of 471 nucleotides were found. In repeat 7, a single...

  13. Variant of a volume-of-fluid method for surface tension-dominant two ...

    Indian Academy of Sciences (India)

    The capabilities of the volume-of-fluid method for the calculation of surface tension-dominant two-phase flows are explained. The accurate calculation of the interface remains a problem for the volume-of-fluid method if the density ratios of the fluids in different phases are high. The simulations of bubble growth is performed ...

  14. Variant of a volume-of-fluid method for surface tension-dominant two ...

    Indian Academy of Sciences (India)

    2013-12-27

    Dec 27, 2013 ... J. Heat Transfer (ASME) 126: 329–338. Brackbill J U, Kothe D B and Zemach C 1992 A continuum method for modeling surface tension. J. Comput. Phys. 100: 335–354. Chakraborty I, Ray B, Biswas G, Durst F, Sharma A and Ghoshdastidar P S 2009 Computational inves- tigation on bubble detachement ...

  15. Identification of surface antigens of schistosomula of Schistosoma mansoni recognized by antibodies from mice immunized by chronic infection and by exposure to highly irradiated cercariae

    Energy Technology Data Exchange (ETDEWEB)

    Simpson, A.J.; James, S.L.; Sher, A.

    1983-08-01

    Surface components of mechanically transformed schistosomula of Schistosoma mansoni were labeled by lactoperoxidase-catalyzed iodination. After solubilization with Triton X-100, antigens were identified by immunoprecipitation. Serum from chronically infected Swiss mice reproducibly precipitated seven major polypeptides with approximate molecular weights (X 10/sup 3/) of 94, 68, 45, 40 to 32, 22, and 16. The antigens of molecular weights (X 10/sup 3/) of 94, 40 to 32, 22, and 16 were shown to be exposed on the parasite surface by interaction of the antibodies with intact labeled schistosomula. Sera from several strains of infected inbred mice precipitated the same polypeptides. The antibodies produced during chronic infection were found to be stimulated by adult worms since sera from 6-week-infected animals precipitated none of the surface antigens, and the pattern produced by precipitation with antibodies from a mouse infected with male worms only was indistinguishable from the pattern obtained with sera from mice with bisexual infections. Antibodies from mice immunized with highly irradiated cercariae reproducibly precipitated major polypeptides of approximately (X 10/sup 3/) 94, 68, 45, 32, 22, 19, and 15 daltons. The antigens of (X 10/sup 3/) 94, 43, 32, 22, and 15 daltons were shown to be exposed on the parasite surface by interaction of the antibodies with intact labeled schistosomula. The 15 X 10(3)-dalton surface protein was recognized by sera from vaccinated, but not chronically infected, mice, suggesting that it represents a stage-specific immunogen present on schistosomula but not on adult worms. Sera from two inbred strains of mice which develop different degrees of immunity recognized the same antigens.

  16. Evaluation of hepatitis B virus transmission and antiviral therapy among hepatitis B surface antigen-positive pregnant women.

    Science.gov (United States)

    Tekin Koruk, Suda; Batirel, Ayse; Kose, Sukran; Cetin Akhan, Sila; Aygen, Bilgehan; Tulek, Necla; Hatipoglu, Çigdem; Bulut, Cemal; Yıldız, Orhan; Sacligil, Cahide; Sirmatel, Fatma; Altunok, Elif

    2015-12-01

    The aim of the present study was to assess the potential risk of hepatitis B virus (HBV) vertical transmission among Turkish parturient women and to evaluate the efficacy and safety of antiviral agents. Data were collected retrospectively from 114 HBV-infected pregnant women and their infants in eight health institutions in Turkey. The baseline characteristics of the women were: mean age, 28.3 ± 5.2 years; alanine aminotransferase, 57.4 ± 139.0 U/L; aspartate aminotransferase, 56.6 ± 150.0 U/L; and HBV DNA, 8.3 × 10(7) ± 2.6 × 10(8) copies/mL. Family history of HBV infection was detected in 53.5% (n = 61). In total, 60 (52.6%) pregnant women received tenofovir (60.0%), lamivudine (33.3%) or telbivudine (6.7%) therapy at the median gestational age of 22.2 ± 8.5 (1-36) weeks. All infants were vaccinated and hepatitis B immune globulin was administered, with 81 of them (71.1%) available for follow-up. After completion of HBV vaccination course, 71 (87.7%) infants had protective anti-HBs levels, three (3.7%) were hepatitis B surface antigen-positive, and seven (8.6%) were hepatitis B surface antigen-negative with nonprotective anti-HBs levels. Five of the infants had low gestational birthweight but no other birth defects were observed. According to our results, viral load may not be the only effecting factor for transmission of HBV to children of infected mothers. Pregnant women with high viral load should be followed-up closely during pregnancy. They should begin to take tenofovir or telbivudine, which are category B drugs for pregnancy, at the beginning of the third trimester at the latest. We need new treatment strategies; and close follow-up of mothers and children is another important issue. © 2015 Japan Society of Obstetrics and Gynecology.

  17. Detection of specific antibodies directed against a consistently expressed surface antigen of Mycoplasma gallisepticum using a monoclonal blocking enzyme-linked immunosorbent assay.

    Science.gov (United States)

    Czifra, G; Kleven, S H; Engström, B; Stipkovits, L

    1995-01-01

    Sera from 14 groups of chickens inoculated with different laboratory and field strains of Mycoplasma gallisepticum (MG) were used to compare the diagnostic potential of the hemagglutination-inhibition (HI) test and a recently developed monoclonal blocking enzyme-linked immunosorbent assay (ELISA). HI was performed with strain A5969, commonly used as hemagglutinating antigen, and it could detect 62.7% of the inoculated chickens as positive. Of all sera, 83% proved to be positive when examined with the blocking ELISA. The difference between the sensitivities of the two methods was due to group-specific insensitivity of the HI test. None of the sera from groups inoculated with strains K 1501, K 1503, K 503, or K 703 and only half of the sera from groups inoculated with K 1453 or 236C could inhibit the activity of the A5969 hemagglutinating antigen, indicating antigenic differences between these challenge strains and the diagnostic strain. ELISA detected MG-specific antibodies in every group of sera, although inoculation with variant strains K 503 or K 703 resulted in lower level of antibody production than inoculation with other strains. The monoclonal blocking ELISA can be useful in the serological diagnosis of MG infections, because it is based on a consistently expressed, specific region of MG.

  18. Active late-appearing variable surface antigen genes in Trypanosoma equiperdum are constructed entirely from pseudogenes.

    Science.gov (United States)

    Roth, C; Bringaud, F; Layden, R E; Baltz, T; Eisen, H

    1989-12-01

    The expression of genes coding for variable surface glycoproteins (VSGs) in Trypanosoma equiperdum is linked to duplicative transpositions of silent, basic copy sequences into telomere-linked expression sites. Examination of three independently derived late-appearing trypanosome clones expressing VSG-78 revealed that the expressed gene in all cases is composed of sequences derived from three or four individual silent genes. The 182 base pairs at the 3' end of the coding sequence are derived from one silent gene, the 3' donor. The remaining 5' segment is a mosaic structure containing variable-length segments derived from two, or perhaps three, related silent genes. All of the silent genes that participate in the construction of the VSG-78 expression-linked copy (ELC) genes contain multiple stop codons and are unable to code for VSGs. Individual silent pseudogenes complement one another in the mosaic structure of the 5' portions of the ELC genes and create functional VSG genes. The joining of the 3' and 5' portions of the composite genes occurs in short regions of homology and suggests a mechanism by which the ordered expression of the VSG genes is generated.

  19. Unexpected interference in cell surface staining by monoclonal antibodies to unrelated antigens.

    Science.gov (United States)

    De Vita, Martina; Catzola, Valentina; Buzzonetti, Alexia; Fossati, Marco; Battaglia, Alessandra; Zamai, Loris; Fattorossi, Andrea

    2015-01-01

    The possible occurrence of an erroneous immunophenotyping due to interference between monoclonal antibodies (MoAbs) is often overlooked when the epitopes are assumed to be not close to each other. This is particularly important when exploring immune cell populations whose identification is still investigational. The commonly held view is that myeloid derived suppressor cells can be identified as either HLA-DR(neg/dim) cells or interleukin-4 receptor-α (CD124)(+) cells among peripheral blood monocytes. We made the serendipitous observation that the fluorescence signal provided by the PE-CD124 MoAb was attenuated when the PE-CF594-HLA-DR MoAb was added to the staining tube. Peripheral blood mononuclear cells from healthy donors were stained with the PE-CD124 MoAb and, as control, PE -CD40, -CD4 and -CD14, and either the PE-CF594-HLA-DR MoAb or its unlabeled form. B cells, which also express CD124, were analyzed for comparison. The PE-CF594-HLA-DR MoAb but not its unlabeled form reduced PE-CD124 MoAb staining on monocytes and B cells. No other monocyte and B cell surface marker staining was affected by the PE-CF594-HLA-DR MoAb. The PE-CF594-HLA-DR MoAb interfered with the PE-CD124 MoAb likely because of steric hindrance by bulky fluorochromes, although a quenching due to fluorescence resonance energy transfer might also cooperate to the PE-CD124 MoAb staining attenuation. Present observations highlight the importance of interference between MoAbs as a source of error when analyzing multicolor flow cytometry data. © 2014 International Clinical Cytometry Society.

  20. Analysis of cosmid clones of nuclear DNA from Trypanosome brucei shows that the genes for variant surface glycoproteins are clustered in the genome.

    NARCIS (Netherlands)

    D. Valerio (Dinko); T. de Lange; P. Borst (Piet); F.G. Grosveld (Frank); L.H.T. van der Ploeg

    1982-01-01

    textabstractTrypanosoma brucei contains more than a hundred genes coding for the different variant surface glycoproteins (VSGs). Activation of some of these genes involves the duplication of the gene (the basic copy or BC) and transposition of the duplicate to an expression site (yielding the

  1. A D+ blood donor with a novel RHD*D-CE(5-6)-D gene variant exhibits the low-frequency antigen RH23 (D(W) ) characteristic of the partial DVa phenotype.

    Science.gov (United States)

    Lopez, Genghis H; McGowan, Eunike C; McGrath, Kelli A; Abaca-Cleopas, Maria E; Schoeman, Elizna M; Millard, Glenda M; O'Brien, Helen; Liew, Yew-Wah; Flower, Robert L; Hyland, Catherine A

    2016-09-01

    Blood donors whose red blood cells (RBCs) exhibit a partial RhD phenotype, lacking some D epitopes, present as D+ in routine screening. Such phenotypes can exhibit low-frequency antigens (LFAs) of clinical significance. The aim of this study was to describe the serologic and genetic profile for a blood donor with an apparent D+ phenotype carrying a variant RHD gene where D Exons 5 and 6 are replaced by RHCE Exon (5-6). Anti-D monoclonal antibodies were used to characterize the presentation of RhD epitopes on the RBCs. RHD exon scanning and DNA sequencing of short- and long-range polymerase chain reaction amplicons were used to determine the RHD structure and sequence. Extended phenotyping for LFAs RH23 (D(W) ) and Rh32 was performed. The donor serology profile was consistent with partial RhD epitope presentation. The donor was hemizygous for an RHD variant allele described as RHD*D-CE(5-6)-D hybrid. The RHCE gene insert is at least 3.868 kb with 5' and 3' breakpoints between IVS4 + 132-c.667 and IVS6 + 1960-IVS6 + 2099, respectively. The sequence for this hybrid was assigned GenBank Accession Number KT099190.2. The RBCs were RH23 (D(W) )+ and Rh32-. A novel RHD*D-CE(5-6)-D hybrid allele encodes a partial RhD epitope and carries the LFA RH23 (D(W) ). This and the epitope profile resemble the partial DVa phenotype. Given that RBCs from this individual lack some RhD epitopes, there is an alloimmunization risk if the donor is exposed to D+ RBCs. Conversely, transfusions of RH23 (D(W) )+ cells to RH23 (D(W) )- recipients also pose an alloimmunization risk. © 2016 AABB.

  2. Hepatitis B Vaccination Coverage and Prevalence of Hepatitis B Surface Antigen Among Children in French Polynesia, 2014.

    Science.gov (United States)

    Patel, Minal K; Le Calvez, Evelyne; Wannemuehler, Kathleen; Ségalin, Jean-Marc

    2016-06-01

    French Polynesia is considered to be moderately endemic for chronic hepatitis B virus infection, with an estimated 3% of the population having hepatitis B surface antigen (HBsAg). From 1990 to 1992, a 3-dose hepatitis B vaccination series was introduced into the routine infant immunization schedule in French Polynesia, including a birth dose (BD). In 2014, a nationally representative 2-stage cluster survey was undertaken to evaluate the impact of the vaccination program on HBsAg prevalence among school children (∼6 years of age) in Cours Préparatoire (CP). Documented vaccination data were reviewed for all eligible children; children with consent were tested for HBsAg with a rapid point-of-care test. In total, 1,660 students were identified; 1,567 (94%) had vaccination data for review and 1,196 (72%) participated in the serosurvey. Three-dose vaccination coverage was 98%, while timely BD coverage, defined as a dose administered within 24 hours of life, was 89%. Receipt of the second and third doses was often delayed, with 75% and 55% receiving a second and third dose within 1 month of the recommended age, respectively. No children tested positive for HBsAg. French Polynesia's vaccination program has achieved high coverage and an HBsAg seroprevalence of 0% (0-0.5%) among CP school children, but timeliness of vaccination could be improved. © The American Society of Tropical Medicine and Hygiene.

  3. Anti-idiotype antibodies to Marek's disease-associated tumour surface antigen in protection against Marek's disease.

    Science.gov (United States)

    Dandapat, S; Pradhan, H K; Mohanty, G C

    1994-04-01

    Marek's disease-associated tumour surface antigen (MATSA) removed by enzymatic (papain) digestion of Marek's disease tumour cells was fractionated by gel filtration chromatography. The first peak (F1) was used to raise antibody in rabbits. Monoclonal antibody (RPH-6) directed against MATSA and the anti-F1 IgG were used as idiotypic antibodies to raise polyclonal anti-idiotype serum in heterologous hosts; rabbit and goat, respectively. The anti-idiotypes (anti-Id) were purified by affinity chromatography and characterized by competitive binding assay using immunofluorescent (IF) tests. Day-old white Leghorn chicks were immunized with anti-Id to MATSA (Group 1) or anti-Id to F1 (Group 3) and challenged with virulent Marek's disease virus (MDV) on the tenth day post immunization. In positive control groups, the day-old chicks were inoculated with anti-BALB/c mouse globulin (Group 2) and anti-rabbit globulin (Group 4) and challenged with virulent MDV on the tenth day post inoculation. As compared with positive control groups, the vaccinated groups (1 and 3) had considerably lower level of MATSA positive cells during the post challenge observation period. The protection level against MD in the immunized groups was 66.6% (Group 1) and 86.6% (Group 3).

  4. Characterization of surface antigen protein 1 (SurA1) from Acinetobacter baumannii and its role in virulence and fitness.

    Science.gov (United States)

    Liu, Dong; Liu, Zeng-Shan; Hu, Pan; Cai, Ling; Fu, Bao-Quan; Li, Yan-Song; Lu, Shi-Ying; Liu, Nan-Nan; Ma, Xiao-Long; Chi, Dan; Chang, Jiang; Shui, Yi-Ming; Li, Zhao-Hui; Ahmad, Waqas; Zhou, Yu; Ren, Hong-Lin

    2016-04-15

    Acinetobacter baumannii is a Gram-negative bacillus that causes nosocomial infections, such as bacteremia, pneumonia, and meningitis and urinary tract and wound infections. In the present study, the surface antigen protein 1 (SurA1) gene of A. baumannii strain CCGGD201101 was identified, cloned and expressed, and then its roles in fitness and virulence were investigated. Virulence was observed in the human lung cancer cell lines A549 and HEp-2 at one week after treatment with recombinant SurA1. One isogenic SurA1 knock-out strain, GR0015, which was derived from the A. baumannii strain CCGGD201101 isolated from diseased chicks in a previous study, highlighted the effect of SurA1 on fitness and growth. Its growth rate in LB broth and killing activity in human sera were significantly decreased compared with strain CCGGD201101. In the Galleria mellonella insect model, the isogenic SurA1 knock-out strain exhibited a lower survival rate and decreased dissemination. These results suggest that SurA1 plays an important role in the fitness and virulence of A. baumannii. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. [Prevalence of hepatitis B surface antigen and its associated factors in Senegalese military personnel sent on mission to Darfur].

    Science.gov (United States)

    Diop, Moustapha; Diouf, Assane; Seck, Said Malaobé; Lo, Gora; Ka, Daye; Massaly, Aminata; Dieye, Alassane; Fall, Ndeye Maguette; Cisse-Diallo, Viviane Marie Pierre; Diallo-Mbaye, Khardiata; Lakhe, Ndèye Aissatou; Fortes-Déguénonvo, Louise; Ndour, Cheikh Tidiane; Soumaré, Maserigne; Seydi, Moussa

    2017-01-01

    In Senegal, 85% of the adult population have been exposed to the hepatitis B virus and about 11% of them are chronic surface antigen (HBsAg) carriers. This infection is poorly documented among Senegalese Armed Forces. The aim of this study was to assess the prevalence of HBsAg in Senegalese military personnel on mission to Darfur (Sudan) and to identify its associated factors. We conducted a cross-sectional study among Senegalese military personnel stationed in Darfur from 1 July 2014 to 31 July 2014. HBsAg test was performed on serum of participants using immunochromatographic method. The search for associated factors was carried out using multivariate logistic regression. Our study included 169 male military personnel. The average age was 36.6 ± 9.5 years. A history of familial chronic liver disease, blood exposure and sexual exposure were found in 12.4%, 24.9% and 45.6% of the study population respectively. HBsAg was found in 24 participants [14.2% (CI 95% = 8.9-19.5)]. After adjusting for potential confounding factors, age (OR = 0.9 CI 95% = 0.9-1.0), university level (OR = 9.5 CI 95% = 1.3 - 67 , 1>) and sexual exposure (OR = 3.3 <; CI 95% = 1.0 - 10.3) were independently associated with hepatitis B. Our study shows high prevalence of HBsAg and underlines the need for further evaluation of hepatitis B in this population.

  6. A molecular assembly system for presentation of antigens on the surface of HBc virus-like particles

    Energy Technology Data Exchange (ETDEWEB)

    Blokhina, Elena A.; Kuprianov, Victor V. [Centre ' Bioengineering' , Russian Academy of Sciences, 117312 Prosp. 60-letya Oktyabrya 7-1, Moscow (Russian Federation); Stepanova, Ludmila A.; Tsybalova, Ludmila M. [Research Institute of Influenza, Russian Federation Ministry of Health and Social Development, St. Petersburg (Russian Federation); Kiselev, Oleg I. [Research Institute of Influenza, Russian Federation Ministry of Health and Social Development, St. Petersburg (Russian Federation); GenNanotech Ltd, St. Petersburg (Russian Federation); Ravin, Nikolai V., E-mail: nravin@biengi.ac.ru [Centre ' Bioengineering' , Russian Academy of Sciences, 117312 Prosp. 60-letya Oktyabrya 7-1, Moscow (Russian Federation); GenNanotech Ltd, St. Petersburg (Russian Federation); Skryabin, Konstantin G. [Centre ' Bioengineering' , Russian Academy of Sciences, 117312 Prosp. 60-letya Oktyabrya 7-1, Moscow (Russian Federation); GenNanotech Ltd, St. Petersburg (Russian Federation)

    2013-01-20

    Hepatitis B virus-like particles, icosahedral structures formed by multiple core protein dimers, are promising immune-enhancing vaccine carriers for foreign antigens. Insertions into the surface-exposed immunodominant loop are especially immunogenic. However, the need to conserve the particulate structure to ensure high immunogenicity imposes restraints on the nature of the heterologous sequence that can be inserted. We propose a new approach to constructing HBc particles linked to the target epitopes that relies on non-covalent interactions between the epitope and pre-assembled unmodified HBc particles. Interaction was enabled by fusion of the epitope to the GSLLGRMKGA peptide, binding to the spike tips. This peptide may be used as a 'binding tag' allowing in vitro construction of HBc particles carrying the target peptide. Such virus-like particles carrying multiple copies of the extracellular domain of the M2 protein of different influenza strains appeared to be highly immunogenic and protected immunised mice against a lethal influenza challenge.

  7. Establishment of an in vivo potency assay for the recombinant hepatit is B surface antigen in monovalent and combined vaccines

    Directory of Open Access Journals (Sweden)

    Mabel Izquierdo-López

    2014-12-01

    Full Text Available In this paper the development of potency assay in animals (mice was made, with the objective of demonstrating the immunogenic power of the recombinant Hepatitis B surface antigen in monovalent and combined vaccines, produced at the Center of Genetic Engineering and Biotechnology. The potency test is a parameter in quality control and it is also a tool to demonstrate the consistency of the production process. Parameters such as duration of the test, number of animals in the test, as well as different areas for the maintenance of the animals were evaluated. The results on the applicability of the potency test, to two presentations of the vaccines; monovalent Heberbiovac HB and pentavalent liquid in one vial Heberpenta-L are shown, for which specificity studies, evaluating different vaccine lots, the behavior of linearity, and parallelism, as well as establishing quality specification of the test were performed. This assay led to the obtainment of reliable results for the vaccines evaluated, the consistent evaluation of the immunogenic power and the monitoring of different production processes.

  8. Pectinesterase Inhibitor from Jelly Fig (Ficus awkeotsang Makino Achene Inhibits Surface Antigen Expression by Human Hepatitis B Virus

    Directory of Open Access Journals (Sweden)

    Yu-Chuen Huang

    2013-01-01

    Full Text Available Pectinesterase inhibitor (PEI isolated from jelly fig (Ficus awkeotsang Makino is an edible component of a popular drink consumed in Asia. Hepatitis B virus (HBV infection is prevalent in Asia, and current treatments for HBV infection need improvement. This study aimed to evaluate the effect of PEI on the surface antigen expression by HBV (HBsAg. Human hepatoma cell lines Hep3B and Huh7 served as in vitro models for assessing the cytotoxicity and HBsAg expression. A culture of primary hepatocytes cultured from mice served as the normal counterpart. Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT colorimetric assay. HBsAg expression was evaluated by measuring HBsAg secretion into the culture medium using an enzyme-linked immunosorbent assay. The results showed that PEI did not affect the viability of the human hepatoma cell lines or primary mouse hepatocytes. PEI inhibited the expression of HBsAg in hepatoma cell lines harboring endogenous (Hep3B and integrated (Huh7 HBV genomes in a concentration- and time-dependent manner, thus implicating a universal activity against HBV gene expression. In conclusion, it suggests that PEI from jelly fig inhibits the expression of human HBsAg in host cells without toxic effects on normal primary hepatocytes.

  9. Detection of Rubisco and mycotoxins as potential contaminants of a plantibody against the hepatitis B surface antigen purified from tobacco.

    Science.gov (United States)

    Geada, Déborah; Valdés, Rodolfo; Escobar, Arturo; Ares, Dulce M; Torres, Edel; Blanco, Reinaldo; Ferro, Williams; Dorta, Dayamí; González, Marcos; Alemán, María R; Padilla, Sigifredo; Gómez, Leonardo; Del Castillo, Norma; Mendoza, Otto; Urquiza, Dioslaida; Soria, Yordanka; Brito, José; Leyva, Alberto; Borroto, Carlos; Gavilondo, Jorge V

    2007-10-01

    Antibodies have been one of the proteins widely expressed in tobacco plants for pharmaceutical purposes, which demand contaminant free preparations. Rubisco constitutes 40-60% of tobacco leaf soluble proteins; therefore it is the major potential protein contaminant of plantibodies, while mycotoxins are toxic compounds that could be introduced during the biomass production and post-harvest stages with important consequences to human health. The objective of this paper was to investigate whether Rubisco and mycotoxins are present in Plantibody HB-01 preparations used in the immunopurification of the hepatitis B surface antigen. Rubisco was purified from Nicotiana tabacum yielding 154 microg of protein per gram of leaves and purity over 95%. Among mouse monoclonal antibodies generated against this enzyme, the CBSS.Rub-2 was selected for its immunodetection. It recognizes a conserved sequential epitope of Rubisco large subunit with an affinity constant of 0.13 x 10(8)M(-1). Rubisco quantification limit was 1 microg spreading to the measurement of this contaminant less than 4% of plantibodies samples. Additionally, according to a Reverse Phase-HPLC used to measure the level of adventitiously introduced contaminants, it can be concluded that aflatoxins B1, B2, G1 and G2 were undetected in the purified Plantibody HB-01 samples.

  10. Long term follow-up and outcome of liver transplantation from hepatitis B surface antigen positive donors

    Science.gov (United States)

    Ballarin, Roberto; Cucchetti, Alessandro; Russo, Francesco Paolo; Magistri, Paolo; Cescon, Matteo; Cillo, Umberto; Burra, Patrizia; Pinna, Antonio Daniele; Di Benedetto, Fabrizio

    2017-01-01

    Liver transplant for hepatitis B virus (HBV) currently yields excellent outcomes: it allows to rescue patients with an HBV-related advanced liver disease, resulting in a demographical modification of the waiting list for liver transplant. In an age of patient-tailored treatments, in liver transplantation as well the aim is to offer the best suitable graft to the patient who can benefit from it, also expanding the criteria for organ acceptance and allocation. With the intent of developing strategies to increase the donor pool, we set-up a multicenter study involving 3 Liver Transplant Centers in Italy: patients undergoing liver transplantation between March 03, 2004, and May 21, 2010, were retrospectively evaluated. 1408 patients underwent liver transplantation during the study period, 28 (2%) received the graft from hepatitis B surface antigen positive (HBsAg)-positive deceased donors. The average follow-up after liver transplantation was 63.7 mo [range: 0.1-119.4; SD ± 35.8]. None Primary non-function, re-liver transplantation, early or late hepatic artery thrombosis occurred. The 1-, 3- and 5-year graft and patient survival resulted of 85.7%, 82.1%, 78.4%. Our results suggest that the use of HBsAg-positive donors liver grafts is feasible, since HBV can be controlled without affecting graft stability. However, the selection of grafts and the postoperative antiviral therapy should be managed appropriately. PMID:28405138

  11. Europium nanoparticle-based simple to perform dry-reagent immunoassay for the detection of hepatitis B surface antigen.

    Science.gov (United States)

    Talha, Sheikh M; Salminen, Teppo; Juntunen, Etvi; Spangar, Anni; Gurramkonda, Chandrasekhar; Vuorinen, Tytti; Khanna, Navin; Pettersson, Kim

    2016-03-01

    Hepatitis B infection, caused by hepatitis B virus (HBV), presents a huge global health burden. Serological diagnosis of HBV mainly relies on the detection of hepatitis B surface antigen (HBsAg). Although there are high sensitivity commercial HBsAg enzyme immunoassays (EIAs) available, many low-resource laboratories lacking trained technicians continue to use rapid point-of-care assays with low sensitivities for HBsAg detection, due to their simplicity to operate. We developed a time-resolved fluorometric dry-reagent HBsAg immunoassay which meets the detection limit of high sensitivity EIAs but is simple to operate. To develop the assay, anti-HBsAg monoclonal antibody coated on europium nanoparticles was dried atop of biotinylated anti-HBsAg polyclonal antibody immobilized on streptavidin-coated microtiter wells. To test a sample in dry-reagent assay, serum sample and assay buffer were added to the wells, incubated, washed and europium signals were measured. The assay showed a detection limit of 0.25 ng/ml using HBsAg spiked in serum sample. When evaluated with 24 HBV positive and 37 negative serum samples, assay showed 100% sensitivity and specificity. Assay wells are stable for at least 26 weeks when stored at 4°C, and can tolerate elevated temperatures of up to 35°C for two weeks. The developed assay has high potential to be used in low-resource laboratories. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. First evaluation of the serum level of anti-hepatitis B surface antigen after vaccination in Libya.

    Science.gov (United States)

    Madour, A; Alkout, A; Vanin, S

    2013-12-01

    The hepatitis B virus (HBV) vaccination schedule in Libya follows international recommendations (1st dose at birth, 2nd after 1 month and 3rd after 6 months). This research aimed to evaluate the long-term protection of the HBV immunization programme in Tripoli and to determine the best age to administer booster doses. Serum levels of hepatitis B surface antigen were determined in 277 randomly selected children aged 1-12 years. The response to HBV vaccine in 1-3-year-olds was 93.2%, but this declined with age and at 7-9 years after initial vaccination only 53.1% of children had protective titres (> or = 10 mIU/mL). No significant differences between males and females in antibody persistence or response to vaccine were observed. We recommend continuing the HBV vaccination programme and that a booster dose be given to 6-year-old children to ensure maximum protection during the period of school entry and beyond.

  13. 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.

  14. The crystal structure of Haloferax volcanii proliferating cell nuclear antigen reveals unique surface charge characteristics due to halophilic adaptation

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    Morroll Shaun

    2009-08-01

    Full Text Available Abstract Background The high intracellular salt concentration required to maintain a halophilic lifestyle poses challenges to haloarchaeal proteins that must stay soluble, stable and functional in this extreme environment. Proliferating cell nuclear antigen (PCNA is a fundamental protein involved in maintaining genome integrity, with roles in both DNA replication and repair. To investigate the halophilic adaptation of such a key protein we have crystallised and solved the structure of Haloferax volcanii PCNA (HvPCNA to a resolution of 2.0 Å. Results The overall architecture of HvPCNA is very similar to other known PCNAs, which are highly structurally conserved. Three commonly observed adaptations in halophilic proteins are higher surface acidity, bound ions and increased numbers of intermolecular ion pairs (in oligomeric proteins. HvPCNA possesses the former two adaptations but not the latter, despite functioning as a homotrimer. Strikingly, the positive surface charge considered key to PCNA's role as a sliding clamp is dramatically reduced in the halophilic protein. Instead, bound cations within the solvation shell of HvPCNA may permit sliding along negatively charged DNA by reducing electrostatic repulsion effects. Conclusion The extent to which individual proteins adapt to halophilic conditions varies, presumably due to their diverse characteristics and roles within the cell. The number of ion pairs observed in the HvPCNA monomer-monomer interface was unexpectedly low. This may reflect the fact that the trimer is intrinsically stable over a wide range of salt concentrations and therefore additional modifications for trimer maintenance in high salt conditions are not required. Halophilic proteins frequently bind anions and cations and in HvPCNA cation binding may compensate for the remarkable reduction in positive charge in the pore region, to facilitate functional interactions with DNA. In this way, HvPCNA may harness its environment as

  15. Mechanisms Mediating Antigenic Variation in Trypanosoma brucei

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    Rudenko Gloria

    1999-01-01

    Full Text Available Antigenic variation in Trypanosoma brucei is a highly sophisticated survival strategy involving switching between the transcription of one of an estimated thousand variant surface glycoprotein (VSG genes. Switching involves either transcriptional control, resulting in switching between different VSG expression sites; or DNA rearrangement events slotting previously inactive VSG genes into an active VSG expression site. In recent years, considerable progress has been made in techniques allowing us to genetically modify infective bloodstream form trypanosomes. This is allowing us to reengineer VSG expression sites, and look at the effect on the mechanisms subsequently used for antigenic variation. We can now begin a dissection of a highly complicated survival strategy mediated by many different mechanisms operating simultaneously.

  16. In vitro evaluation of a soluble Leishmania promastigote surface antigen as a potential vaccine candidate against human leishmaniasis.

    Science.gov (United States)

    Chamakh-Ayari, Rym; Bras-Gonçalves, Rachel; Bahi-Jaber, Narges; Petitdidier, Elodie; Markikou-Ouni, Wafa; Aoun, Karim; Moreno, Javier; Carrillo, Eugenia; Salotra, Poonam; Kaushal, Himanshu; Negi, Narender Singh; Arevalo, Jorge; Falconi-Agapito, Francesca; Privat, Angela; Cruz, Maria; Pagniez, Julie; Papierok, Gérard-Marie; Rhouma, Faten Bel Haj; Torres, Pilar; Lemesre, Jean-Loup; Chenik, Mehdi; Meddeb-Garnaoui, Amel

    2014-01-01

    PSA (Promastigote Surface Antigen) belongs to a family of membrane-bound and secreted proteins present in several Leishmania (L.) species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice. We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S) produced in a L. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm) or L. braziliensis (CCLb) or visceral leishmaniasis due to L. donovani (CVLd) and in healthy individuals. Healthy individuals were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection) or non immune/naive individuals (HLR: Healthy Low Responders), depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection.

  17. In Vitro Evaluation of a Soluble Leishmania Promastigote Surface Antigen as a Potential Vaccine Candidate against Human Leishmaniasis

    Science.gov (United States)

    Bahi-Jaber, Narges; Petitdidier, Elodie; Markikou-Ouni, Wafa; Aoun, Karim; Moreno, Javier; Carrillo, Eugenia; Salotra, Poonam; Kaushal, Himanshu; Negi, Narender Singh; Arevalo, Jorge; Falconi-Agapito, Francesca; Privat, Angela; Cruz, Maria; Pagniez, Julie; Papierok, Gérard-Marie; Rhouma, Faten Bel Haj; Torres, Pilar; Lemesre, Jean-Loup; Chenik, Mehdi; Meddeb-Garnaoui, Amel

    2014-01-01

    PSA (Promastigote Surface Antigen) belongs to a family of membrane-bound and secreted proteins present in several Leishmania (L.) species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice. We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S) produced in a L. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm) or L. braziliensis (CCLb) or visceral leishmaniasis due to L. donovani (CVLd) and in healthy individuals. Healthy individuals were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection) or non immune/naive individuals (HLR: Healthy Low Responders), depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection. PMID:24786587

  18. In vitro evaluation of a soluble Leishmania promastigote surface antigen as a potential vaccine candidate against human leishmaniasis.

    Directory of Open Access Journals (Sweden)

    Rym Chamakh-Ayari

    Full Text Available PSA (Promastigote Surface Antigen belongs to a family of membrane-bound and secreted proteins present in several Leishmania (L. species. PSA is recognized by human Th1 cells and provides a high degree of protection in vaccinated mice. We evaluated humoral and cellular immune responses induced by a L. amazonensis PSA protein (LaPSA-38S produced in a L. tarentolae expression system. This was done in individuals cured of cutaneous leishmaniasis due to L. major (CCLm or L. braziliensis (CCLb or visceral leishmaniasis due to L. donovani (CVLd and in healthy individuals. Healthy individuals were subdivided into immune (HHR-Lm and HHR-Li: Healthy High Responders living in an endemic area for L. major or L. infantum infection or non immune/naive individuals (HLR: Healthy Low Responders, depending on whether they produce high or low levels of IFN-γ in response to Leishmania soluble antigen. Low levels of total IgG antibodies to LaPSA-38S were detected in sera from the studied groups. Interestingly, LaPSA-38S induced specific and significant levels of IFN-γ, granzyme B and IL-10 in CCLm, HHR-Lm and HHR-Li groups, with HHR-Li group producing TNF-α in more. No significant cytokine response was observed in individuals immune to L. braziliensis or L. donovani infection. Phenotypic analysis showed a significant increase in CD4+ T cells producing IFN-γ after LaPSA-38S stimulation, in CCLm. A high positive correlation was observed between the percentage of IFN-γ-producing CD4+ T cells and the released IFN-γ. We showed that the LaPSA-38S protein was able to induce a mixed Th1 and Th2/Treg cytokine response in individuals with immunity to L. major or L. infantum infection indicating that it may be exploited as a vaccine candidate. We also showed, to our knowledge for the first time, the capacity of Leishmania PSA protein to induce granzyme B production in humans with immunity to L. major and L. infantum infection.

  19. Carbohydrate antigen microarrays.

    Science.gov (United States)

    Wang, Denong

    2012-01-01

    This chapter describes one of my laboratory's working protocols for carbohydrate-based microarrays. Using a standard microarray spotter, we print carbohydrate antigens directly on the nitrocellulose-coated bioarray substrates. Because these substrates support noncovalent immobilization of many spotted antigens, in general no chemical modification of the antigen is needed for microarray production. Thus, this bioarray platform is technically simple and applicable for high-throughput construction of carbohydrate antigen microarrays. A number of nitrocellulose-coated glass slides with different technical characteristics are commercially available. Given the structural diversity of carbohydrate antigens, examining each antigen preparation to determine the efficacy of its immobilization in a given type of substrate and the surface display of the desired glycoepitopes in a microarray assay is essential.

  20. Development of a nasal vaccine for chronic hepatitis B infection that uses the ability of hepatitis B core antigen to stimulate a strong Th1 response against hepatitis B surface antigen.

    Science.gov (United States)

    Aguilar, J C; Lobaina, Y; Muzio, V; García, D; Pentón, E; Iglesias, E; Pichardo, D; Urquiza, D; Rodríguez, D; Silva, D; Petrovsky, N; Guillén, G

    2004-10-01

    There are estimated to be 350 million chronic carriers of hepatitis B infection worldwide. Patients with chronic hepatitis B are at risk of liver cirrhosis with associated mortality because of hepatocellular carcinoma and other complications. An important goal, therefore, is the development of an effective therapeutic vaccine against chronic hepatitis B virus (HBV). A major barrier to the development of such a vaccine is the impaired immune response to HBV antigens observed in the T cells of affected patients. One strategy to overcome these barriers is to activate mucosal T cells through the use of nasal vaccination because this may overcome the systemic immune downregulation that results from HBV infection. In addition, it may be beneficial to present additional HBV epitopes beyond those contained in the traditional hepatitis B surface antigen (HbsAg) vaccine, for example, by using the hepatitis B core antigen (HBcAg). This is advantageous because HBcAg has a unique ability to act as a potent Th1 adjuvant to HbsAg, while also serving as an immunogenic target. In this study we describe the effect of coadministration of HBsAg and HBcAg as part of a strategy to develop a more potent and effective HBV therapeutic vaccine. Copyright 2004 Australasian Society for Immunology Inc.

  1. Expression of Hepatitis B virus surface antigen (HBsAg from genotypes A, D and F and influence of amino acid variations related or not to genotypes on HBsAg detection

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    Natalia M. Araujo

    Full Text Available The impact of hepatitis B virus (HBV genotypes on the sensitivity of surface antigen (HBsAg detection assays has been poorly investigated. Here, plasmids carrying consensus or variant coding sequences for HBV surface proteins from genotypes A, D and F, were constructed. HBsAg levels were evaluated in medium and extracts of transfected CHO cells by a commercial polyclonal-based assay. We show that HBsAg detection values of consensus forms from genotypes D and F were, respectively, 37% and 30% lower than those obtained by genotype A. However, the presence of two single variations, T143M in genotype A, and T125M in genotype D, produced a decrease of 44% and an increase of 34%, respectively, on HBsAg mean values in comparison with their consensus forms. In conclusion, HBsAg detection levels varied among HBV genotypes. However, unique amino acid substitutions not linked to genotypes, such as T125M and T143M described here, should have more implications in HBV immunological diagnostics than the set of variations characteristic of each HBV genotype.

  2. Expression of Hepatitis B virus surface antigen (HBsAg) from genotypes A, D and F and influence of amino acid variations related or not to genotypes on HBsAg detection.

    Science.gov (United States)

    Araujo, Natalia M; Vianna, Carlos O A; Moraes, Marcia T B; Gomes, Selma A

    2009-08-01

    The impact of hepatitis B virus (HBV) genotypes on the sensitivity of surface antigen (HBsAg) detection assays has been poorly investigated. Here, plasmids carrying consensus or variant coding sequences for HBV surface proteins from genotypes A, D and F, were constructed. HBsAg levels were evaluated in medium and extracts of transfected CHO cells by a commercial polyclonal-based assay. We show that HBsAg detection values of consensus forms from genotypes D and F were, respectively, 37% and 30% lower than those obtained by genotype A. However, the presence of two single variations, T143M in genotype A, and T125M in genotype D, produced a decrease of 44% and an increase of 34%, respectively, on HBsAg mean values in comparison with their consensus forms. In conclusion, HBsAg detection levels varied among HBV genotypes. However, unique amino acid substitutions not linked to genotypes, such as T125M and T143M described here, should have more implications in HBV immunological diagnostics than the set of variations characteristic of each HBV genotype.

  3. Stem Cell Physics. Laser Manipulation of Blood Types: Laser-Stripping-Away of Red Blood Cell Surface Antigens

    Science.gov (United States)

    Stefan, V. Alexander

    2014-03-01

    A novel mechanism of importance for the transfusion medicine[2] is proposed. The interaction of ultrashort wavelength multilaser beams with the flowing blood thin films can lead to a conversion of blood types A, B, and AB into O type.[3] The stripping away of antigens is done by the scanning-multiple-lasers of a high repetition rate in the blue-purple frequency domain. The guiding-lasers are in the red-green frequency domain. The laser force, (parametric interaction with the antigen eigen-oscillation),[4] upon the antigen protein molecule must exceed its weight. Supported by Nikola Tesla Labs, La Jolla, CA.

  4. A Role of Sp1 Binding Motifs in Basal and Large T-Antigen-Induced Promoter Activities of Human Polyomavirus HPyV9 and Its Variant UF-1

    Directory of Open Access Journals (Sweden)

    Ugo Moens

    2017-11-01

    Full Text Available Human polyomavirus 9 (HPyV9 was originally detected in the serum of a renal transplant patient. Seroepidemiological studies showed that ~20–50% of the human population have antibodies against this virus. HPyV9 has not yet been associated with any disease and little is known about the route of infection, transmission, host cell tropism, and genomic variability in circulating strains. Recently, the HPyV9 variant UF-1 with an eight base-pair deletion, a thirteen base-pair insertion and with point mutations, creating three putative Sp1 binding sites in the late promoter was isolated from an AIDS patient. Transient transfection studies with a luciferase reporter plasmid driven by HPyV9 or UF1 promoter demonstrated that UF1 early and late promoters were stronger than HPyV9 promoters in most cell lines, and that the UF1 late promoter was more potently activated by HPyV9 large T-antigen (LTAg. Mutation of two Sp1 motifs strongly reduced trans-activation of the late UF1 promoter by HPyV9 LTAg in HeLa cells. In conclusion, the mutations in the UF1 late promoter seem to strengthen its activity and its response to stimulation by HPyV9 LTAg in certain cells. It remains to be investigated whether these promoter changes have an influence on virus replication and affect the possible pathogenic properties of the virus.

  5. Case of rhesus antigen weak D type 4.2. (DAR category detection

    Directory of Open Access Journals (Sweden)

    L. L. Golovkina

    2015-01-01

    Full Text Available Serological methods of Rhesus antigens identification in humans cannot identify D-antigen variants. In this article the serological characteristics of Rhesus antigen D weak type 4.2. (Category DAR are described.

  6. Focused antibody response to influenza linked to antigenic drift.

    Science.gov (United States)

    Huang, Kuan-Ying A; Rijal, Pramila; Schimanski, Lisa; Powell, Timothy J; Lin, Tzou-Yien; McCauley, John W; Daniels, Rodney S; Townsend, Alain R

    2015-07-01

    The selective pressure that drives antigenic changes in influenza viruses is thought to originate from the human immune response. Here, we have characterized the B cell repertoire from a previously vaccinated donor whose serum had reduced neutralizing activity against the recently evolved clade 6B H1N1pdm09 viruses. While the response was markedly polyclonal, 88% of clones failed to recognize clade 6B viruses; however, the ability to neutralize A/USSR/90/1977 influenza, to which the donor would have been exposed in childhood, was retained. In vitro selection of virus variants with representative monoclonal antibodies revealed that a single amino acid replacement at residue K163 in the Sa antigenic site, which is characteristic of the clade 6B viruses, was responsible for resistance to neutralization by multiple monoclonal antibodies and the donor serum. The K163 residue lies in a part of a conserved surface that is common to the hemagglutinins of the 1977 and 2009 H1N1 viruses. Vaccination with the 2009 hemagglutinin induced an antibody response tightly focused on this common surface that is capable of selecting current antigenic drift variants in H1N1pdm09 influenza viruses. Moreover, amino acid replacement at K163 was not highlighted by standard ferret antisera. Human monoclonal antibodies may be a useful adjunct to ferret antisera for detecting antigenic drift in influenza viruses.

  7. Recombinant hepatitis B surface antigen production in Aspergillus niger: evaluating the strategy of gene fusion to native glucoamylase.

    Science.gov (United States)

    James, Er; van Zyl, Wh; van Zyl, Pj; Görgens, Jf

    2012-10-01

    This study demonstrates the potential of Aspergillus niger as a candidate expression system for virus-like particle production using gene fusion. Hepatitis B surface antigen (HBsAg) production, targeted through the secretory pathway in A. niger, resulted in completely assembled and properly folded HBsAg. This was achieved by implementing a gene fusion strategy using the highly expressed catalytic domain of the native glucoamylase gene (GlaA ( G2 )) fused to the HBsAg S gene. The inducible glucoamylase promoter (GlaA ( p )) was used to control transcription in the A. niger D15 host. The gene fusion strategy was designed for cleavage of the fused product by the KEX2-like protease, resulting in intracellular accumulation of HBsAg and extracellular secretion of glucoamylase. Immunodetection using a monoclonal HBsAg antibody could not detect the fused GlaA ( G2 ) ::S product in intracellular and extracellular fractions, indicating that full assembly and maturation of HBsAg occurred after cleavage of the fused product in the Golgi complex. Several breakdown products showing an immunoreactive response to the glucoamylase polyclonal antibody indicated a level of intracellular degradation. The choice of carbon source used in cultivation significantly affected HBsAg production levels through induction of the glucoamylase promoter. The highest specific HBsAg production was observed during growth on inducing substrates of starch and its degradation products (maltodextrin and maltose), although residual glucose accumulation in the mid-exponential phase reduced HBsAg production. HBsAg production in starch-based cultures may be improved further by optimization of the rates of starch hydrolysis by glucoamylase and subsequent glucose consumption by the host.

  8. The trophoblast cell surface antigen 2 and miR-125b axis in urothelial bladder cancer.

    Science.gov (United States)

    Avellini, Chiara; Licini, Caterina; Lazzarini, Raffaella; Gesuita, Rosaria; Guerra, Emanuela; Tossetta, Giovanni; Castellucci, Clara; Giannubilo, Stefano Raffaele; Procopio, Antonio; Alberti, Saverio; Mazzucchelli, Roberta; Olivieri, Fabiola; Marzioni, Daniela

    2017-08-29

    Human trophoblast cell surface antigen 2 (Trop-2) is a 40-kDa transmembrane glycoprotein that was first identified as a marker of human trophoblast cells. Trop-2 acts on cell proliferation, adhesion, and migration by activating a number of intracellular signalling pathways. Elevated Trop-2 expression has been demonstrated in several types of cancer and correlated with aggressiveness and poor prognosis. Since no data are available on Trop-2 in bladder cancer (BC), the purpose of the study was to determine its levels in tissue specimens from normal individuals and patients with BC at different stages. Moreover, since according to recent evidence Trop-2 is a miR-125b target, miR-125b expression was also assessed in tissue specimens. Finally, the effect of the Trop-2/miR-125b axis on the proliferation and migration of BC cells was evaluated in vitro. The Trop-2/miR-125b axis was seen to be differentially expressed in normal urothelium, non-invasive BC and invasive BC tissue. Significant miR-125b down-regulation was associated with a significant increase in Trop-2 protein levels in BC tissue and correlated with disease severity. In vitro analysis confirmed the role of miR-125b in down-modulation of Trop-2 protein levels and showed that Trop-2/miR-125b axis affects cellular proliferation in bladder tissue. In conclusion, our findings highlight a role for the Trop-2/miR-125b axis in BC progression and suggest Trop-2 and miR-125b as diagnostic/prognostic marker candidates as well as druggable targets for innovative therapeutic approaches.

  9. Genetic variations in merozoite surface antigen genes of Babesia bovis detected in Vietnamese cattle and water buffaloes.

    Science.gov (United States)

    Yokoyama, Naoaki; Sivakumar, Thillaiampalam; Tuvshintulga, Bumduuren; Hayashida, Kyoko; Igarashi, Ikuo; Inoue, Noboru; Long, Phung Thang; Lan, Dinh Thi Bich

    2015-03-01

    The genes that encode merozoite surface antigens (MSAs) in Babesia bovis are genetically diverse. In this study, we analyzed the genetic diversity of B. bovis MSA-1, MSA-2b, and MSA-2c genes in Vietnamese cattle and water buffaloes. Blood DNA samples from 258 cattle and 49 water buffaloes reared in the Thua Thien Hue province of Vietnam were screened with a B. bovis-specific diagnostic PCR assay. The B. bovis-positive DNA samples (23 cattle and 16 water buffaloes) were then subjected to PCR assays to amplify the MSA-1, MSA-2b, and MSA-2c genes. Sequencing analyses showed that the Vietnamese MSA-1 and MSA-2b sequences are genetically diverse, whereas MSA-2c is relatively conserved. The nucleotide identity values for these MSA gene sequences were similar in the cattle and water buffaloes. Consistent with the sequencing data, the Vietnamese MSA-1 and MSA-2b sequences were dispersed across several clades in the corresponding phylogenetic trees, whereas the MSA-2c sequences occurred in a single clade. Cattle- and water-buffalo-derived sequences also often clustered together on the phylogenetic trees. The Vietnamese MSA-1, MSA-2b, and MSA-2c sequences were then screened for recombination with automated methods. Of the seven recombination events detected, five and two were associated with the MSA-2b and MSA-2c recombinant sequences, respectively, whereas no MSA-1 recombinants were detected among the sequences analyzed. Recombination between the sequences derived from cattle and water buffaloes was very common, and the resultant recombinant sequences were found in both host animals. These data indicate that the genetic diversity of the MSA sequences does not differ between cattle and water buffaloes in Vietnam. They also suggest that recombination between the B. bovis MSA sequences in both cattle and water buffaloes might contribute to the genetic variation in these genes in Vietnam. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Changes in seroprevalence of hepatitis B surface antigen and epidemiologic characteristics in the Republic of Korea, 1998-2013.

    Science.gov (United States)

    Lee, Hyerin; Lee, Hyungmin; Cho, Yumi; Oh, Kyungwon; Ki, Moran

    2015-01-01

    This study investigated changes in hepatitis B seroprevalence from 1998 to 2013, and to identify differences in epidemiologic characteristics between hepatitis B surface antigen (HBsAg)-positive and HBsAg-negative people. HBsAg seropositive rates were compared by year, sex, and age using the blood test data from the periods I to VI (1998-2013) of the Korea National Health and Nutrition Examination Survey. Interviews and self-administered surveys were conducted to collect data on health behavior, quality of life, comorbidities, and health care utilization. HBsAg seropositive rates in the Republic of Korea decreased from 4.6% in 1998 to 2.9% in 2008, and then remained the same for the next five years. While seropositive rates by age were the highest at 35 to 39 years of age in 1998, it peaked at 50 to 54 years of age in 2013. HBsAg-positive people showed high values from two liver function tests, including glutamic-oxaloacetic transaminase and glutamic-pyruvic transaminase, and the prevalence rates of liver cirrhosis and liver cancer were also significantly high. Indices for health behavior and quality of life showed no significant differences between HBsAg-positive and HBsAg-negative people. While HBsAg seropositive rates tended to decline after 1998, there have been no significant changes over the most recent five years. We should focus on treatment of existing hepatitis B patients along with immunization programmes to prevent new hepatitis B infections. In addition, it may be necessary to encourage HBsAg-positive people to follow healthier life-styles in order to prevent further progression of hepatitis B to liver cirrhosis and liver cancer.

  11. Interferon-alpha-induced changes in surface antigens in a hairy-cell leukemia (JOK-1), and a Burkitt's lymphoma cell line (Daudi) during in vitro culture

    DEFF Research Database (Denmark)

    Nielsen, B; Madsen, P S; Jensen, A W

    1992-01-01

    In further studying the mechanism of action of IFN-alpha in HCL, we cultured the HCL cell line JOK-1 and the IFN-sensitive Burkitt cell line Daudi with and without IFN-alpha and investigated the changes in density of a number of surface antigens by use of mAb and flow cytometry analyses. During...... culture with IFN-alpha, reproducible changes were induced in both cell lines, which were qualitatively similar but differed quantitatively with small and transient changes in JOK-1. Significant decreases in surface antigen expression were observed for CD 19, 23, 37, and for IgM on both cell lines...... of IFN-alpha in HCL was not paralleled by a specific direct effect on JOK-1 in vitro. Our findings therefore do not support the theory that IFN's mechanism of action in vivo is a direct effect on HC, but suggest that indirect effects are involved. Udgivelsesdato: 1992-Mar...

  12. A Novel Pan-Genome Reverse Vaccinology Approach Employing a Negative-Selection Strategy for Screening Surface-Exposed Antigens against leptospirosis.

    Science.gov (United States)

    Zeng, LingBing; Wang, Dongliang; Hu, NiYa; Zhu, Qing; Chen, Kaishen; Dong, Ke; Zhang, Yan; Yao, YuFeng; Guo, XiaoKui; Chang, Yung-Fu; Zhu, YongZhang

    2017-01-01

    Reverse vaccinology (RV) has been widely used for screening of surface-exposed proteins (PSEs) of important pathogens, including outer membrane proteins (OMPs), and extracellular proteins (ECPs) as potential vaccine candidates. In this study, we applied a novel RV negative strategy and a pan-genome analysis for screening of PSEs from 17 L. interrogans strains covering 11 predominately epidemic serovars and 17 multilocus typing (MLST) sequence types (STs) worldwide. Our results showed, for instance, out of a total of 633 predicted PSEs in strain 56601, 92.8% were OMPs or ECPs (588/633). Among the 17 strains, 190 core PSEs, 913 dispensable PSEs and 861 unique PSEs were identified. Of the 190 PSEs, 121 were further predicted to be highly antigenic and thus may serve as potential vaccine candidates against leptospirosis. With the exception of LipL45, OmpL1, and LigB, the majority of the 121 PSEs were newly identified antigens. For example, hypothetical proteins BatC, LipL71, and the OmpA family proteins sharing many common features, such as surface-exposed localization, universal conservation, and eliciting strong antibody responses in patients, are regarded as the most promising vaccine antigens. Additionally, a wide array of potential virulence factors among the predicted PSEs including TonB-dependent receptor, sphingomyelinase 2, leucine-rich repeat protein, and 4 neighboring hypothetical proteins were identified as potential antigenicity, and deserve further investigation. Our results can contribute to the prediction of suitable antigens as potential vaccine candidates against leptospirosis and also provide further insights into mechanisms of leptospiral pathogenicity. In addition, our novel negative-screening strategy combined with pan-genome analysis can be a routine RV method applied to numerous other pathogens.

  13. 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 mouse cerebellar cells by Fab fragments of both antibodies are at least additive, when compared with equal concentrations of the individual antibodies....

  14. A Conserved DNA Repeat Promotes Selection of a Diverse Repertoire of Trypanosoma brucei Surface Antigens from the Genomic Archive.

    Directory of Open Access Journals (Sweden)

    Galadriel Hovel-Miner

    2016-05-01

    Full Text Available African trypanosomes are mammalian pathogens that must regularly change their protein coat to survive in the host bloodstream. Chronic trypanosome infections are potentiated by their ability to access a deep genomic repertoire of Variant Surface Glycoprotein (VSG genes and switch from the expression of one VSG to another. Switching VSG expression is largely based in DNA recombination events that result in chromosome translocations between an acceptor site, which houses the actively transcribed VSG, and a donor gene, drawn from an archive of more than 2,000 silent VSGs. One element implicated in these duplicative gene conversion events is a DNA repeat of approximately 70 bp that is found in long regions within each BES and short iterations proximal to VSGs within the silent archive. Early observations showing that 70-bp repeats can be recombination boundaries during VSG switching led to the prediction that VSG-proximal 70-bp repeats provide recombinatorial homology. Yet, this long held assumption had not been tested and no specific function for the conserved 70-bp repeats had been demonstrated. In the present study, the 70-bp repeats were genetically manipulated under conditions that induce gene conversion. In this manner, we demonstrated that 70-bp repeats promote access to archival VSGs. Synthetic repeat DNA sequences were then employed to identify the length, sequence, and directionality of repeat regions required for this activity. In addition, manipulation of the 70-bp repeats allowed us to observe a link between VSG switching and the cell cycle that had not been appreciated. Together these data provide definitive support for the long-standing hypothesis that 70-bp repeats provide recombinatorial homology during switching. Yet, the fact that silent archival VSGs are selected under these conditions suggests the 70-bp repeats also direct DNA pairing and recombination machinery away from the closest homologs (silent BESs and toward the rest of

  15. A Conserved DNA Repeat Promotes Selection of a Diverse Repertoire of Trypanosoma brucei Surface Antigens from the Genomic Archive.

    Science.gov (United States)

    Hovel-Miner, Galadriel; Mugnier, Monica R; Goldwater, Benjamin; Cross, George A M; Papavasiliou, F Nina

    2016-05-01

    African trypanosomes are mammalian pathogens that must regularly change their protein coat to survive in the host bloodstream. Chronic trypanosome infections are potentiated by their ability to access a deep genomic repertoire of Variant Surface Glycoprotein (VSG) genes and switch from the expression of one VSG to another. Switching VSG expression is largely based in DNA recombination events that result in chromosome translocations between an acceptor site, which houses the actively transcribed VSG, and a donor gene, drawn from an archive of more than 2,000 silent VSGs. One element implicated in these duplicative gene conversion events is a DNA repeat of approximately 70 bp that is found in long regions within each BES and short iterations proximal to VSGs within the silent archive. Early observations showing that 70-bp repeats can be recombination boundaries during VSG switching led to the prediction that VSG-proximal 70-bp repeats provide recombinatorial homology. Yet, this long held assumption had not been tested and no specific function for the conserved 70-bp repeats had been demonstrated. In the present study, the 70-bp repeats were genetically manipulated under conditions that induce gene conversion. In this manner, we demonstrated that 70-bp repeats promote access to archival VSGs. Synthetic repeat DNA sequences were then employed to identify the length, sequence, and directionality of repeat regions required for this activity. In addition, manipulation of the 70-bp repeats allowed us to observe a link between VSG switching and the cell cycle that had not been appreciated. Together these data provide definitive support for the long-standing hypothesis that 70-bp repeats provide recombinatorial homology during switching. Yet, the fact that silent archival VSGs are selected under these conditions suggests the 70-bp repeats also direct DNA pairing and recombination machinery away from the closest homologs (silent BESs) and toward the rest of the archive.

  16. A lactate dehydrogenase (LDH)-based immunoassay for detection of cell surface antigens and its application to the study of MHC class I-binding peptides.

    Science.gov (United States)

    Sigal, L J; Berens, S; Wylie, D

    1994-12-28

    A lactate dehydrogenase (LDH)-based immunoassay, referred to as CPEIA (cell panning enzyme immunoassay), has been developed for the detection of cell-surface antigens. CPEIA is similar to a panning assay, in that it is based on the capture of cells bearing an antigen of interest by means of an antibody immobilized to a 96-well microtiter plate. Attachment of the cells is then measured by addition of a substrate for the intracellular enzyme lactate dehydrogenase. The substrate solution also contains the nonionic detergent Triton X-100 to lyse the cells and release LDH, which converts the substrate p-iodonitrotetrazolium violet (INT) from yellow to red. The intensity of the color resulting from the LDH-catalyzed reaction is proportional to the number of cells bound to the plate. The procedure does not require fixation of the cells, centrifugation, and blocking steps, resulting in a more convenient assay. CPEIA has been used for the detection of MHC class I antigens and other molecules on the surfaces of mouse cell lines and concanavalin A (ConA)-stimulated T lymphocytes. In addition, the assay has been used to detect peptide binding to Db and Kb MHC class I molecules on the surface of the mutant cell line RMA-S. The half-maximal responses for peptide-MHC class I interactions at different peptide concentrations can be determined with the assay, allowing the apparent dissociation constants to be calculated.

  17. The Taenia saginata homologue of the major surface antigen of Echinococcus spp. is immunogenic and 97% identical to its Taenia solium homologue.

    Science.gov (United States)

    González, Luis Miguel; Ferrer, Elizabeth; Spickett, Andrea; Michael, Lynne M; Vatta, Adriano F; Gárate, Teresa; Harrison, Leslie J S; Parkhouse, R Michael E

    2007-11-01

    The TEG-Tsag gene of Taenia saginata is homologous to the genes expressing the two major surface antigens of Echinococcus spp. (EM10 and EG10). Surface antigens of parasites are logical candidates for vaccines, and in this paper we demonstrate that cattle vaccinated with the recombinant TEG-Tsag protein, either used singly or in conjunction with the recombinant HP6-Tsag protein, the major 18 kDa surface/secreted antigen of T. saginata oncospheres, produce excellent antibody responses to both these recombinant proteins. Thus TEG-Tsag may have utility as a vaccine and also as a diagnostic tool for bovine cysticercosis. In addition, as we now demonstrate a 97% homology between TEG-Tsag and its Taenia solium homologue, TEG-Tsol, this latter molecule may have similar potential in the control of human and porcine cysticercosis. The TEG molecule is characterized by an N-terminal FERM domain and a C-terminal ERM domain which are found in a number of cytoskeletal-associated proteins located at the interface between the plasma membrane and the cytoskeleton and in proteins that interact with lipid membranes. The FERM domain is also postulated to bind to adhesion proteins, in a PIP2-regulated fashion, providing a link between cytoskeletal signals and membrane dynamics. Thus TEG protein may play a role in tegument function and interaction with the host.

  18. An Extended Surface Loop on Toxoplasma gondii Apical Membrane Antigen 1 (AMA1 Governs Ligand Binding Selectivity.

    Directory of Open Access Journals (Sweden)

    Michelle L Parker

    Full Text Available Apicomplexan parasites are the causative agents of globally prevalent diseases including malaria and toxoplasmosis. These obligate intracellular pathogens have evolved a sophisticated host cell invasion strategy that relies on a parasite-host cell junction anchored by interactions between apical membrane antigens (AMAs on the parasite surface and rhoptry neck 2 (RON2 proteins discharged from the parasite and embedded in the host cell membrane. Key to formation of the AMA1-RON2 complex is displacement of an extended surface loop on AMA1 called the DII loop. While conformational flexibility of the DII loop is required to expose the mature RON2 binding groove, a definitive role of this substructure has not been elucidated. To establish a role of the DII loop in Toxoplasma gondii AMA1, we engineered a form of the protein where the mobile portion of the loop was replaced with a short Gly-Ser linker (TgAMA1ΔDIIloop. Isothermal titration calorimetry measurements with a panel of RON2 peptides revealed an influential role for the DII loop in governing selectivity. Most notably, an Eimeria tenella RON2 (EtRON2 peptide that showed only weak binding to TgAMA1 bound with high affinity to TgAMA1ΔDIIloop. To define the molecular basis for the differential binding, we determined the crystal structure of TgAMA1ΔDIIloop in complex with the EtRON2 peptide. When analyzed in the context of existing AMA1-RON2 structures, spatially distinct anchor points in the AMA1 groove were identified that, when engaged, appear to provide the necessary traction to outcompete the DII loop. Collectively, these data support a model where the AMA1 DII loop serves as a structural gatekeeper to selectively filter out ligands otherwise capable of binding with high affinity in the AMA1 apical groove. These data also highlight the importance of considering the functional implications of the DII loop in the ongoing development of therapeutic intervention strategies targeting the AMA1-RON

  19. Human Leukocyte Antigen (HLA Class I Down-Regulation by Human Immunodeficiency Virus Type 1 Negative Factor (HIV-1 Nef: What Might We Learn From Natural Sequence Variants?

    Directory of Open Access Journals (Sweden)

    Philip Mwimanzi

    2012-09-01

    Full Text Available HIV-1 causes a chronic infection in humans that is characterized by high plasma viremia, progressive loss of CD4+ T lymphocytes, and severe immunodeficiency resulting in opportunistic disease and AIDS. Viral persistence is mediated in part by the ability of the Nef protein to down-regulate HLA molecules on the infected cell surface, thereby allowing HIV-1 to evade recognition by antiviral CD8+ T lymphocytes. Extensive research has been conducted on Nef to determine protein domains that are required for its immune evasion activities and to identify critical cellular co-factors, and our mechanistic understanding of this process is becoming more complete. This review highlights our current knowledge of Nef-mediated HLA class I down-regulation and places this work in the context of naturally occurring sequence variation in this protein. We argue that efforts to fully understand the critical role of Nef for HIV-1 pathogenesis will require greater analysis of patient-derived sequences to elucidate subtle differences in immune evasion activity that may alter clinical outcome.

  20. A microRNA derived from an apparent canonical biogenesis pathway regulates variant surface protein gene expression in Giardia lamblia

    Science.gov (United States)

    Saraiya, Ashesh A.; Li, Wei; Wang, Ching C.

    2011-01-01

    We have previously shown that a snoRNA-derived microRNA, miR2, in Giardia lamblia potentially regulates the expression of 22 variant surface protein (VSP) genes. Here, we identified another miRNA, miR4, also capable of regulating the expression of several VSPs but derived from an unannotated open reading frame (ORF) rather than a snoRNA, suggesting a canonical miRNA biogenesis pathway in Giardia. miR4 represses expression of a reporter containing two miR4 antisense sequences at the 3′ UTR without causing a corresponding decrease in the mRNA level. This repression requires the presence of the Giardia Argonaute protein (GlAgo) and is reversed by 2′ O–methylated antisense oligo to miR4, suggesting an RNA-induced silencing complex (RISC)–mediated mechanism. Furthermore, in vivo and in vitro evidence suggested that the Giardia Dicer protein (GlDcr) is required for miR4 biogenesis. Coimmunoprecipitation of miR4 with GlAgo further verified miR4 as a miRNA. A total of 361 potential target sites for miR4 were bioinformatically identified in Giardia, out of which 69 (32.7%) were associated with VSP genes. miR4 reduces the expression of a reporter containing two copies of the target site from VSP (GL50803_36493) at the 3′ UTR. Sixteen of the 69 VSP genes were further found to contain partially overlapping miR2 and miR4 targeting sites. Expression of a reporter carrying the two overlapping sites was inhibited by either miR2 or miR4, but the inhibition was neither synergistic nor additive, suggesting a complex mechanism of miRNA regulation of VSP expression and the presence of a rich miRNAome in Giardia. PMID:22033329

  1. Phage display used for gene cloning of human recombinant antibody against the erythrocyte surface antigen, rhesus D

    DEFF Research Database (Denmark)

    Dziegiel, Morten Hanefeld; Nielsen, L K; Andersen, P S

    1995-01-01

    A novel phage display system has been developed for PCR amplification and cloning of the Fab fragments of human immunoglobulin genes. Using this system, we have cloned an antibody from a mouse-human hybridoma cell line directed against the erythrocyte antigen rhesus D. Intact erythrocytes were used...... Fab phages demonstrates that it is possible to by-pass purification of the antigen of interest. Comparison with published germline sequences demonstrated that the immunoglobulin coding regions had the highest homology to the VH 1.9III and V kappa Hum kappa v325 germline genes, respectively....

  2. The antigen presentation function of bone marrow-derived mast cells is spatiotemporally restricted to a subset expressing high levels of cell surface FcεRI and MHC II

    Directory of Open Access Journals (Sweden)

    Sun Liangwu

    2010-06-01

    Full Text Available Abstract Background At present, it is highly controversial whether pure mast cells can serve as antigen presenting cells, and it is not known whether the capacity of antigen presenting function is temporally restricted to a particular subset of differentiated mast cells. Evidence is presented for a novel surface FcεRIhi , MHC II +, and c-kit + pure mast cell subset, temporally restricted as antigen-presenting cells in the immune axis of T-cell activation. Results Bone marrow-derived mast cells (BMMC cultured in the presence of IL-3 for three weeks are pure mast cells based on surface expression of lineage-specific marker, c-kit and FcεRI. Herein we present the first demonstration that approximately 98.7% c-kit + and FcεRI expressing BMMC, further depleted of any contaminated professional antigen-presenting cells, are still fully capable of presenting antigens, i.e., OVA protein, OVA peptide, and IgE-TNP-OVA, to OVA peptide-specific T-cell hybridomas. Notably, IgE-dependent antigen presentation is more efficient compared to that resulting from direct antigen uptake. Importantly, we present the novel finding that only surface FcεRIhi mast cells, also expressing surface MHC II exhibited antigen-presenting function. In contrast, surface FcεRIlo mast cells without expressing surface MHC II were not capable of antigen presentation. Interestingly, the antigen-presenting function of BMMC was irrevocably lost during the third and fourth week in IL-3 or SCF containing cultures. Conclusions This is the first observation to attribute a spatiotemporally restricted antigen-presenting function to a subset of three-week old pure BMMC expressing both high levels of surface FcεRI and surface MHC II. We propose that mast cells play an important role in immune deviating and/or sustaining the activation of infiltrating CD4 T-cells, and modulating T-cell mediated allergic inflammation via its flexibility to present antigens and antigen-IgE complexes.

  3. Diagnostic accuracy of tests to detect hepatitis B surface antigen: a systematic review of the literature and meta-analysis

    Directory of Open Access Journals (Sweden)

    Ali Amini

    2017-11-01

    Full Text Available Abstract Background Chronic Hepatitis B Virus (HBV infection is characterised by the persistence of hepatitis B surface antigen (HBsAg. Expanding HBV diagnosis and treatment programmes into low resource settings will require high quality but inexpensive rapid diagnostic tests (RDTs in addition to laboratory-based enzyme immunoassays (EIAs to detect HBsAg. The purpose of this review is to assess the clinical accuracy of available diagnostic tests to detect HBsAg to inform recommendations on testing strategies in 2017 WHO hepatitis testing guidelines. Methods The systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA guidelines using 9 databases. Two reviewers independently extracted data according to a pre-specified plan and evaluated study quality. Meta-analysis was performed. HBsAg diagnostic accuracy of rapid diagnostic tests (RDTs was compared to enzyme immunoassay (EIA and nucleic-acid test (NAT reference standards. Subanalyses were performed to determine accuracy among brands, HIV-status and specimen type. Results Of the 40 studies that met the inclusion criteria, 33 compared RDTs and/or EIAs against EIAs and 7 against NATs as reference standards. Thirty studies assessed diagnostic accuracy of 33 brands of RDTs in 23,716 individuals from 23 countries using EIA as the reference standard. The pooled sensitivity and specificity were 90.0% (95% CI: 89.1, 90.8 and 99.5% (95% CI: 99.4, 99.5 respectively, but accuracy varied widely among brands. Accuracy did not differ significantly whether serum, plasma, venous or capillary whole blood was used. Pooled sensitivity of RDTs in 5 studies of HIV-positive persons was lower at 72.3% (95% CI: 67.9, 76.4 compared to that in HIV-negative persons, but specificity remained high. Five studies evaluated 8 EIAs against a chemiluminescence immunoassay reference standard with a pooled sensitivity and specificity of 88.9% (95% CI: 87.0, 90.6 and

  4. Serum HBV surface antigen positivity is associated with low prevalence of metabolic syndrome: A meta-analysis.

    Directory of Open Access Journals (Sweden)

    Yuanyuan Li

    Full Text Available As there is conflicting evidence for the relationship between hepatitis B virus surface antigen (HBsAg positivity and the prevalence of metabolic syndrome (MetS, we performed a meta-analysis to investigate whether HBsAg positivity affects the incidence of MetS.Observational studies on the relationship between HBsAg positivity and MetS were obtained from PubMed, Web of Science, and the Cochrane Library in April 2016. The pooled odds ratios (ORs of MetS and its components (central obesity, increased fasting glucose, increased blood pressure, dyslipidemia for subjects with or without HBsAg positivity were synthesized. The standardized mean difference of MetS components between HBsAg-positive participants and healthy controls was calculated. Heterogeneity was explored with subgroup analysis and sensitivity analysis. Publication bias was detected using Egger's test and Begg's test.Thirty studies were eligible for meta-analysis. The MetS OR for HBsAg-positive participants was significantly decreased compared with the controls [OR = 0.80, 95% confidence interval (CI, 0.70-0.90]. The negative effect of HBsAg positivity on elevated triglycerides (OR = 0.62, 95% CI, 0.59-0.64 was strong, while that for increased fasting blood glucose was weak (OR = 0.94, 95% CI, 0.90-0.98. The pooled ORs of central obesity (OR = 0.97, 95% CI, 0.91-1.04, reduced high-density lipoprotein cholesterol (OR = 0.98, 95% CI, 0.83-1.14, and elevated blood pressure (OR = 1.00, 95% CI, 0.80-1.25 for HBsAg-positive participants were all not significantly different compared with the controls. No publication bias was detected.Serum HBsAg positivity is inversely associated with the prevalence of MetS. Among the five components of MetS, elevated triglycerides had the strongest inverse relationship with HBsAg positivity.

  5. Effects of Diabetes and Glycemia Control on Risk of Hepatocellular Carcinoma After Seroclearance of Hepatitis B Surface Antigen.

    Science.gov (United States)

    Cheuk-Fung Yip, Terry; Wai-Sun Wong, Vincent; Lik-Yuen Chan, Henry; Tse, Yee-Kit; Pik-Shan Kong, Alice; Long-Yan Lam, Kelvin; Chung-Yan Lui, Grace; Lai-Hung Wong, Grace

    2017-12-12

    Diabetes is associated with a 2-fold increase in risk of hepatocellular carcinoma (HCC) among patients with chronic hepatitis B virus (HBV) infection. However, we know little about the effect of diabetes on HCC risk after seroclearance of hepatitis B surface antigen (HBsAg). We evaluated the effect of diabetes and glycemic control on HCC development after HBsAg seroclearance in a population-wide study in Hong Kong. We performed a retrospective study of 4568 patients with chronic HBV infection who cleared HBsAg from January 2000 through August 2016, using the Clinical Data Analysis and Reporting System of the Hospital Authority, Hong Kong. We collected and analyzed data on patient demographics, comorbidities, medications, laboratory test results, and subsequent development of HCC. The presence of diabetes was defined by International Classification of Diseases, Ninth Revision, Clinical Modification diagnosis code, with level of hemoglobin; A1c (HbA1c) above 6.5%, fasting glucose level of 7 mmol/L or more, and/or treatment with; any anti-diabetic agent. We identified 1560 patients with diabetes; 29 patients (1.9%) developed HCC after a median follow-up time of 3.4 years (interquartile range, 1.5-5.0 years). Diabetes was associated with increased risk of HCC after adjustment of age, sex, presence of cirrhosis, and the use of medications (adjusted hazard ratio, 1.85; 95% CI 1.04-3.28; P=.036). Among patients with diabetes, time-weighted average level of HbA1c was an independent risk factor for HCC, after adjustment for age at clearance, use of statins, and other important covariates (adjusted hazard ratio, 1.51, 95% CI 1.20-1.91; P<.001). A time-weighted average level of HbA1c of 7% or more was associated with a higher 5-year cumulative incidence of HCC (4.0%) than a time-weighted average HbA1c level below 7% (1.8%) (log-rank test P=.035). In a population-based analysis of patients with chronic HBV infection in Hong Kong, we found diabetes to be an independent risk

  6. Molecular cloning of a putative agglutination/immobilization antigen located on the surface of a novel agglutination/immobilization serotype of Cryptocaryon irritans.

    Science.gov (United States)

    Hatanaka, A; Umeda, N; Hirazawa, N

    2008-08-01

    A surface agglutination/immobilization antigen was purified from the novel agglutination/immobilization serotype (serotype G37) of the ciliated protozoan Cryptocaryon irritans, a parasite of seawater fishes. Serum from fish immunized with C. irritans theronts had agglutination/immobilization activity against theronts in vitro. However, fish and rabbit antisera raised against serotype G32 (reported previously) caused little agglutination/immobilization of serotype G37 theronts. Immunological analysis indicated that the 37 kDa theront surface membrane protein may be the agglutination/immobilization antigen of this serotype. The full-length 37 kDa antigen cDNA contained 1171 base pairs, encoding a 331-amino acid protein with hydrophobic N- and C-termini, which are characteristically found in proteins containing a C-terminal glycosylphosphatidylinositol anchor. In addition, the genetically characterized nucleotide sequences of the first internal transcribed spacer region of ribosomal DNA of these 2 serotypes were compared. The internal transcribed spacer rDNA sequence of serotype G32 was identical to that of isolates from Pingtung, Taiwan, and from the USA. On the other hand, the sequences of serotype G37 were not identical to those of any C. irritans isolate.

  7. Chronic pneumonia in calves after experimental infection with Mycoplasma bovis strain 1067: Characterization of lung pathology, persistence of variable surface protein antigens and local immune response

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    Hermeyer Kathrin

    2012-02-01

    Full Text Available Abstract Background Mycoplasma bovis is associated with pneumonia in calves characterized by the development of chronic caseonecrotic lesions with the agent persisting within the lesion. The purposes of this study were to characterize the morphology of lung lesions, examine the presence of M. bovis variable surface protein (Vsp antigens and study the local immune responses in calves after infection with M. bovis strain 1067. Methods Lung tissue samples from eight calves euthanased three weeks after experimental infection with M. bovis were examined by bacteriology and pathology. Lung lesions were evaluated by immunohistochemical (IHC staining for wide spectrum cytokeratin and for M. bovis Vsp antigens and pMB67 antigen. IHC identification and quantitative evaluation of CD4+ and CD8+ T lymphocytes and immunoglobulin (IgG1, IgG2, IgM, IgA-containing plasma cells was performed. Additionally, expression of major histocompatibility complex class II (MHC class II was studied by IHC. Results Suppurative pneumonic lesions were found in all calves. In two calves with caseonecrotic pneumonia, necrotic foci were surrounded by epithelial cells resembling bronchial or bronchiolar epithelium. In all calves, M. bovis Vsp antigens were constantly present in the cytoplasm of macrophages and were also present extracellularly at the periphery of necrotic foci. There was a considerable increase in numbers of IgG1- and IgG2-positive plasma cells among which IgG1-containing plasma cells clearly predominated. Statistical evaluation of the numbers of CD4+ and CD8+ T cells, however, did not reveal statistically significant differences between inoculated and control calves. In M. bovis infected calves, hyperplasia of bronchus-associated lymphoid tissue (BALT was characterized by strong MHC class II expression of lymphoid cells, but only few of the macrophages demarcating the caseonecrotic foci were positive for MHC class II. Conclusions The results from this study show

  8. Proteasomal targeting and minigene repetition improve cell-surface presentation of a transfected, modified melanoma tumour antigen

    DEFF Research Database (Denmark)

    Rasmussen, A B; Zocca, M-B; Bonefeld, C M

    2004-01-01

    Melanoma antigen recognized by T cell 1 (MART-1) is regarded as a candidate peptide for vaccination against malignant melanoma, and it is of importance to develop strategies to improve the vaccine-elicited T-cell activation towards MART-1. T-cell activation is, among other determinants, dependent...

  9. Expression and characterization of an anti-(hepatitis B surface antigen) glycosylated mouse antibody in transgenic tobacco (Nicotiana tabacum) plants and its use in the immunopurification of its target antigen.

    Science.gov (United States)

    Ramírez, Nadia; Rodríguez, Meilyn; Ayala, Marta; Cremata, José; Pérez, Marlene; Martínez, Anazuria; Linares, Marbelis; Hevia, Yasser; Páez, Rolando; Valdés, Rodolfo; Gavilondo, Jorge V; Selman-Housein, Guillermo

    2003-12-01

    Transgenic plants expressing recombinant immunoglobulins have arisen as an alternative technology for the large-scale production of antibodies useful in therapeutics and in industrial processes. In the present paper we report the expression in transgenic tobacco ( Nicotiana tabacum ) of an anti-HBsAg [anti-(hepatitis B virus surface antigen)] mouse IgG1 mAb (monoclonal antibody), currently used for the industrial purification of the recombinant vaccine antigen. Using the sweet potato sporamin signal peptide, a KDEL (Lys-Asp-Glu-Leu) ER (endoplasmic reticulum) anchorage domain, and a heavy- and light-chain gene tandem construction, we generated F1 plants in which the expression of the antibody accounted for 0.5% of the total soluble proteins. The 'plantibody' (functional IgG antibody produced in plants) was easily purified by Protein A-Sepharose chromatography with a yield of approximately 35 microg/g of fresh leaf material, and its glycosylation indicated that, irrespective of the KDEL signal, the molecule is modified in both the ER and Golgi. Finally, a successful comparison of the plantibody with the ascites-derived mAb in the immunoaffinity purification of the vaccine recombinant HBsAg was performed. Taken as a whole, our results show that the large-scale production of this antibody of industrial relevance in transgenic tobacco is feasible.

  10. Facile synthesis of MoS2@Cu2O-Pt nanohybrid as enzyme-mimetic label for the detection of the Hepatitis B surface antigen.

    Science.gov (United States)

    Li, Faying; Li, Yueyun; Feng, Jinhui; Gao, Zengqiang; Lv, Hui; Ren, Xiang; Wei, Qin

    2018-02-15

    An ultrasensitive sandwich-type electrochemical immunosensor was proposed for quantitative detection of hepatitis B surface antigen, which is a representative biomarker of the Hepatitis B virus. First, the porous graphene oxide/Au composites with good conductive ability were employed to accelerate the electron transfer on the electrode interface. Furthermore, the amino functionalized molybdenum disulfide @ cuprous oxide hybrid with coral morphology was prepared to combine platinum nanoparticles for achieving signal amplification strategy. The resulting nanocomposites (molybdenum disulfide @ cuprous oxide - platinum) demonstrated uniform coral morphology, which effectively improved the specific surface area available for loading the secondary antibody and the number of catalytically active sites, even also increased the electrical conductivity. Based on these advantages, this composite system yielded a superior electrocatalytic current response toward the reduction of hydrogen peroxide. In addition, porous graphene oxide/Au composites were used to modify the glassy carbon electrode, thereby presenting a large surface area and becoming biocompatible, for improving the loading capacity of the primary antibody. Under optimal conditions, we obtained a linear relationship between current signal and hepatitis B surface antigen concentration in the broad range from 0.5pg/mL to 200ng/mL, with a detection limit of 0.15pg/mL (signal-to-noise ratio of 3). These values are promising towards clinical applications. Copyright © 2017. Published by Elsevier B.V.

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

    Science.gov (United States)

    Paulus, Aneel; Chitta, Kasyapa S; Wallace, Paul K; Advani, Pooja P; Akhtar, Sharoon; Kuranz-Blake, Maja; Ailawadhi, Sikander; Chanan-Khan, Asher A

    2015-01-01

    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 therapeutic exploit.

  12. The profile of snoRNA-derived microRNAs that regulate expression of variant surface proteins in Giardia lamblia

    Science.gov (United States)

    Li, Wei; Saraiya, Ashesh A.; Wang, Ching C.

    2012-01-01

    Summary In the current investigation, we analyzed all the known small nucleolar RNAs (snoRNAs) in the deeply branching protozoan parasite Giardia lamblia for potential microRNAs (miRNAs) that might be derived from them. Two putative miRNAs have since been identified by Northern blot, primer extension, 3′-RACE and co-immunoprecipitation with Giardia Argonaute (GlAgo), and designated miR6 and miR10. Giardia Dicer (GlDcr) is capable of processing the snoRNAs into the corresponding miRNAs in vitro. Potential miR6 and miR10 binding sites in Giardia genome were predicted bioinformatically. A miR6 binding site was found at the 3′-untranslated regions (UTR) of 44 variant surface protein (vsp) genes, whereas a miR10 binding site was identified at the 3′-end of 159 vsp open-reading frames. Thirty-three of these vsp genes turned out to contain binding sites for both miR6 and miR10. A reporter mRNA tagged with the 3′ end of vsp1267, which contains the target sites for both miRNAs, was translationally repressed by both miRNAs in Giardia. Episomal expression of an N-terminal c-myc tagged VSP1267 was found significantly repressed by introducing either miR6 or miR10 into the cells and the repressive effects were additive. When the 2′-O-methyl antisense oligos (ASOs) of either miR6 or miR10 was introduced, however, there was an enhancement of tagged VSP1267 expression suggesting an inhibition of the repressive effects of endogenous miR6 or miR10 by the ASOs. Of the total 220 vsp genes in Giardia, we have now found 178 of them carrying putative binding sites for all the miRNAs that have been currently identified, suggesting that miRNAs are likely the regulators of VSP expression in Giardia. PMID:22568619

  13. Seroprevalence of hepatitis B e antigen (HBe antigen and B core antibodies (IgG anti-HBcore and IgM anti-HBcore among hepatitis B surface antigen positive blood donors at a Tertiary Centre in Nigeria

    Directory of Open Access Journals (Sweden)

    Akinbami Akinsegun A

    2012-03-01

    Full Text Available Abstract Background Hepatitis B virus (HBV is a common cause of liver disease throughout the world. HBV is transmitted through blood and other body fluids, including semen and saliva. Chronic replication of HBV virons is characterized by persistence circulation of HBsAg, HBeAg and HBV DNA; usually with anti-HBc and occasionally with anti-HBs. Aim: To determine the prevalence of HBeAg, IgG anti-HBcore and IgM anti-HBcore amongst HBsAg positive blood donors. These parameters are reflective of transmissibility and active hepatitis B infection. A cross sectional study was carried out at the blood donor clinics of Lagos State University Teaching Hospital Ikeja and Lagos University Teaching Hospital Idiaraba. A total of 267 donors were recruited to determine HBe antigen, IgG and IgM anti-HBcore antibodies amongst hepatitis BsAg positive donors. Five milliliters of blood was collected from those who tested positive to HBsAg screen during donation. The sera were subjected to enzyme linked immunosorbent assay (ELISA. Pearson chi-squared test was used for the analytical assessment. Findings A total number of 267 HBsAg positive blood donors were studied. A seroprevalence of 8.2% (22 of 267 HBeAg was obtained, 4 of 267 (1.5% were indeterminate while 241 (90.3% tested negative. Only 27 out of 267 donors (10.1% tested positive to IgM anti-HBcore, 234(87.6% tested negative, while 6(2.2% were indeterminate. A higher percentage of 60.7% (162 of 267 tested positive to IgG anti-HBcore, while 39.3% (105 of 267 tested negative. Conclusion There is a low seroprevalence rate of HBeAg-positive chronic hepatitis and relatively high IgG anti-HBcore and IgM anti-HBcore rates in South West Nigeria.

  14. Six host range variants of the xenotropic/polytropic gammaretroviruses define determinants for entry in the XPR1 cell surface receptor

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    Kozak Christine A

    2009-10-01

    Full Text Available Abstract Background The evolutionary interactions between retroviruses and their receptors result in adaptive selection of restriction variants that can allow natural populations to evade retrovirus infection. The mouse xenotropic/polytropic (X/PMV gammaretroviruses rely on the XPR1 cell surface receptor for entry into host cells, and polymorphic variants of this receptor have been identified in different rodent species. Results We screened a panel of X/PMVs for infectivity on rodent cells carrying 6 different XPR1 receptor variants. The X/PMVs included 5 well-characterized laboratory and wild mouse virus isolates as well as a novel cytopathic XMV-related virus, termed Cz524, isolated from an Eastern European wild mouse-derived strain, and XMRV, a xenotropic-like virus isolated from human prostate cancer. The 7 viruses define 6 distinct tropisms. Cz524 and another wild mouse isolate, CasE#1, have unique species tropisms. Among the PMVs, one Friend isolate is restricted by rat cells. Among the XMVs, two isolates, XMRV and AKR6, differ from other XMVs in their PMV-like restriction in hamster cells. We generated a set of Xpr1 mutants and chimeras, and identified critical amino acids in two extracellular loops (ECLs that mediate entry of these different viruses, including 3 residues in ECL3 that are involved in PMV entry (E500, T507, and V508 and can also influence infectivity by AKR6 and Cz524. Conclusion We used a set of natural variants and mutants of Xpr1 to define 6 distinct host range variants among naturally occurring X/PMVs (2 XMV variants, 2 PMVs, 2 different wild mouse variants. We identified critical amino acids in XPR1 that mediate entry of these viruses. These gammaretroviruses and their XPR1 receptor are thus highly functionally polymorphic, a consequence of the evolutionary pressures that favor both host resistance and virus escape mutants. This variation accounts for multiple naturally occurring virus resistance phenotypes and

  15. The O-Antigen Capsule of Salmonella enterica Serovar Typhimurium Facilitates Serum Resistance and Surface Expression of FliC

    OpenAIRE

    Marshall, Joanna M.; Gunn, John S.

    2015-01-01

    Group IV polysaccharide capsules are common in enteric bacteria and have more recently been described in nontyphoidal Salmonella species. Such capsules are known as O-antigen (O-Ag) capsules, due to their high degree of similarity to the O-Ag of the lipopolysaccharide (LPSO-Ag). Capsular polysaccharides are known virulence factors of many bacterial pathogens, facilitating evasion of immune recognition and systemic dissemination within the host. Previous studies on the O-Ag capsule of salmonel...

  16. Energy filtering transmission electron microscopy immunocytochemistry and antigen retrieval of surface layer proteins from Tannerella forsythensis using microwave or autoclave heating with citraconic anhydride.

    Science.gov (United States)

    Moriguchi, K; Mitamura, Y; Iwami, J; Hasegawa, Y; Higuchi, N; Murakami, Y; Maeda, H; Yoshimura, F; Nakamura, H; Ohno, N

    2012-11-01

    Tannerella forsythensis (Bacteroides forsythus), an anaerobic Gram-negative species of bacteria that plays a role in the progression of periodontal disease, has a unique bacterial protein profile. It is characterized by two unique protein bands with molecular weights of more than 200 kDa. It also is known to have a typical surface layer (S-layer) consisting of regularly arrayed subunits outside the outer membrane. We examined the relationship between high molecular weight proteins and the S-layer using electron microscopic immunolabeling with chemical fixation and an antigen retrieval procedure consisting of heating in a microwave oven or autoclave with citraconic anhydride. Immunogold particles were localized clearly at the outermost cell surface. We also used energy-filtering transmission electron microscopy (EFTEM) to visualize 3, 3'-diaminobenzidine tetrahydrochloride (DAB) reaction products after microwave antigen retrieval with 1% citraconic anhydride. The three-window method for electron spectroscopic images (ESI) of nitrogen by the EFTEM reflected the presence of moieties demonstrated by the DAB reaction with horseradish peroxidase (HRP)-conjugated secondary antibodies instead of immunogold particles. The mapping patterns of net nitrogen were restricted to the outermost cell surface.

  17. Antigenic variation in the intestinal parasite Giardia lamblia.

    Science.gov (United States)

    Gargantini, Pablo Rubén; Serradell, Marianela Del Carmen; Ríos, Diego Nicolás; Tenaglia, Albano Heraldo; Luján, Hugo Daniel

    2016-08-01

    Giardia lamblia trophozoites undergo antigenic variation, where one member of the Variant-specific Surface Protein (VSP) family is expressed on the surface of proliferating trophozoites and periodically replaced by another one. Two main questions have challenged researchers since antigenic switching was discovered in Giardia: What are the mechanisms involved? How are they influenced by other cellular processes or by the environment? Two molecular mechanisms have been proposed, both involving small non-coding RNAs. Here we postulate that (a) chromatin remodeling, triggered by environmental factors, also plays an important role in selecting the VSP that will be expressed and (b) the particular VSP structure may not only protect the parasite in the small intestine but also signal the need to exchange the existing VSP for another. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Computer simulation of histo-blood group oligosaccharides: energy maps of all constituting disaccharides and potential energy surfaces of 14 ABH and Lewis carbohydrate antigens.

    Science.gov (United States)

    Imberty, A; Mikros, E; Koca, J; Mollicone, R; Oriol, R; Pérez, S

    1995-06-01

    The three-dimensional structures of fourteen histo-blood groups carbohydrate antigens have been established through a combination of molecular mechanics and conformational searching methods. The conformational space available for each disaccharide, constituents of these determinants, has been throroughly characterized. The results have been organized in a data bank fashion. Larger relatives, i.e. 14 tri- and tetrasaccharides of histo-blood group antigens, have been modelled using a different method for exploring the complex potential energy surface. This approach is aimed at establishing all the possible families of conformations, along with the conformational pathways. Different conformational behaviours are exhibited by these oligosaccharides. Some of them, i.e. Le(x) and Le(y) tri and tetrasaccharides, are very rigid; 99% of their populations belong to the same conformational family. Others, like H type 1, H type 2 or H type 6 oligosaccharides, are essentially rigid, but a secondary conformational family, corresponding to 3-4% of the total population, can arise. Finally, the H types 3 and 4 trisaccharides, and the A type 1 and A type 2 tetrasaccharides are predicted to behave rather flexibly. The information gathered in the present investigation has been used to analyse the body of experimental evidence, either physical or biological, available for this series of carbohydrate antigens. Of special interest are the several different alignments that can be proposed for these molecules. They yield a realistic definition of the three-dimensional features of the epitopes thereby providing essential information about how carbohydrate antigens are recognized by proteins.

  19. Removing N-terminal sequences in pre-S1 domain enhanced antibody and B-cell responses by an HBV large surface antigen DNA vaccine.

    Directory of Open Access Journals (Sweden)

    Guohong Ge

    Full Text Available Although the use of recombinant hepatitis B virus surface (HBsAg protein vaccine has successfully reduced global hepatitis B infection, there are still a number of vaccine recipients who do not develop detectable antibody responses. Various novel vaccination approaches, including DNA vaccines, have been used to further improve the coverage of vaccine protection. Our previous studies demonstrated that HBsAg-based DNA vaccines could induce both humoral and CMI responses in experimental animal models. However, one form of the the HBsAg antigen, the large S antigen (HBs-L, expressed by DNA vaccine, was not sufficiently immunogenic in eliciting antibody responses. In the current study, we produced a modified large S antigen DNA vaccine, HBs-L(T, which has a truncated N-terminal sequence in the pre-S1 region. Compared to the original HBs-L DNA vaccine, the HBs-L(T DNA vaccine improved secretion in cultured mammalian cells and generated significantly enhanced HBsAg-specific antibody and B cell responses. Furthermore, this improved HBsL DNA vaccine, along with other HBsAg-expressing DNA vaccines, was able to maintain predominantly Th1 type antibody responses while recombinant HBsAg protein vaccines produced in either yeast or CHO cells elicited mostly Th2 type antibody responses. Our data indicate that HBsAg DNA vaccines with improved immunogenicity offer a useful alternative choice to recombinant protein-based HBV vaccines, particularly for therapeutic purposes against chronic hepatitis infection where immune tolerance led to poor antibody responses to S antigens.

  20. Flow cytometric measurement of RNA synthesis based on bromouridine labelling and combined with measurement of DNA content or cell surface antigen

    DEFF Research Database (Denmark)

    Jensen, P O; Larsen, J; Larsen, J K

    1993-01-01

    RNA synthesis can be analysed in nuclei or cells labelled with 5-bromouridine (BrUrd) and stained using cross-reacting anti-bromodeoxyuridine (BrdUrd) antibody. Flow cytometric dual parameter analysis of BrUrd incorporation and DNA content in nuclear suspensions of human blood lymphocytes showed...... that RNA synthesis increased within the first 24 hours of phytohemagglutinin (PHA) stimulation, reaching a maximum at 48 hours, when cells had entered the cell cycle. Using a new method for flow cytometric dual parameter analysis of BrUrd incorporation and a cell surface antigen, spontaneous RNA synthesis...

  1. Novel atomic force microscopy based biopanning for isolation of morphology specific reagents against TDP-43 variants in amyotrophic lateral sclerosis.

    Science.gov (United States)

    Williams, Stephanie M; Venkataraman, Lalitha; Tian, Huilai; Khan, Galam; Harris, Brent T; Sierks, Michael R

    2015-02-12

    Because protein variants play critical roles in many diseases including TDP-43 in Amyotrophic Lateral Sclerosis (ALS), alpha-synuclein in Parkinson's disease and beta-amyloid and tau in Alzheimer's disease, it is critically important to develop morphology specific reagents that can selectively target these disease-specific protein variants to study the role of these variants in disease pathology and for potential diagnostic and therapeutic applications. We have developed novel atomic force microscopy (AFM) based biopanning techniques that enable isolation of reagents that selectively recognize disease-specific protein variants. There are two key phases involved in the process, the negative and positive panning phases. During the negative panning phase, phages that are reactive to off-target antigens are eliminated through multiple rounds of subtractive panning utilizing a series of carefully selected off-target antigens. A key feature in the negative panning phase is utilizing AFM imaging to monitor the process and confirm that all undesired phage particles are removed. For the positive panning phase, the target antigen of interest is fixed on a mica surface and bound phages are eluted and screened to identify phages that selectively bind the target antigen. The target protein variant does not need to be purified providing the appropriate negative panning controls have been used. Even target protein variants that are only present at very low concentrations in complex biological material can be utilized in the positive panning step. Through application of this technology, we acquired antibodies to protein variants of TDP-43 that are selectively found in human ALS brain tissue. We expect that this protocol should be applicable to generating reagents that selectively bind protein variants present in a wide variety of different biological processes and diseases.

  2. Phage display used for gene cloning of human recombinant antibody against the erythrocyte surface antigen, rhesus D

    DEFF Research Database (Denmark)

    Dziegiel, M; Nielsen, L K; Andersen, P S

    1995-01-01

    A novel phage display system has been developed for PCR amplification and cloning of the Fab fragments of human immunoglobulin genes. Using this system, we have cloned an antibody from a mouse-human hybridoma cell line directed against the erythrocyte antigen rhesus D. Intact erythrocytes were used...... for absorption of the Fab phages. Soluble Fab fragments produced from the cloned material showed identical performance to the parental antibody in agglutination assays. Gel filtration confirmed that the Fab fragment consists of a kappa-Fd heterodimer. The successful use of intact cells for selection of specific...

  3. HD-03/ES: A Herbal Medicine Inhibits Hepatitis B Surface Antigen Secretion in Transfected Human Hepatocarcinoma PLC/PRF/5 Cells

    Directory of Open Access Journals (Sweden)

    Sandeep R. Varma

    2013-01-01

    Full Text Available HD-03/ES is a herbal formulation used for the treatment of hepatitis B. However, the molecular mechanism involved in the antihepatitis B (HBV activity of this drug has not been studied using in vitro models. The effect of HD-03/ES on hepatitis B surface antigen (HBsAg secretion and its gene expression was studied in transfected human hepatocarcinoma PLC/PRF/5 cells. The anti-HBV activity was tested based on the inhibition of HBsAg secretion into the culture media, as detected by HBsAg-specific antibody-mediated enzyme assay (ELISA at concentrations ranging from 125 to 1000 μg/mL. The effect of HD-03/ES on HBsAg gene expression was analyzed using semiquantitative multiplex RT-PCR by employing specific primers. The results showed that HD-03/ES suppressed HBsAg production with an IC50 of 380 μg/mL in PLC/PRF/5 cells for a period of 24 h. HD-03/ES downregulated HBsAg gene expression in PLC/PRF/5 cells. In conclusion, HD-03/ES exhibits strong anti-HBV properties by inhibiting the secretion of hepatitis B surface antigen in PLC/PRF/5 cells, and this action is targeted at the transcription level. Thus, HD-03/ES could be beneficial in the treatment of acute and chronic hepatitis B infections.

  4. HD-03/ES: A Herbal Medicine Inhibits Hepatitis B Surface Antigen Secretion in Transfected Human Hepatocarcinoma PLC/PRF/5 Cells.

    Science.gov (United States)

    Varma, Sandeep R; Sundaram, R; Gopumadhavan, S; Vidyashankar, Satyakumar; Patki, Pralhad S

    2013-01-01

    HD-03/ES is a herbal formulation used for the treatment of hepatitis B. However, the molecular mechanism involved in the antihepatitis B (HBV) activity of this drug has not been studied using in vitro models. The effect of HD-03/ES on hepatitis B surface antigen (HBsAg) secretion and its gene expression was studied in transfected human hepatocarcinoma PLC/PRF/5 cells. The anti-HBV activity was tested based on the inhibition of HBsAg secretion into the culture media, as detected by HBsAg-specific antibody-mediated enzyme assay (ELISA) at concentrations ranging from 125 to 1000  μ g/mL. The effect of HD-03/ES on HBsAg gene expression was analyzed using semiquantitative multiplex RT-PCR by employing specific primers. The results showed that HD-03/ES suppressed HBsAg production with an IC50 of 380  μ g/mL in PLC/PRF/5 cells for a period of 24 h. HD-03/ES downregulated HBsAg gene expression in PLC/PRF/5 cells. In conclusion, HD-03/ES exhibits strong anti-HBV properties by inhibiting the secretion of hepatitis B surface antigen in PLC/PRF/5 cells, and this action is targeted at the transcription level. Thus, HD-03/ES could be beneficial in the treatment of acute and chronic hepatitis B infections.

  5. Seroprevalence of Hepatitis B virus surface antigen and factors associated among pregnant women in Dawuro zone, SNNPR, Southwest Ethiopia: a cross sectional study.

    Science.gov (United States)

    Chernet, Asrat; Yesuf, Aman; Alagaw, Amsalu

    2017-08-22

    Hepatitis B is the world's most common serious liver infection. Infection by hepatitis B virus during pregnancy has high rate of vertical transmission and adverse effect on both the mother and child. Data on seroprevalence and associated factors among pregnant women in Dawuro or surrounding was scarce. Therefore, the aim of this study was to assess prevalence of hepatitis B virus and factors associated among pregnant women in Dawuro Zone. Institution based cross-sectional study was conducted from 1st April to May 31, 2015 in Dawuro zone. Venous blood was collected, plasma was separated and hepatitis B virus surface antigen was screened using rapid test. Logistic regression was employed with 95% CI and p value of women. The overall prevalence from this study was (p = 3.5%, 95% CI 1.4-5.6%) multivariate analysis using logistic regression showed multiple sexual partner (AOR = 6.923; 95% CI 1.685-28.441), and abortion history (AOR = 4.975; 95% CI 1.21-20.456), were significantly associated with hepatitis B virus surface antigen (HBsAg) infection. This seroprevalence was categorized as intermediate endemicity according to WHO classification criteria. Health education on sexual transmission of hepatitis B virus and inclusion of screening hepatitis b virus as routine antenatal care service is recommended.

  6. Isolation of recombinant Hepatitis B surface antigen with antibody-conjugated superparamagnetic Fe3O4/SiO2 core-shell nanoparticles.

    Science.gov (United States)

    Mostafaei, Mehdi; Hosseini, Seyed Nezamedin; Khatami, Maryam; Javidanbardan, Amin; Sepahy, Abbas Akhavan; Asadi, Ebadullah

    2017-12-12

    In the production process of recombinant Hepatitis B surface antigen (rHBsAg) various separation techniques are used to purify this virus-like particle (VLP). In this study, we developed antibody-conjugated super-paramagnetic Fe3O4/SiO2 core-shell nanoparticles as a highly selective method for isolation of expressed rHBsAg in yeast Pichia pastoris. For this purpose, first, iron oxide magnetic nanoparticles (MNPs) were prepared by co-precipitation method in alkali media and coated with silica. Then the surface was activated by amine groups and conjugated with oxidized antibodies. X-ray diffraction (XRD), transmission electron microscopy (TEM), and vibrating sample magnetometer (VSM) were used to study the physical properties of MNPs. To evaluate the efficacy of these MNPs as a purification technique successfully synthesized MNPs were added to the rHBsAg sample to couple with the antigen and then be isolated based on their magnetic property. In the present research, in the optimum condition, we could isolate 65% of total rHBsAg from the final vaccine sample with purity above 95%. In this procedure, the maximum obtained specific yield (mg HBsAg/mg MNPs) was equal to 37.6. These results underline the potential application of the immune-magnetic separation (IMS) in the future bioseparation systems. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. A Sheep in Wolf's Clothing: Listeria innocua Strains with Teichoic Acid-Associated Surface Antigens and Genes Characteristic of Listeria monocytogenes Serogroup 4

    Science.gov (United States)

    Lan, Zheng; Fiedler, Franz; Kathariou, Sophia

    2000-01-01

    Listeria monocytogenes serotype 4b has been implicated in numerous food-borne epidemics and in a substantial fraction of sporadic listeriosis. A unique lineage of the nonpathogenic species Listeria innocua was found to express teichoic acid-associated surface antigens that were otherwise expressed only by L. monocytogenes of serotype 4b and the rare serotypes 4d and 4e. These L. innocua strains were also found to harbor sequences homologous to the gene gtcA, which has been shown to be essential for teichoic acid glycosylation in L. monocytogenes serotype 4b. Transposon mutagenesis and genetic studies revealed that the gtcA gene identified in this lineage of L. innocua was functional in serotype 4b-like glycosylation of the teichoic acids of these organisms. The genomic organization of the gtcA region was conserved between this lineage of L. innocua and L. monocytogenes serotype 4b. Our data are in agreement with the hypothesis that, in this lineage of L. innocua, gtcA was acquired by lateral transfer from L. monocytogenes serogroup 4. The high degree of nucleotide sequence conservation in the gtcA sequences suggests that such transfer was relatively recent. Transfer events of this type may alter the surface antigenic properties of L. innocua and may eventually lead to evolution of novel pathogenic lineages through additional acquisition of genes from virulent listeriae. PMID:11029438

  8. Human microRNA hsa-miR-125a-5p interferes with expression of hepatitis B virus surface antigen

    Science.gov (United States)

    Potenza, Nicoletta; Papa, Umberto; Mosca, Nicola; Zerbini, Francesca; Nobile, Valentina; Russo, Aniello

    2011-01-01

    MicroRNAs are small non-coding RNAs that modulate gene expression at post-transcriptional level, playing a crucial role in cell differentiation and development. Recently, some reports have shown that a limited number of mammalian microRNAs are also involved in anti-viral defense. In this study, the analysis of the hepatitis B virus (HBV) genome by the computer program MiRanda led to the identification of seven sites that are potential targets for human liver microRNAs. These sites were found to be clustered in a 995-bp segment within the viral polymerase ORF and the overlapping surface antigen ORF, and conserved among the most common HBV subtypes. The HBV genomic targets were then subjected to a validation test based on cultured hepatic cells (HepG2, HuH-7 and PLC/PRF/5) and luciferase reporter genes. In this test, one of the selected microRNAs, hsa-miR-125a-5p, was found to interact with the viral sequence and to suppress the reporter activity markedly. The microRNA was then shown to interfere with the viral translation, down-regulating the expression of the surface antigen. Overall, these results support the emerging concept that some mammalian microRNAs play a role in virus-host interaction. Furthermore, they provide the basis for the development of new strategies for anti-HBV intervention. PMID:21317190

  9. Glycosyl-sn-1,2-dimyristylphosphatidylinositol is the membrane anchor for Trypanosoma equiperdum and T. (Nannomonas) congolense variant surface glycoproteins.

    Science.gov (United States)

    Lamont, G S; Fox, J A; Cross, G A

    1987-06-01

    We have analysed the structures of the Trypanosoma (Nannomonas) congolense and T. equiperdum variant surface glycoprotein (VSG) membrane anchors. Myristic acid uptake, phospholipase treatment, and nitrous acid deamination showed that, for each species, the anchor is glycosyl-sn-1,2-dimyristylphosphatidylinositol, as has been previously described for T. brucei. Osmotic lysis of these trypanosomes resulted in the release of soluble VSG, lacking fatty acid. In both species and in T. evansi, an endogenous phospholipase C, which cleaved diacylglycerol from membrane form VSG, was identified.

  10. Three-Dimensional Structure and Biophysical Characterization of Staphylococcus aureus Cell Surface Antigen-Manganese Transporter MntC

    Energy Technology Data Exchange (ETDEWEB)

    Gribenko, Alexey; Mosyak, Lidia; Ghosh, Sharmistha; Parris, Kevin; Svenson, Kristine; Moran, Justin; Chu, Ling; Li, Sheng; Liu, Tong; Woods, Jr., Virgil L.; Jansen, Kathrin U.; Green, Bruce A.; Anderson, Annaliesa S.; Matsuka, Yury V. [Pfizer; (UCSD)

    2013-08-23

    MntC is a metal-binding protein component of the Mn2 +-specific mntABC transporter from the pathogen Staphylococcus aureus. The protein is expressed during the early stages of infection and was proven to be effective at reducing both S. aureus and Staphylococcus epidermidis infections in a murine animal model when used as a vaccine antigen. MntC is currently being tested in human clinical trials as a component of a multiantigen vaccine for the prevention of S. aureus infections. To better understand the biological function of MntC, we are providing structural and biophysical characterization of the protein in this work. The three-dimensional structure of the protein was solved by X-ray crystallography at 2.2 Å resolution and suggests two potential metal binding modes, which may lead to reversible as well as irreversible metal binding. Precise Mn2 +-binding affinity of the protein was determined from the isothermal titration calorimetry experiments using a competition approach. Differential scanning calorimetry experiments confirmed that divalent metals can indeed bind to MntC reversibly as well as irreversibly. Finally, Mn2 +-induced structural and dynamics changes have been characterized using spectroscopic methods and deuterium–hydrogen exchange mass spectroscopy. Results of the experiments show that these changes are minimal and are largely restricted to the structural elements involved in metal coordination. Therefore, it is unlikely that antibody binding to this antigen will be affected by the occupancy of the metal-binding site by Mn2 +.

  11. Quantification of large and middle proteins of hepatitis B virus surface antigen (HBsAg) as a novel tool for the identification of inactive HBV carriers.

    Science.gov (United States)

    Pfefferkorn, Maria; Böhm, Stephan; Schott, Tina; Deichsel, Danilo; Bremer, Corinna M; Schröder, Kathrin; Gerlich, Wolfram H; Glebe, Dieter; Berg, Thomas; van Bömmel, Florian

    2017-09-26

    Among individuals with chronic hepatitis B, those with hepatitis B e-antigen (HBeAg)-negative chronic hepatitis (CHB) can be difficult to distinguish from those with HBeAg-negative chronic HBV infection, also referred to as inactive HBV carriers (ICs), but both require different medical management. The level of HBV surface antigen (HBsAg) has been proposed as a marker to discriminate between chronic infection and hepatitis stages. HBsAg consists of large, middle and small HBs. The aim of this study was to determine whether the composition of HBsAg improved the identification of ICs among HBsAg-positive subjects with different phases of HBV infections. HBV large surface proteins (LHBs) and HBV middle surface proteins (MHBs) were quantified in serum samples from 183 clinically well-characterised untreated patients with acute (n=14) HBV infection, ICs (n=44), CHBs (n=46), chronic HBeAg-positive phase (n=68) and hepatitis delta coinfection (n=11) using an ELISA, with well-defined monoclonal antibodies against the preS1 domain (LHBs) and the preS2-domain (MHBs). A Western blot analysis was used to verify the quantitation of the components of HBsAg. Total HBsAg was quantified using a modified commercially available assay (HBsAg V.6.0, Enzygnost, Siemens, Erlangen). The composition of HBsAg showed specific patterns across different phases of hepatitis B. Individuals in the IC phase had significantly lower proportions of LHBs and MHBs than patients in acute or chronic phases irrespective of their HBV e-antigen status (p<0.0001) or HBsAg level. Both LHBs and MHBs ratios better predicted the IC phase than total HBsAg levels. Quantification of MHBs, particularly LHBs represents a novel tool for the identification of the IC stage. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  12. A Novel Platform for the Potentiation of Therapeutic Antibodies Based on Antigen-Dependent Formation of IgG Hexamers at the Cell Surface.

    Directory of Open Access Journals (Sweden)

    Rob N de Jong

    2016-01-01

    Full Text Available IgG antibodies can organize into ordered hexamers on cell surfaces after binding their antigen. These hexamers bind the first component of complement C1 inducing complement-dependent target cell killing. Here, we translated this natural concept into a novel technology platform (HexaBody technology for therapeutic antibody potentiation. We identified mutations that enhanced hexamer formation and complement activation by IgG1 antibodies against a range of targets on cells from hematological and solid tumor indications. IgG1 backbones with preferred mutations E345K or E430G conveyed a strong ability to induce conditional complement-dependent cytotoxicity (CDC of cell lines and chronic lymphocytic leukemia (CLL patient tumor cells, while retaining regular pharmacokinetics and biopharmaceutical developability. Both mutations potently enhanced CDC- and antibody-dependent cellular cytotoxicity (ADCC of a type II CD20 antibody that was ineffective in complement activation, while retaining its ability to induce apoptosis. The identified IgG1 Fc backbones provide a novel platform for the generation of therapeutics with enhanced effector functions that only become activated upon binding to target cell-expressed antigen.

  13. Hepatitis B virus-associated diffuse large B-cell lymphoma: unique clinical features, poor outcome, and hepatitis B surface antigen-driven origin.

    Science.gov (United States)

    Deng, Lijuan; Song, Yuqin; Young, Ken H; Hu, Shimin; Ding, Ning; Song, Weiwei; Li, Xianghong; Shi, Yunfei; Huang, Huiying; Liu, Weiping; Zheng, Wen; Wang, Xiaopei; Xie, Yan; Lin, Ningjing; Tu, Meifeng; Ping, Lingyan; Ying, Zhitao; Zhang, Chen; Sun, Yingli; Zhu, Jun

    2015-09-22

    While the epidemiologic association between hepatitis B virus (HBV) infection and diffuse large B-cell lymphoma (DLBCL) is established, little is known more than this epidemiologic evidence. We studied a cohort of 587 patients with DLBCL for HBV infection status, clinicopathologic features, and the immunoglobulin variable region in HBV surface antigen (HBsAg)-positive patients. Eighty-one (81/587, 13.8%) patients were HBsAg-positive. Compared with HBsAg-negative DLBCL, HBsAg-positive DLBCL displayed a younger median onset age (45 vs. 55 years), more frequent involvement of spleen or retroperitoneal lymph node (40.7% vs. 16.0% and 61.7% vs. 31.0% respectively, both p < 0.001), more advanced disease (stage III/IV: 76.5% vs 59.5%, p = 0.003), and significantly worse outcome (2-year overall survival: 47% versus 70%, p < 0.001). In HBsAg-positive DLBCL patients, almost all (45/47, 96%) amino acid sequences of heavy and light chain complementarity determining region 3 exhibited a high homology to antibodies specific for HBsAg, and the majority (45/50, 90%) of IgHV and IgLV genes were mutated. We conclude that 13.8% of DLBCL cases are HBV-associated in HBV-endemic China and show unique clinical features and poor outcomes. Furthermore, our study strongly suggests that HBV-associated DLBCL might arise from HBV antigen-selected B cells.

  14. Distinct domains of the CD3-gamma chain are involved in surface expression and function of the T cell antigen receptor

    DEFF Research Database (Denmark)

    Wegener, A M; Hou, X; Dietrich, J

    1995-01-01

    The T cell antigen receptor (TcR) is a multisubunit complex that consists of at least six different polypeptides. We have recently demonstrated that the CD3-delta subunit cannot substitute for the CD3-gamma subunit in TcR cell surface expression, in spite of significant amino acid homology between...... these two subunits. To identify CD3-gamma-specific domains that are required for assembly of the complete TcR and for surface expression and function of the TcR, chimeric CD3-gamma/CD3-delta molecules were constructed and expressed in T cells devoid of endogenous CD3-gamma. Substitution of the extracellular...... domain of CD3-gamma with that of CD3-delta did not allow cell surface expression of the TcR. In contrast, substitution of the transmembrane and/or the intracellular domains of CD3-gamma with those of CD3-delta did allow TcR cell surface expression. These results conclusively demonstrate...

  15. The Leishmania promastigote surface antigen-2 (PSA-2) is specifically recognised by Th1 cells in humans with naturally acquired immunity to L. major

    DEFF Research Database (Denmark)

    Kemp, M; Handman, E; Kemp, K

    1998-01-01

    The promastigote surface antigen-2 (PSA-2) is a Leishmania parasite antigen, which can induce Th1-mediated protection against murine leishmaniasis when used as a vaccine. To evaluate PSA-2 as a human vaccine candidate the specific T-cell response to PSA-2 was characterised in individuals immune...... to cutaneous leishmaniasis. Peripheral blood mononuclear cells from Sudanese individuals with a past history of self-healing cutaneous leishmaniasis proliferated vigorously in response to PSA-2 isolated from Leishmania major, whereas the antigen did not activate cells from presumably unexposed Danes...... and that these cells were the major source of interferon-gamma. The results show that Th1-like cells recognising PSA-2 are expanded during infection by L. major and that they maintain their Th1-like cytokine profile upon reactivation in vitro. Since immunity to cutaneous leishmaniasis is mediated by antigen...

  16. Sero-prevalence of hepatitis B surface antigen amongst pregnant women attending an antenatal clinic, Volta region, Ghana

    Directory of Open Access Journals (Sweden)

    Arnold Luuse

    2017-01-01

    Full Text Available Hepatitis B virus (HBV infection remains a global challenge, although there is currently a safe and effective vaccine available. HBV prevalence in Ghana is not well documented, but vary regionally from 4.8% to 12.3% in the general population, 10.8% to 12.7% in blood donors and about 10.6% in pregnant women. This puts Ghana among the high endemic countries in Africa. The study objective was to determine the sero-prevalence of HBs antigen (Ag and HBeAg among pregnant women in the Ho municipality. Two hundred and eight participants (pregnant women, attending Ho Municipal antenatal clinic were enrolled into the study. This study recorded a HBsAg seroprevalence rate of 2.4% among the pregnant women, with primigravida pregnant women recording (0.98% and multigravida (1.42%. The prevalence of HBsAg among the pregnant women can be classified as Low Intermediate; therefore there is still the need for routine screening of pregnant women during antenatal visits. Amongst HBsAg positives, HBeAg positivity was significantly high (40% of all HBsAg positive women, which suggests high chances of carrier and vertical transmission (mother to child state.

  17. Sero-Prevalence of Hepatitis B Surface Antigen Amongst Pregnant Women Attending an Antenatal Clinic, Volta Region, Ghana.

    Science.gov (United States)

    Luuse, Arnold; Dassah, Sylvester; Lokpo, Sylvester; Ameke, Louise; Noagbe, Mark; Adatara, Peter; Hagan, Oheneba; Binka, Fred

    2016-12-31

    Hepatitis B virus (HBV) infection remains a global challenge, although there is currently a safe and effective vaccine available. HBV prevalence in Ghana is not well documented, but vary regionally from 4.8% to 12.3% in the general population, 10.8% to 12.7% in blood donors and about 10.6% in pregnant women. This puts Ghana among the high endemic countries in Africa. The study objective was to determine the sero-prevalence of HBs antigen (Ag) and HBeAg among pregnant women in the Ho municipality. Two hundred and eigh participants (pregnant women), attending Ho Municipal antenatal clinic were enrolled into the study. This study recorded a HBsAg sero-prevalence rate of 2.4% among the pregnant women, with primigravida pregnant women recording (0.98%) and multigravida (1.42%). The prevalence of HBsAg among the pregnant women can be classified as Low Intermediate; therefore there is still the need for routine screening of pregnant women during antenatal visits. Amongst HBsAg positives, HBeAg positivity was significantly high (40% of all HBsAg positive women), which suggests high chances of carrier and vertical transmission (mother to child) state.

  18. Stable overproducer of hepatitis B surface antigen in the methylotrophic yeast Hansenula polymorpha due to multiple integration of heterologous auxotrophic selective markers and defect in peroxisome biogenesis.

    Science.gov (United States)

    Krasovska, Olena S; Stasyk, Oleh V; Sibirny, Andriy A

    2013-12-01

    Two methods of multicopy integrant selection in the methylotrophic yeast Hansenula polymorpha based on the use of heterologous yeast auxotrophic genes have been used to isolate effective overproducers of hepatitis B surface antigen (HBsAg). One selection marker was described earlier for this yeast, the Saccharomyces cerevisiae URA3 gene, whereas the second selection marker was developed by us, the Pichia pastoris ADE1 gene with shortened native promoter. Sequential use of both selection markers produced stable transformants containing up to 30 integration cassettes with HBsAg gene. Deletion of PEX3 gene coding for peroxine involved in the early step of peroxisome formation substantially increased the production of HBsAg in glucose medium as compared to the parental strain. Maximal production of HBsAg in Δpex3 strain was nearly 8-9% of the total cell protein.

  19. A Mutant Library Approach to Identify Improved Meningococcal Factor H Binding Protein Vaccine Antigens.

    Directory of Open Access Journals (Sweden)

    Monica Konar

    Full Text Available Factor H binding protein (FHbp is a virulence factor used by meningococci to evade the host complement system. FHbp elicits bactericidal antibodies in humans and is part of two recently licensed vaccines. Using human complement Factor H (FH transgenic mice, we previously showed that binding of FH decreased the protective antibody responses to FHbp vaccination. Therefore, in the present study we devised a library-based method to identify mutant FHbp antigens with very low binding of FH. Using an FHbp sequence variant in one of the two licensed vaccines, we displayed an error-prone PCR mutant FHbp library on the surface of Escherichia coli. We used fluorescence-activated cell sorting to isolate FHbp mutants with very low binding of human FH and preserved binding of control anti-FHbp monoclonal antibodies. We sequenced the gene encoding FHbp from selected clones and introduced the mutations into a soluble FHbp construct. Using this approach, we identified several new mutant FHbp vaccine antigens that had very low binding of FH as measured by ELISA and surface plasmon resonance. The new mutant FHbp antigens elicited protective antibody responses in human FH transgenic mice that were up to 20-fold higher than those elicited by the wild-type FHbp antigen. This approach offers the potential to discover mutant antigens that might not be predictable even with protein structural information and potentially can be applied to other microbial vaccine antigens that bind host proteins.

  20. In vitro selection of Plasmodium falciparum 3D7 for expression of variant surface antigens associated with severe malaria in African children

    DEFF Research Database (Denmark)

    Staalsoe, Trine; Nielsen, Morten A; Vestergaard, Lasse S

    2003-01-01

    . Parasites isolated from young children with severe malaria (SM) tend to express a limited and conserved set of VSA (VSASM) that are both stronger and more commonly recognized by IgG in the plasma of malaria-exposed individuals than VSA (VSAUM) expressed by parasites causing uncomplicated malaria (UM...... and epidemiologically diverse areas of endemic parasite transmission. The described selection method appears a useful tool in the identification of genes encoding VSA involved in severe and life-threatening P. falciparum malaria....

  1. Maternally transmitted antibodies to pregnancy-associated variant antigens on the surface of erythrocytes infected with Plasmodium falciparum: relation to child susceptibility to malaria

    DEFF Research Database (Denmark)

    Cot, Michel; Le Hesran, Jean Yves; Staalsoe, Trine

    2003-01-01

    The consequences of pregnancy-associated malaria on a child's health have been poorly investigated. Malarial infection of the placenta seems to result in a higher susceptibility of children to the parasite during their first year of life. In 1993-1995, the authors investigated the role of antibod...

  2. The immuno-epidemiology of pregnancy-associated Plasmodium falciparum malaria: a variant surface antigen-specific perspective

    DEFF Research Database (Denmark)

    Hviid, L

    2004-01-01

    Women living in areas of intense P. falciparum transmission have acquired substantial protective immunity to malaria when they reach childbearing age. Nevertheless, pregnancies in such areas are associated with substantial malaria-related morbidity and mortality, particularly among women of low p...... understanding of how protective immunity to P. falciparum malaria operates and is acquired, have provided important insights into this enigma....

  3. Human monoclonal IgG selection of Plasmodium falciparum for the expression of placental malaria-specific variant surface antigens

    DEFF Research Database (Denmark)

    Soerli, J; Barfod, L; Lavstsen, T

    2009-01-01

    Pregnancy-associated Plasmodium falciparum malaria (PAM) is a major cause of morbidity and mortality in African women and their offspring. PAM is characterized by accumulation of infected erythrocytes (IEs) that adhere to chondroitin sulphate A (CSA) in the placental intervillous space. We show...

  4. Acquisition and decay of antibodies to pregnancy-associated variant antigens on the surface of Plasmodium falciparum-infected erythrocytes that protect against placental parasitemia

    DEFF Research Database (Denmark)

    Staalsoe, T; Megnekou, R; Fievét, N

    2001-01-01

    Otherwise clinically immune women in areas endemic for malaria are highly susceptible to Plasmodium falciparum malaria during their first pregnancy. Pregnancy-associated malaria (PAM) is characterized by placental accumulation of infected erythrocytes that adhere to chondroitin sulfate A (CSA). S...

  5. The Trypomastigote Small Surface Antigen from Trypanosoma cruzi improves treatment evaluation and diagnosis in pediatric Chagas disease.

    Science.gov (United States)

    Balouz, Virginia; Melli, Luciano J; Volcovich, Romina; Moscatelli, Guillermo; Moroni, Samanta; González, Nicolás; Ballering, Griselda; Bisio, Margarita; Ciocchini, Andrés; Buscaglia, Carlos A; Altcheh, Jaime

    2017-10-04

    Chagas disease is caused by the protozoan parasite Trypanosoma cruzi. Assessment of parasitological cure upon treatment with available drugs relies on achieving consistent negative results in conventional parasitological and serological tests, which may take years to assess. Here, we evaluated the use of a recombinant T. cruzi antigen termed TSSA as an early serological marker of drug efficacy in T. cruzi-infected children. A cohort of 78 pediatric patients born to T. cruzi-infected mothers was included in this study. Solely 39 of them were infected with T. cruzi, and were immediately treated with trypanocidal drugs. Serological responses against TSSA were evaluated in infected and non-infected populations during the follow-up period using an in-house ELISA test, and compared to conventional serological methods. Anti-TSSA antibody titers decreased significantly faster than anti-whole parasite antibodies detected by conventional serology in both T. cruzi-infected patients undergoing effective treatment and in those not infected. This differential kinetics allowed a significant reduction in the required follow-up periods to evaluate therapeutic responses or to rule out maternal-fetal transmission, respectively. Finally, we present the case of a congenitally-infected patient with atypical course, in which TSSA provided an early marker for T. cruzi infection. In conclusion, we showed that TSSA was efficacious both for rapid assessment of treatment efficiency and for early negative diagnosis in infants at risk of congenital T. cruzi infection. Based upon these findings we propose the inclusion of TSSA for refining the post-therapeutic cure criterion and other diagnostic needs in pediatric Chagas disease. Copyright © 2017 American Society for Microbiology.

  6. Curcumin induced nanoscale CD44 molecular redistribution and antigen-antibody interaction on HepG2 cell surface

    Energy Technology Data Exchange (ETDEWEB)

    Wang Mu [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Ruan Yuxia [Department of Ophthalmology, The First Affiliated Hospital, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Xing Xiaobo; Chen Qian; Peng, Yuan [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China); Cai Jiye, E-mail: tjycai@jnu.edu.cn [Department of Chemistry, Jinan University, 601 Huangpu Road West, Tianhe District, Guangzhou 510632 (China)

    2011-07-04

    Graphical abstract: Highlights: > In this study, we investigate the changes of CD44 expression and distribution on HepG2 cells after curcumin treatment. > We find curcumin is able to change the morphology and ultrastructure of HepG2 cells. > Curcumin can reduce the expression of CD44 molecules and induce the nanoscale molecular redistribution on cell surface. > The binding force between CD44-modified AFM tip and the HepG2 cell surface decreases after curcumin-treatment. - Abstract: The cell surface glycoprotein CD44 was implicated in the progression, metastasis and apoptosis of certain human tumors. In this study, we used atomic force microscope (AFM) to monitor the effect of curcumin on human hepatocellular carcinoma (HepG2) cell surface nanoscale structure. High-resolution imaging revealed that cell morphology and ultrastructure changed a lot after being treated with curcumin. The membrane average roughness increased (10.88 {+-} 4.62 nm to 129.70 {+-} 43.72 nm) and the expression of CD44 decreased (99.79 {+-} 0.16% to 75.14 {+-} 8.37%). Laser scanning confocal microscope (LSCM) imaging showed that CD44 molecules were located on the cell membrane. The florescence intensity in control group was weaker than that in curcumin treated cells. Most of the binding forces between CD44 antibodies and untreated HepG2 cell membrane were around 120-220 pN. After being incubated with curcumin, the major forces focused on 70-150 pN (10 {mu}M curcumin-treated) and 50-120 pN (20 {mu}M curcumin-treated). These results suggested that, as result of nanoscale molecular redistribution, changes of the cell surface were in response to external treatment of curcumin. The combination of AFM and LSCM could be a powerful method to detect the distribution of cell surface molecules and interactions between molecules and their ligands.

  7. The mannose receptor acts as hepatitis B virus surface antigen receptor mediating interaction with intrahepatic dendritic cells

    NARCIS (Netherlands)

    Op den Brouw, Marjoleine L.; Binda, Rekha S.; Geijtenbeek, Teunis B. H.; Janssen, Harry L. A.; Woltman, Andrea M.

    2009-01-01

    Dendritic cells (DC) play a key role in anti-viral immunity. Direct interactions between DC and hepatitis B virus (HBV) may explain the impaired DC function and the ineffective anti-viral response of chronic HBV patients resulting in HBV persistence. Here, the interaction between HBV surface

  8. Virus-like particle production with yeast: ultrastructural and immunocytochemical insights into Pichia pastoris producing high levels of the Hepatitis B surface antigen

    Directory of Open Access Journals (Sweden)

    Adnan Ahmad

    2011-06-01

    Full Text Available Abstract Background A protective immune response against Hepatitis B infection can be obtained through the administration of a single viral polypeptide, the Hepatitis B surface antigen (HBsAg. Thus, the Hepatitis B vaccine is generated through the utilization of recombinant DNA technology, preferentially by using yeast-based expression systems. However, the polypeptide needs to assemble into spherical particles, so-called virus-like particles (VLPs, to elicit the required protective immune response. So far, no clear evidence has been presented showing whether HBsAg assembles in vivo inside the yeast cell into VLPs or later in vitro during down-stream processing and purification. Results High level production of HBsAg was carried out with recombinant Pichia pastoris using the methanol inducible AOX1 expression system. The recombinant vaccine was isolated in form of VLPs after several down-stream steps from detergent-treated cell lysates. Search for the intracellular localization of the antigen using electron microscopic studies in combination with immunogold labeling revealed the presence of HBsAg in an extended endoplasmic reticulum where it was found to assemble into defined multi-layered, lamellar structures. The distance between two layers was determined as ~6 nm indicating that these lamellas represent monolayers of well-ordered HBsAg subunits. We did not find any evidence for the presence of VLPs within the endoplasmic reticulum or other parts of the yeast cell. Conclusions It is concluded that high level production and intrinsic slow HBsAg VLP assembly kinetics are leading to retention and accumulation of the antigen in the endoplasmic reticulum where it assembles at least partly into defined lamellar structures. Further transport of HBsAg to the Golgi apparatus is impaired thus leading to secretory pathway disfunction and the formation of an extended endoplasmic reticulum which bulges into irregular cloud-shaped formations. As VLPs were

  9. Serum hepatitis B surface antigen titer and transient elastography in screening for insignificant fibrosis in HBeAg-positive chronic hepatitis B patients

    Directory of Open Access Journals (Sweden)

    Liang LB

    2015-02-01

    Full Text Available Ling-Bo Liang, Xia Zhu, Li-Bo Yan, Ling-Yao Du, Cong Liu, Li-Yu Chen, Juan Liao, Hong Tang Center of Infectious Disease, West China Hospital, West China School of Medicine, and State Key Laboratory of Biotherapy, Sichuan University, Chengdu, People’s Republic of China Objective: To explore the predictive value of serum hepatitis B surface antigen (HBsAg titer and transient elastography in screening for insignificant fibrosis in hepatitis B e antigen (HBeAg-positive chronic hepatitis B patients. Methods: We conducted a cross-sectional study of eligible patients treated from March 2012 to May 2013 at the West China Hospital of Sichuan University. Eligible patients underwent liver transient elastography and liver biopsy. We assessed the serum HBsAg level, serum hepatitis B virus (HBV deoxyribonucleic acid (DNA level, HBV genotypes, liver stiffness measurement (LSM values by transient elastography, and histological fibrosis staging by METAVIR classification. Results: A total of 129 consecutive patients were recruited. The LSM value (P<0.001, odds ratio 14.67, 95% CI 0.158–0.551 and log10HBsAg (P=0.045, odds ratio 4.03, 95% CI 0.136–0.976 correlated with a liver fibrosis score

  10. Cell surface expression of MR1B, a splice variant of the MHC class I-related molecule MR1, revealed with antibodies.

    Science.gov (United States)

    Yamaguchi, Hisateru; Tsukamoto, Kentaro; Hashimoto, Keiichiro

    2014-01-10

    The major histocompatibility complex (MHC) class I-related molecule, MR1, is highly conserved in mammals and can present bacteria-derived vitamin B metabolites to mucosal-associated invariant T (MAIT) cells, possibly having important defense function in the microbial infection. MR1B is a splice variant of MR1 and possesses an intriguing domain structure with only two extracellular domains resembling some NKG2D ligand molecules. Thus far, cell surface expression of MR1B could not be analyzed with flow cytometry due to a lack of appropriate antibodies reactive with MR1B. Here we clarified the expression of MR1B recombinant protein on the cell surface of the transfected cells by flow cytometry analyses using the antiserum against MR1. Consistently, MR1B tagged with FLAG peptide at the N-terminus also could be detected with anti-FLAG monoclonal antibodies. Our result showed that MR1B can be recognized on the cell surface by macromolecules such as antibodies, indicating its potential of interaction with certain receptor(s). We discuss possibility of interaction of MR1B and/or the full-length MR1 with some receptor(s) other than αβ T cell receptor (TCR) of MAIT cells based on the highly conserved characteristic residues of the ligand-binding domains of MR1 and its MAIT cells αβTCR footprints. Copyright © 2013 Elsevier Inc. All rights reserved.

  11. MYD88-DEPENDENT PROTECTIVE IMMUNITY ELICITED BY ADENOVIRUS 5 EXPRESSING THE SURFACE ANTIGEN 1 FROM TOXOPLASMA GONDII IS MEDIATED BY CD8+ T LYMPHOCYTES

    Science.gov (United States)

    Mendes, Érica A.; Caetano, Bráulia C.; Penido, Marcus L. O.; Bruna-Romero, Oscar; Gazzinelli, Ricardo T.

    2011-01-01

    Toxoplasma gondii is an intracellular parasite widely spread around the world. The Surface Antigens (SAG) 1, 2 and 3 are the main proteins expressed on the surface of T. gondii tachyzoites. Replication-defective adenovirus serotype 5 (rAd5) is one of the most potent recombinant viral vectors for eliciting T cell-mediated immunity in mice and humans. Here we show that vaccination with rAd5 expressing SAG1 (AdSAG1), but neither SAG2 nor SAG3, induces protective immunity in the highly susceptible C57BL/6 mice challenged with T. gondii. Furthermore, we evaluated different immunological components involved on viral induced protective immunity. We observed that host protection elicited by AdSAG1 is highly dependent on IL-12, IFN-γ and CD8+ T lymphocytes. Importantly, the induction of protective immunity (T cell-derived IFN-γ) was also dependent on Myeloid Differentiation Factor 88 (MyD88), and thus, likely to involve Toll-Like receptors. We conclude that protective parasite specific-CD8+ T cells are elicited by a mechanism that involves MyD88-dependent induction of IL-12. PMID:21549794

  12. Protective immunity to Haemonchus contortus induced by immunoaffinity isolated antigens that share a phylogenetically conserved carbohydrate gut surface epitope.

    Science.gov (United States)

    Jasmer, D P; Perryman, L E; Conder, G A; Crow, S; McGuire, T

    1993-11-15

    Whole gut homogenates of the blood-sucking nematode Haemonchus contortus induce protective immunity in goats, and some of these gut Ag are conserved among related parasitic nematode species. To identify gut Ag that induce protective immunity and have phylogenetically conserved epitopes, mAb were made to gut-surface Ag of H. contortus. Forty-nine mAb reacted with microvilli of the parasite gut. Two of these mAb (42/10.6.1 and 42/53.3.5) were analyzed here. Both of the mAb bound to the microvillar surface of freshly isolated gut, and each mAb recognized carbohydrate epitopes, based on sensitivity to periodate oxidation. The 42/10.6.1 epitope occurred on at least 18 proteins in Western blots and in several H. contortus tissues. Proteins recognized by this mAb localized to membrane and excretory/secretory fractions of the worm. This epitope was also identified on the gut and other tissues and multiple proteins of related adult and larval nematodes, including larval Ancylostoma caninum and a mixed population of the free-living nematode Caenorhabditis elegans. In contrast, the 42/53.3.5 mAb bound to the gut surface and recognized proteins of 100 and 46 kDa from adult H. contortus gut. Four proteins of 100, 52, 46, and 30 kDa were isolated from the 42/53.3.5 immunoaffinity columns, and except for the 30-kDa protein, each was recognized by both the 42/10.6.1 and 42/53.3.5 mAb. Epitopes recognized by each mAb were distinct from one another and phosphorylcholine. When used to immunize goats, Ag isolated by both mAb induced protection that significantly (p < 0.05) reduced total worm counts after challenge infections compared with the control groups.

  13. Correlation of Hepatitis B surface antigen mutations with clinical status of the chronically infected patients from Kermanshah, West of Iran.

    Science.gov (United States)

    Sayad, B; Anvari, F A; Alavian, S M; Norouzi, M; Hamzelooie, M; Shirvani, M; Pourhoseingholi, A; Khedive, A; Ghamari, S; Judaki, M A; Jazayeri, S M

    2012-03-01

    Hepatitis B virus (HBV) genetic and protein variations have been found in chronic HBV infected patients who did not receive any treatment and active or passive immunizations. The aims of this study were to determine the genotypes as well as the patterns of variations distribution in chronically infected patients from the west part of Iran. Forty-six people with chronic HBV infection were enrolled in the study. The surface genes were amplified, sequenced and subsequently aligned using international and national Iranian database. All strains belonged to genotype D, subgenotype D1 and subtype ayw2. Of all 116 "mutations" that occurred at 59 nucleotide positions, 49 (42.2 %) were missense (amino acid altering) and 67 (57.7%) were silent (no amino acid changing), respectively. At the amino acid level, 38 (79.1%) out of 48 amino acid mutations occurred in different immune epitopes within the surface proteins, of which 2 (5.2%) occurred in B cell; 12 (31.5%) in T helper and 24 (63.1%) inside CTL epitopes. There were significant associations between amino acid mutations (especially within immune epitopes) and anti-HBe positivity and increased ALT levels (P values: 0.014 and 0.04, respectively). The distribution of amino acid mutations as well as the ratio between silent and missense nucleotide mutations showed that a narrowly focused immune pressure had already been on the surface protein T cell epitopes (94.9% of mutations), particularly CTL epitopes which led to the emergence of escape mutants in these patients.

  14. Cationic Lipid-Formulated DNA Vaccine against Hepatitis B Virus : Immunogenicity of MIDGE-Th1 Vectors Encoding Small and Large Surface Antigen in Comparison to a Licensed Protein Vaccine

    NARCIS (Netherlands)

    Endmann, Anne; Klunder, Katharina; Kapp, Kerstin; Riede, Oliver; Oswald, Detlef; Talman, Eduard G.; Schroff, Matthias; Kleuss, Christiane; Ruiters, Marcel H. J.; Juhls, Christiane

    2014-01-01

    Currently marketed vaccines against hepatitis B virus (HBV) based on the small (S) hepatitis B surface antigen (HBsAg) fail to induce a protective immune response in about 10% of vaccinees. DNA vaccination and the inclusion of PreS1 and PreS2 domains of HBsAg have been reported to represent feasible

  15. Natural Killer Cell Characteristics in Patients With Chronic Hepatitis B Virus (HBV) Infection Are Associated With HBV Surface Antigen Clearance After Combination Treatment With Pegylated Interferon Alfa-2a and Adefovir

    NARCIS (Netherlands)

    Stelma, Femke; de Niet, Annikki; Tempelmans Plat-Sinnige, Marjan J.; Jansen, Louis; Takkenberg, R. Bart; Reesink, Hendrik W.; Kootstra, Neeltje A.; van Leeuwen, Ester M. M.

    2015-01-01

    The role of natural killer (NK) cells in the process of hepatitis B virus (HBV) surface antigen (HBsAg) clearance and whether their phenotype is related to treatment outcome in patients with chronic hepatitis B are currently unknown. Patients with chronic hepatitis B (HBV DNA load, >17 000 IU/mL)

  16. Characterization of foot- and mouth disease virus antigen by surface-enhanced laser desorption ionization-time of flight-mass spectrometry in aqueous and oil-emulsion formulations

    NARCIS (Netherlands)

    Harmsen, M.M.; Jansen, J.; Westra, D.F.; Coco-Martin, J.M.

    2010-01-01

    We have used a novel method, surface-enhanced laser desorption ionization-time of flight-mass spectrometry (SELDI-TOF-MS), to characterize foot-and-mouth disease virus (FMDV) vaccine antigens. Using specific capture with FMDV binding recombinant antibody fragments and tryptic digestion of FMDV

  17. Antigenicity and immunogenicity of a novel chimeric peptide antigen based on the P. vivax circumsporozoite protein.

    Science.gov (United States)

    Céspedes, Nora; Arévalo-Herrera, Myriam; Felger, Ingrid; Reed, Steve; Kajava, Andrey V; Corradin, Giampietro; Herrera, Sócrates

    2013-10-01

    Plasmodium vivax circumsporozoite (PvCS) protein is a major sporozoite surface antigen involved in parasite invasion of hepatocytes and is currently being considered as vaccine candidate. PvCS contains a dimorphic central repetitive fragment flanked by conserved regions that contain functional domains. We have developed a chimeric 137-mer synthetic polypeptide (PvCS-NRC) that includes the conserved region I and region II-plus and the two natural repeat variants known as VK210 and VK247. The antigenicity of PvCS-NRC was tested using human sera from PNG and Colombia endemic areas and its immunogenicity was confirmed in mice with different genetic backgrounds, the polypeptide formulated either in Alum or GLA-SE adjuvants was assessed in inbred C3H, CB6F1 and outbred ICR mice, whereas a formulation in Montanide ISA51 was tested in C3H mice. Antigenicity studies indicated that the chimeric peptide is recognized by a high proportion (60-70%) of residents of malaria-endemic areas. Peptides formulated with either GLA-SE or Montanide ISA51 adjuvants induced stronger antibody responses as compared with the Alum formulation. Sera from immunized mice as well as antigen-specific affinity purified human IgG antibodies reacted with sporozoite preparations in immunofluorescence and Western blot assays, and displayed strong in vitro inhibition of sporozoite invasion (ISI) into hepatoma cells. The polypeptide was recognized at high prevalence when tested against naturally induced human antibodies and was able to induce significant immunogenicity in mice. Additionally, specific antibodies were able to recognize sporozoites and were able to block sporozoite invasion in vitro. Further evaluation of this chimeric protein construct in preclinical phase e.g. in Aotus monkeys in order to assess the humoral and cellular immune responses as well as protective efficacy against parasite challenge of the vaccine candidate must be conducted. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Occult hepatitis B virus infection: influence of S protein variants.

    Science.gov (United States)

    Zhang, Zhenhua; Zhang, Ling; Dai, Yu; Zhang, Yafei; Li, Jun; Li, Xu

    2016-01-19

    In occult hepatitis B viral infection (OBI), the persistence of hepatitis B virus (HBV) DNA is associated with a lack of hepatitis B surface antigen (HBsAg). To assess the possible role of HBsAg immune escape variants in OBI patients, variability in the HBV S gene was evaluated for OBI patients as well as chronic HBV infection patients from the same families. We selected 17 HBV DNA-positive/HBsAg-negative patients (OBI group) and 15 HBV DNA- and HBsAg-positive patients from OBI families (control group). The S gene was amplified and cloned, and at least 15 clones per patient were sequenced and analyzed. Although the incidence of stop codon mutations within the S region was higher in the OBI group (13.6 %) than in the control group (1.5 %, P type of mutation, together with insertion and deletion mutations, was prevalent in only three OBI patients. In the major hydrophilic region (MHR), a median of 0.75 residues were altered in every 100 residues for the OBI patients, whereas 0.95 out of 100 residues were changed in the control group (P = 0.428). Furthermore, some variants that are generally considered immune escape variants, such as mutations at positions s145, s147, and s123, were only observed in less than 5 % of all the clones sequenced, in either OBI or control group. Our data suggest that HBsAg variants may not play a major role in OBI pathogenesis.

  19. Changed epitopes drive the antigenic drift for influenza A (H3N2 viruses

    Directory of Open Access Journals (Sweden)

    Yang Jinn-Moon

    2011-02-01

    Full Text Available Abstract Background In circulating influenza viruses, gradually accumulated mutations on the glycoprotein hemagglutinin (HA, which interacts with infectivity-neutralizing antibodies, lead to the escape of immune system (called antigenic drift. The antibody recognition is highly correlated to the conformation change on the antigenic sites (epitopes, which locate on HA surface. To quantify a changed epitope for escaping from neutralizing antibodies is the basis for the antigenic drift and vaccine development. Results We have developed an epitope-based method to identify the antigenic drift of influenza A utilizing the conformation changes on epitopes. A changed epitope, an antigenic site on HA with an accumulated conformation change to escape from neutralizing antibody, can be considered as a "key feature" for representing the antigenic drift. According to hemagglutination inhibition (HI assays and HA/antibody complex structures, we statistically measured the conformation change of an epitope by considering the number of critical position mutations with high genetic diversity and antigenic scores. Experimental results show that two critical position mutations can induce the conformation change of an epitope to escape from the antibody recognition. Among five epitopes of HA, epitopes A and B, which are near to the receptor binding site, play a key role for neutralizing antibodies. In addition, two changed epitopes often drive the antigenic drift and can explain the selections of 24 WHO vaccine strains. Conclusions Our method is able to quantify the changed epitopes on HA for predicting the antigenic variants and providing biological insights to the vaccine updates. We believe that our method is robust and useful for studying influenza virus evolution and vaccine development.

  20. A computational framework for influenza antigenic cartography.

    Directory of Open Access Journals (Sweden)

    Zhipeng Cai

    2010-10-01

    Full Text Available Influenza viruses have been responsible for large losses of lives around the world and continue to present a great public health challenge. Antigenic characterization based on hemagglutination inhibition (HI assay is one of the routine procedures for influenza vaccine strain selection. However, HI assay is only a crude experiment reflecting the antigenic correlations among testing antigens (viruses and reference antisera (antibodies. Moreover, antigenic characterization is usually based on more than one HI dataset. The combination of multiple datasets results in an incomplete HI matrix with many unobserved entries. This paper proposes a new computational framework for constructing an influenza antigenic cartography from this incomplete matrix, which we refer to as Matrix Completion-Multidimensional Scaling (MC-MDS. In this approach, we first reconstruct the HI matrices with viruses and antibodies using low-rank matrix completion, and then generate the two-dimensional antigenic cartography using multidimensional scaling. Moreover, for influenza HI tables with herd immunity effect (such as those from Human influenza viruses, we propose a temporal model to reduce the inherent temporal bias of HI tables caused by herd immunity. By applying our method in HI datasets containing H3N2 influenza A viruses isolated from 1968 to 2003, we identified eleven clusters of antigenic variants, representing all major antigenic drift events in these 36 years. Our results showed that both the completed HI matrix and the antigenic cartography obtained via MC-MDS are useful in identifying influenza antigenic variants and thus can be used to facilitate influenza vaccine strain selection. The webserver is available at http://sysbio.cvm.msstate.edu/AntigenMap.

  1. A computational framework for influenza antigenic cartography.

    Science.gov (United States)

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

    2010-10-07

    Influenza viruses have been responsible for large losses of lives around the world and continue to present a great public health challenge. Antigenic characterization based on hemagglutination inhibition (HI) assay is one of the routine procedures for influenza vaccine strain selection. However, HI assay is only a crude experiment reflecting the antigenic correlations among testing antigens (viruses) and reference antisera (antibodies). Moreover, antigenic characterization is usually based on more than one HI dataset. The combination of multiple datasets results in an incomplete HI matrix with many unobserved entries. This paper proposes a new computational framework for constructing an influenza antigenic cartography from this incomplete matrix, which we refer to as Matrix Completion-Multidimensional Scaling (MC-MDS). In this approach, we first reconstruct the HI matrices with viruses and antibodies using low-rank matrix completion, and then generate the two-dimensional antigenic cartography using multidimensional scaling. Moreover, for influenza HI tables with herd immunity effect (such as those from Human influenza viruses), we propose a temporal model to reduce the inherent temporal bias of HI tables caused by herd immunity. By applying our method in HI datasets containing H3N2 influenza A viruses isolated from 1968 to 2003, we identified eleven clusters of antigenic variants, representing all major antigenic drift events in these 36 years. Our results showed that both the completed HI matrix and the antigenic cartography obtained via MC-MDS are useful in identifying influenza antigenic variants and thus can be used to facilitate influenza vaccine strain selection. The webserver is available at http://sysbio.cvm.msstate.edu/AntigenMap.

  2. Radioimmunoassays of hidden viral antigens

    Energy Technology Data Exchange (ETDEWEB)

    Neurath, A.R. (Lindsley F. Kimbell Research Inst., New York, NY); Strick, N.; Baker, L.; Krugman, S.

    1982-07-01

    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.

  3. New surface-associated heat-labile colonization factor antigen (CFA/II) produced by enterotoxigenic Escherichia coli of serogroups O6 and O8.

    Science.gov (United States)

    Evans, D G; Evans, D J

    1978-08-01

    Enterotoxigenic Escherichia coli (ETEC) belonging to serogroups O6 and O8 do not possess the H-10407-type colonization factor antigen (CFA/I). However, these frequently isolated ETEC were found to possess a second and distinct heat-labile surface-associated colonization factor antigen, termed CFA/II. Whereas CFA/I mediates mannose-resistant hemagglutination of human group A erythrocytes, CFA/II does not. CFA/II mediates mannose-resistant hemagglutination of bovine erythrocytes, and mannose-resistant hemagglutination is rapid only at reduced temperature (4 degrees C). Because CFA/II, like CFA/I, is spontaneously lost by many ETEC isolates in the laboratory, it was possible to produce specific anti-CFA/II serum by preparing antiserum against living cells of a prototype strain (PB-176) and adsorbing this serum with living and heat-treated cells of its CFA/II-negative derivative strain PB-176-P. This serum, which neutralized the colonization factor activity of CFA/II-positive strains in infant rabbits, was employed to confirm the presence of CFA/II on ETEC which exhibited mannose-resistant hemagglutination of bovine but not human erythrocytes. CFA/II, like CFA/I, mediates adherence of the bacteria to the mucosal surface of the small intestine, as demonstrated by indirect immunofluorescence. CFA/II appears to be an important virulence factor for humans since CFA/II-positive ETEC are frequently isolated from diarrhea cases, particularly travelers' diarrhea, in Mexico; these ETEC were not uncommon in a collection of isolates from Bangladesh. The O6:H16 strain of ETEC responsible for an outbreak of diarrhea in the United States was also shown to be CFA/II positive. CFA/I and CFA/II were never found on the same serotypes of ETEC, but 98% of the heat-stable and heat-labile enterotoxin-producing ETEC belonging to the frequently isolated serogroups O6, O8, O15, O25, O63, and O78 were positive for either CFA/I or CFA/II.

  4. Genetic variant as a marker for bladder cancer therapy

    Science.gov (United States)

    Patients who have inherited a specific common genetic variant develop bladder cancer tumors that strongly express a protein known as prostate stem cell antigen (PSCA), which is also expressed in many pancreatic and prostate tumors, according to research a

  5. Trehalose 6,6′-dimycolate on the surface of Mycobacterium tuberculosis modulates surface marker expression for antigen presentation and costimulation in murine macrophages

    OpenAIRE

    Kan-Sutton, Celestine; Jagannath, Chinnaswamy; Hunter, Robert L.

    2008-01-01

    Trehalose 6,6′-dimycolate (TDM) is the most abundant lipid extracted from Mycobacterium tuberculosis (MTB). TDM promotes MTB survival by decreasing phagosomal acidification and phagolysosomal fusion in macrophages. Delipidation of MTB using petroleum ether removes TDM and decreases MTB survival within host cells. TDM reconstituted onto MTB restores its virulent wild-type characteristics. We investigated the role of TDM in regulating surface marker expression in MTB-infected macrophages. Macro...

  6. Detection of the tuberculosis antigenic marker mannose-capped lipoarabinomannan in pretreated serum by surface-enhanced Raman scattering.

    Science.gov (United States)

    Crawford, Alexis C; Laurentius, Lars B; Mulvihill, Timothy S; Granger, Jennifer H; Spencer, John S; Chatterjee, Delphi; Hanson, Kimberly E; Porter, Marc D

    2016-12-19

    The ability to detect tuberculosis (TB) continues to be a global health care priority. This paper describes the development and preliminary assessment of the clinical accuracy of a heterogeneous immunoassay that integrates a serum pretreatment process with readout by surface-enhanced Raman scattering (SERS) for the low-level detection of mannose-capped lipoarabinomannan (ManLAM). ManLAM is a major virulence factor in the infectious pathology of Mycobacterium tuberculosis (Mtb) that has been found in the serum and other body fluids of infected patients. The effectiveness of ManLAM as a TB diagnostic marker, however, remains unproven for reasons not yet well understood. As reported herein, we have found that (1) ManLAM complexes with proteins and possibly other components in serum; (2) these complexes have a strongly detrimental impact on the ability to detect ManLAM using an immunoassay; (3) a simple pretreatment step can disrupt this complexation; and (4) disruption by pretreatment improves detection by 250×. We also describe the results from a preliminary assessment on the utility of serum pretreatment by running immunoassays on archived specimens from 24 TB-positive patients and 10 healthy controls. ManLAM was measurable in 21 of the 24 TB-positive specimens, but not in any of the 10 control specimens. These findings, albeit for a very small specimen set, translate to a clinical sensitivity of 87.5% and a clinical specificity of 100%. Together, these results both provide much needed evidence for the clinical utility of ManLAM as a TB marker, and demonstrate the potential utility of our overall approach to serve as a new strategy for the development of diagnostic tests for this disease.

  7. Hepatitis B vaccination coverage and risk factors associated with incomplete vaccination of children born to hepatitis B surface antigen-positive mothers, Denmark, 2006 to 2010.

    Science.gov (United States)

    Kunoee, Asja; Nielsen, Jens; Cowan, Susan

    2016-01-01

    In Denmark, universal screening of pregnant women for hepatitis B has been in place since November 2005, with the first two years as a trial period with enhanced surveillance. It is unknown what the change to universal screening without enhanced surveillance has meant for vaccination coverage among children born to hepatitis B surface antigen (HBsAg)-positive mothers and what risk factors exist for incomplete vaccination. This retrospective cohort study included 699 children of mothers positive for HBsAg. Information on vaccination and risk factors was collected from central registers. In total, 93% (651/699) of the children were vaccinated within 48 hours of birth, with considerable variation between birthplaces. Only 64% (306/475) of the children had received all four vaccinations through their general practitioner (GP) at the age of two years, and 10% (47/475) of the children had received no hepatitis B vaccinations at all. Enhanced surveillance was correlated positively with coverage of birth vaccination but not with coverage at the GP. No or few prenatal examinations were a risk factor for incomplete vaccination at the GP. Maternity wards and GPs are encouraged to revise their vaccination procedures and routines for pregnant women, mothers with chronic HBV infection and their children.

  8. Detection of Leishmania-specific DNA and surface antigens using a combination of functionalized magnetic beads and cadmium selenite quantum dots.

    Science.gov (United States)

    Andreadou, Margarita; Liandris, Emmanouil; Gazouli, Maria; Mataragka, Antonia; Tachtsidis, Ilias; Goutas, Nikolaοs; Vlachodimitropoulos, Dimitrios; Ikonomopoulos, John

    2016-04-01

    Leishmaniosis is a zoonotic disease that affects millions of people especially in resource-poor settings. The development of reliable diagnostic assays that do not require dedicated equipment or highly trained personnel would improve early diagnosis and effective control. For this purpose, a combination of magnetic bead and cadmium selenite quantum dot probes was applied for the detection of Leishmania-specific surface antigens (proteins) and DNA. Both analytes are isolated from the solution using magnetic bead capture probes whereas the presence of the targeted molecules is demonstrated by quantum dot detection probes. The sensitivity and specificity of this method reached 100% based on an assessment performed on 55 cultured isolates of various microbial pathogens. The low limit of detection was 3125 ng/μl and 10(3)cells/ml for Leishmania DNA and protein, respectively. The method shows considerable potential for clinical application in human and veterinary medicine, especially in resource-poor settings. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Antibody response to a major human Pneumocystis carinii surface antigen in patients without evidence of immunosuppression and in patients with suspected atypical pneumonia

    DEFF Research Database (Denmark)

    Lundgren, Bettina; Lebech, M; Lind, K

    1993-01-01

    IgG and IgM antibodies to a purified human Pneumocystis carinii surface antigen (gp95) were measured in 694 serum specimens from two different population groups using an EIA technique. In a population of 441 patients with no evidence of immunosuppression, the percentage of persons positive for Ig......G antibodies to gp95 was significantly lower in the age group 1 to 9 years (30%, 23/77) compared to persons 10 to 19 years old (56%, 49/88). In the age group 1 to 14 years there was a significant correlation between the percentage of persons with IgG antibodies to gp95 and age. In 106 consecutive patients...... under evaluation due to atypical pneumonia, 76 patients showed no change in the titre of antibodies to Legionella spp. or Mycoplasma pneumoniae in two consecutive serum samples. Three of these 76 patients (4%) demonstrated an increase in the level of IgG antibodies to gp95 in the paired samples. One...

  10. Comparison between Elecsys HBsAg II and Architect assays for quantification of serum hepatitis B surface antigen in Chinese patients with chronic hepatitis B.

    Science.gov (United States)

    Liu, Wei-Wei; Wang, Ai-Zhen; Xie, Fu-Yi; Hu, Ting-Ting; Zhong, Qi; Huang, Sheng

    2015-01-01

    Chronic hepatitis B (CHB) is an important global health problem. Recent innovations have rendered quantification of serum hepatitis B surface antigen (HBsAg) a valuable tool in hepatitis B virus (HBV) disease management and for determining the effectiveness of drug treatment. The aim of the present study was to compare the performances of the Elecsys HBsAg II and Abbott Architect HBsAg assays in Chinese patients with CHB, with predominantly genotypes B and C. A dilution protocol was developed for the Elecsys assay to allow quantification of HBsAg levels. Sera were obtained from patients with various HBV genotypes, including HBV mutants, and longitudinal samples were obtained from patients undergoing antiviral treatment. There was a significant overall correlation between the Elecsys and Architect assays (r = 0.9881; p Architect assay were higher than those obtained by the Elecsys assay. There was a good correlation between the results of the Elecsys HBsAg II and Abbott Architect HBsAg assays in patients with CHB, especially those with genotype C.

  11. Correlation between the Elecsys HBsAg II assay and the Architect assay for the quantification of hepatitis B surface antigen (HBsAg) in the serum.

    Science.gov (United States)

    Wursthorn, Karsten; Jaroszewicz, Jerzy; Zacher, Behrend J; Darnedde, Martina; Raupach, Regina; Mederacke, Ingmar; Cornberg, Markus; Manns, Michael P; Wedemeyer, Heiner

    2011-04-01

    Hepatitis B surface antigen (HBsAg) clearance during chronic hepatitis B (CHB) infection is associated with improved long-term clinical outcome, so is considered an important therapeutic goal in CHB. Studies have shown that serum HBsAg quantification during, and at end of, treatment may predict long-term HBsAg loss. Performance comparison of the qualitative Elecsys HBsAg II assay using a quantitative research protocol and an established quantitative HBsAg assay. A dilution algorithm was developed for the Elecsys HBsAg II assay to allow quantification of HBsAg levels; this was used to measure HBsAg levels in a range of samples including sera from patients infected with different HBV genotypes, HBV mutants, and longitudinal samples from patients undergoing antiviral treatment. Results were compared with those from the quantitative Architect HBsAg assay. There was significant overall correlation between Elecsys and Architect assays (correlation coefficient [r]=0.97; pArchitect HBsAg assay. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Comparison of the Elecsys HBsAg II Assay and the Architect Assay for Quantification of hepatitis B surface antigen in patients with chronic hepatitis B.

    Science.gov (United States)

    Karagoz, Ergenekon; Selek, Mehmet Burak; Tanoglu, Alpaslan; Hatipoglu, Mustafa; Ulçay, Asim; Turhan, Vedat

    2016-12-01

    Quantitative hepatitis B surface antigen (HBsAg) is a valuable tool in hepatitis B virus (HBV) disease diagnosis and management for evaluating the effectiveness of antiviral therapy. The aim of the current research was to compare the performances of the Elecsys HBsAg II and Abbott Architect HBsAg assays in chronic hepatitis B (CHB) patients. Between May 2014 and December 2014, 72 CHBs were tested using Abbott Architect HBsAg QT and Roche Elecsys HBsAg II assay. After transformation to log (10) IU/mL, the results of the two assays were compared using the interclass correlation test, the Pearson correlation test and Bland-Altman analyses. We also analyzed all the parameters in on-treatment patients and naive patients with Pearson correlation test. There was a significant overall correlation between the Elecsys and Architect assays. We also analyzed all the parameters in naive and on-treatment patients. There was a significantly good correlation between the two assays in untreated patients and on-treatment patients. In this study, there was a significant correlation between the results of the Elecsys HBsAg II and Abbott Architect HBsAg assays in the overall and naive/on-treatment CHB patients. Additionally, we found that mean HBsAg levels resulting from the Architect assay were higher than those obtained by Elecsys assay.

  13. Enhanced immunogenicity of an anti-caries vaccine encoding a cell-surface protein antigen of Streptococcus mutans by intranasal DNA prime-protein boost immunization.

    Science.gov (United States)

    Li, Yuhong; Jin, Jie; Yang, Yaping; Bian, Zhuan; Chen, Zhi; Fan, Mingwen

    2009-11-01

    The present study aimed to enhance the specific anti-caries immunity induced by DNA prime-protein boost strategy for an A-P fragment of a cell-surface protein antigen of Streptococcus mutans (PAc). BALB/c mice were immunized with DNA prime-protein boost, DNA-DNA or protein-protein regimens by the intranasal route, using combinations of plasmid vector (pCIA-P) that express PAc protein and a pure secretec recombinant PAc protein (rPAc). Then, a gnotobiotic mouse model was constructed 2 weeks after the last immunization, and specific immune responses in vivo and their protection against dental caries were observed. The present study revealed stronger antibody responses in the DNA prime-protein boost group compared to those elicited by either DNA-DNA vaccination or protein-protein vaccination. In particular, PAc-specific antibody concentrations were improved significantly after boosting the pCIA-P DNA-primed mice with rPAc. Moreover, protection against S. mutans challenge was obtained in the mice treated with the DNA prime-protein boost vaccination, as demonstrated by a significant reduction in S. mutans colonization compared to control mice and animals immunized with the DNA-DNA vaccination or protein-protein vaccination. The results obtained in the present study suggest that the intranasal DNA prime-protein boost vaccination regimen is a novel strategy for the practical application of DNA vaccine against dental caries.

  14. Immunogenicity of Leishmania-derived hepatitis B small surface antigen particles exposing highly conserved E2 epitope of hepatitis C virus.

    Science.gov (United States)

    Czarnota, Anna; Tyborowska, Jolanta; Peszyńska-Sularz, Grażyna; Gromadzka, Beata; Bieńkowska-Szewczyk, Krystyna; Grzyb, Katarzyna

    2016-04-13

    Hepatitis C virus (HCV) infection is a major health problem worldwide, affecting an estimated 2-3 % of human population. An HCV vaccine, however, remains unavailable. High viral diversity poses a challenge in developing a vaccine capable of eliciting a broad neutralizing antibody response against all HCV genotypes. The small surface antigen (sHBsAg) of hepatitis B virus (HBV) has the ability to form highly immunogenic subviral particles which are currently used as an efficient anti-HBV vaccine. It also represents an attractive antigen carrier for the delivery of foreign sequences. In the present study, we propose a bivalent vaccine candidate based on novel chimeric particles in which highly conserved epitope of HCV E2 glycoprotein (residues 412-425) was inserted into the hydrophilic loop of sHBsAg. The expression of chimeric protein was performed in an unconventional, Leishmania tarentolae expression system resulting in an assembly of particles which retained immunogenicity of both HCV epitope and sHBsAg protein. Direct transmission electron microscopy observation and immunogold staining confirmed the formation of spherical particles approximately 22 nm in diameter, and proper foreign epitope exposition. Furthermore, the sera of mice immunized with chimeric particles proved reactive not only to purified yeast-derived sHBsAg proteins but also HCV E2 412-425 synthetic peptide. Most importantly, they were also able to cross-react with E1E2 complexes from different HCV genotypes. For the first time, we confirmed successful assembly of chimeric sHBsAg virus-like particles (VLPs) in the L. tarentolae expression system which has the potential to produce high-yields of properly N-glycosylated mammalian proteins. We also proved that chimeric Leishmania-derived VLPs are highly immunogenic and able to elicit cross-reactive antibody response against HCV. This approach may prove useful in the development of a bivalent prophylactic vaccine against HBV and HCV and opens up a new

  15. Hepatitis B surface antigen titer is a good indicator of durable viral response after entecavir off-treatment for chronic hepatitis B

    Directory of Open Access Journals (Sweden)

    Han Ah Lee

    2016-09-01

    Full Text Available Background/Aims Clear indicators for stopping antiviral therapy in chronic hepatitis B (CHB patients are not yet available. Since the level of hepatitis B surface antigen (HBsAg is correlated with covalently closed circular DNA, the HBsAg titer might be a good indicator of the off-treatment response. This study aimed to determine the relationship between the HBsAg titer and the entecavir (ETV off-treatment response. Methods This study analyzed 44 consecutive CHB patients (age, 44.6±11.4 years, mean±SD; men, 63.6%; positive hepatitis B envelope antigen (HBeAg at baseline, 56.8%; HBV DNA level, 6.8±1.3 log10 IU/mL treated with ETV for a sufficient duration and in whom treatment was discontinued after HBsAg levels were measured. A virological relapse was defined as an increase in serum HBV DNA level of >2000 IU/mL, and a clinical relapse was defined as a virological relapse with a biochemical flare, defined as an increase in the serum alanine aminotransferase level of >2 × upper limit of normal. Results After stopping ETV, virological relapse and clinical relapse were observed in 32 and 24 patients, respectively, during 20.8±19.9 months of follow-up. The cumulative incidence rates of virological relapse were 36.2% and 66.2%, respectively, at 6 and 12 months, and those of clinical relapse were 14.3% and 42.3%. The off-treatment HBsAg level was an independent factor associated with clinical relapse (hazard ratio, 2.251; 95% confidence interval, 1.076–4.706; P=0.031. When patients were grouped according to off-treatment HBsAg levels, clinical relapse did not occur in patients with an off-treatment HBsAg level of ≤2 log10 IU/mL (n=5, while the incidence rates of clinical relapse at 12 months after off-treatment were 28.4% and 55.7% in patients with off-treatment HBsAg levels of >2 and ≤3 log10 IU/mL (n=11 and >3 log10 IU/mL (n=28, respectively. Conclusion The off-treatment HBsAg level is closely related to clinical relapse after treatment

  16. A Survey of ABO, Rhesus (D) Antigen and Haemoglobin Genes ...

    African Journals Online (AJOL)

    Summary: A survey of ABO and Rhesus (Rh D) antigens and variants of haemoglobin genes (HbGen) in Oyo state was carried out. This longitudinal study involved the determination of ABO and Rh(D) antigens in 3241 and HbGen in 2622 male and female adults (aged 26-65years) respectively using standard methods.

  17. Cancer vaccine--Antigenics.

    Science.gov (United States)

    2002-01-01

    purified complexes of tumour-derived HSPs linked to tumour antigen peptides. When these HSPPC are readministered to a patient following surgery or biopsy of the tumour, the antigenic tumour peptides are expressed on the surface of potent antigen-presenting cells of the immune system, such as macrophages and dendritic cells. This stimulates a much more powerful anti-tumour immune response than that generated by expression of the same antigens by the tumour cell. Thus, Antigenics autologous HSP technology is attractive because it is highly specific for individual patients and circumvents the need for identification of specific antigens for individual cancers (i.e. it does not require definition of the antigenic epitopes on cancer cells) and it overcomes the immune tolerance associated with various tumours. Oncophage is manufactured in a 10-hour process from surgically resected autologous tumour. A minimum of 1-3g of tumour tissue is required to produce enough Oncophage for a course of treatment. The major limiting factor for producing Oncophage from a particular cancer is the ability to purify HSP from that cancer. From clinical studies to date, Antigenics has been able to produce HSP from 100, 98, 90, 71 and 30% of colorectal carcinoma, renal cell carcinoma, melanoma, gastric cancer and pancreatic cancer tumours, respectively. The low success rate with pancreatic cancers is because of the high concentration of proteases in that tissue type. HSPs are a family of highly conserved proteins present in the cells of all organisms. They function as molecular chaperones, assisting the correct folding of polypeptides and aiding intracellular protein transport. In addition, HSPs associate with a broad range of peptides derived from intracellular protein degradation, including antigenic peptides produced in tumour cells. Antigenics has exclusively licensed worldwide rights to its HSP immunotherapeutic complexes from Mount Sinai School of Medicine and Fordham University in the USA. On

  18. Antigenic variation in Trypanosoma equiperdum.

    Science.gov (United States)

    Roth, C; Jacquemot, C; Giroud, C; Bringaud, F; Eisen, H; Baltz, T

    1991-01-01

    Trypanosoma equiperdum is an African trypanosome that causes dourine in horses. Like the other African trypanosomes, T. equiperdum escapes elimination by the immune system of its host by using an elaborate system of antigenic variant. The trypanosomes are covered by a coat consisting of a single protein called the variable surface glycoprotein (VSG) that acts as the major trypanosome immunogen. As the host responds to one VSG, trypanosomes covered with another VSG become dominant. There is a loose order of appearance of these VSG during the infection. The factors that affect the timing of VSG expression and the effective size of the VSG repertoire in T. equiperdum are reviewed. The VSG genes are generally activated by a process of duplicative transposition involving the duplication of a silent VSG gene and inserting a copy of the gene into an expression site. The order of VSG expression is related to the amount of homology between the silent gene and the expression site. The genes expressed late in infection lack extensive homology with the expression site and depend on homology with the gene in the expression site. The genes coding for VSG expressed late in infection are hybrid genes because of this mode of transfer. This transfer mechanism allows the trypanosome to create complex VSG genes from parts of several different silent genes that are each pseudogenes. Additionally, data are presented showing that only a limited portion of the VSG is actually seen by the host immune system. These factors indicate that the effective VSG repertoire is greater than the number of VSG genes in the trypanosome genome.

  19. Antigenic distance measurements for seasonal influenza vaccine selection.

    Science.gov (United States)

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

    2012-01-05

    Influenza vaccination is one of the major options to counteract the effects of influenza diseases. Selection of an effective vaccine strain is the key to the success of an effective vaccination program since vaccine protection can only be achieved when the selected influenza vaccine strain matches the antigenic variants causing future outbreaks. Identification of an antigenic variant is the first step to determine whether vaccine strain needs to be updated. Antigenic distance derived from immunological assays, such as hemagglutination inhibition, is commonly used to measure the antigenic closeness between circulating strains and the current influenza vaccine strain. Thus, consensus on an explicit and robust antigenic distance measurement is critical in influenza surveillance. Based on the current seasonal influenza surveillance procedure, we propose and compare three antigenic distance measurements, including Average antigenic distance (A-distance), Mutual antigenic distance (M-distance), and Largest antigenic distance (L-distance). With the assistance of influenza antigenic cartography, our simulation results demonstrated that M-distance is a robust influenza antigenic distance measurement. Experimental results on both simulation and seasonal influenza surveillance data demonstrate that M-distance can be effectively utilized in influenza vaccine strain selection. Copyright © 2011 Elsevier Ltd. All rights reserved.

  20. 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

  1. Rapid and Highly Sensitive Detection of Variant Creutzfeldt - Jakob Disease Abnormal Prion Protein on Steel Surfaces by Protein Misfolding Cyclic Amplification: Application to Prion Decontamination Studies

    Science.gov (United States)

    Belondrade, Maxime; Nicot, Simon; Béringue, Vincent; Coste, Joliette; Lehmann, Sylvain; Bougard, Daisy

    2016-01-01

    The prevalence of variant Creutzfeldt-Jakob disease (vCJD) in the population remains uncertain, although it has been estimated that 1 in 2000 people in the United Kingdom are positive for abnormal prion protein (PrPTSE) by a recent survey of archived appendix tissues. The prominent lymphotropism of vCJD prions raises the possibility that some surgical procedures may be at risk of iatrogenic vCJD transmission in healthcare facilities. It is therefore vital that decontamination procedures applied to medical devices before their reprocessing are thoroughly validated. A current limitation is the lack of a rapid model permissive to human prions. Here, we developed a prion detection assay based on protein misfolding cyclic amplification (PMCA) technology combined with stainless-steel wire surfaces as carriers of prions (Surf-PMCA). This assay allowed the specific detection of minute quantities (10−8 brain dilution) of either human vCJD or ovine scrapie PrPTSE adsorbed onto a single steel wire, within a two week timeframe. Using Surf-PMCA we evaluated the performance of several reference and commercially available prion-specific decontamination procedures. Surprisingly, we found the efficiency of several marketed reagents to remove human vCJD PrPTSE was lower than expected. Overall, our results demonstrate that Surf-PMCA can be used as a rapid and ultrasensitive assay for the detection of human vCJD PrPTSE adsorbed onto a metallic surface, therefore facilitating the development and validation of decontamination procedures against human prions. PMID:26800081

  2. Rapid and Highly Sensitive Detection of Variant Creutzfeldt-Jakob Disease Abnormal Prion Protein on Steel Surfaces by Protein Misfolding Cyclic Amplification: Application to Prion Decontamination Studies.

    Directory of Open Access Journals (Sweden)

    Maxime Belondrade

    Full Text Available The prevalence of variant Creutzfeldt-Jakob disease (vCJD in the population remains uncertain, although it has been estimated that 1 in 2000 people in the United Kingdom are positive for abnormal prion protein (PrPTSE by a recent survey of archived appendix tissues. The prominent lymphotropism of vCJD prions raises the possibility that some surgical procedures may be at risk of iatrogenic vCJD transmission in healthcare facilities. It is therefore vital that decontamination procedures applied to medical devices before their reprocessing are thoroughly validated. A current limitation is the lack of a rapid model permissive to human prions. Here, we developed a prion detection assay based on protein misfolding cyclic amplification (PMCA technology combined with stainless-steel wire surfaces as carriers of prions (Surf-PMCA. This assay allowed the specific detection of minute quantities (10-8 brain dilution of either human vCJD or ovine scrapie PrPTSE adsorbed onto a single steel wire, within a two week timeframe. Using Surf-PMCA we evaluated the performance of several reference and commercially available prion-specific decontamination procedures. Surprisingly, we found the efficiency of several marketed reagents to remove human vCJD PrPTSE was lower than expected. Overall, our results demonstrate that Surf-PMCA can be used as a rapid and ultrasensitive assay for the detection of human vCJD PrPTSE adsorbed onto a metallic surface, therefore facilitating the development and validation of decontamination procedures against human prions.

  3. Luteolin-7-O-Glucoside Present in Lettuce Extracts Inhibits Hepatitis B Surface Antigen Production and Viral Replication by Human Hepatoma Cells in Vitro

    Directory of Open Access Journals (Sweden)

    Xiao-Xian Cui

    2017-12-01

    Full Text Available Hepatitis B virus (HBV infection is endemic in Asia and chronic hepatitis B (CHB is a major public health issue worldwide. Current treatment strategies for CHB are not satisfactory as they induce a low rate of hepatitis B surface antigen (HBsAg loss. Extracts were prepared from lettuce hydroponically cultivated in solutions containing glycine or nitrate as nitrogen sources. The lettuce extracts exerted potent anti-HBV effects in HepG2 cell lines in vitro, including significant HBsAg inhibition, HBV replication and transcription inhibition, without exerting cytotoxic effects. When used in combination interferon-alpha 2b (IFNα-2b or lamivudine (3TC, the lettuce extracts synergistically inhibited HBsAg expression and HBV replication. By using differential metabolomics analysis, Luteolin-7-O-glucoside was identified and confirmed as a functional component of the lettuce extracts and exhibited similar anti-HBV activity as the lettuce extracts in vitro. The inhibition rate on HBsAg was up to 77.4%. Moreover, both the lettuce extracts and luteolin-7-O-glucoside functioned as organic antioxidants and, significantly attenuated HBV-induced intracellular reactive oxygen species (ROS accumulation. Luteolin-7-O-glucoside also normalized ROS-induced mitochondrial membrane potential damage, which suggests luteolin-7-O-glucoside inhibits HBsAg and HBV replication via a mechanism involving the mitochondria. Our findings suggest luteolin-7-O-glucoside may have potential value for clinical application in CHB and may enhance HBsAg and HBV clearance when used as a combination therapy.

  4. Intracellular trafficking of bio-nanocapsule-liposome complex: Identification of fusogenic activity in the pre-S1 region of hepatitis B virus surface antigen L protein.

    Science.gov (United States)

    Somiya, Masaharu; Sasaki, Yasuo; Matsuzaki, Takashi; Liu, Qiushi; Iijima, Masumi; Yoshimoto, Nobuo; Niimi, Tomoaki; Maturana, Andrés Daniel; Kuroda, Shun'ichi

    2015-08-28

    Bio-nanocapsules (BNCs) are a hollow nanoparticle consisting of about 100-nm liposome (LP) embedding about 110 molecules of hepatitis B virus (HBV) surface antigen (HBsAg) L protein as a transmembrane protein. Owing to the human hepatocyte-recognizing domains on the N-terminal region (pre-S1 region), BNCs have recently been shown to attach and enter into human hepatic cells using the early infection mechanism of HBV. Since BNCs could form a complex with an LP containing various drugs and genes, BNC-LP complexes have been used as a human hepatic cell-specific drug and gene-delivery system in vitro and in vivo. However, the role of BNCs in cell entry and intracellular trafficking of payloads in BNC-LP complexes has not been fully elucidated. In this study, we demonstrate that low pH-dependent fusogenic activity resides in the N-terminal part of pre-S1 region (NPLGFFPDHQLDPAFG), of which the first FF residues are essential for the activity, and which facilitates membrane fusion between LPs in vitro. Moreover, BNC-LP complexes can bind human hepatic cells specifically, enter into the cells via clathrin-mediated endocytosis, and release their payloads mostly into the cytoplasm. Taken together, the BNC portion of BNC-LP complexes can induce membrane fusion between LPs and endosomal membranes under low pH conditions, and thereby facilitate the endosomal escape of payloads. Furthermore, the fusogenic domain of the pre-S1 region of HBsAg L protein may play a pivotal role in the intracellular trafficking of not only BNC-LP complexes but also of HBV. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Kidney disease in hepatitis B surface antigen-positive children: experience from a centre in south-west Nigeria and a review of the Nigerian literature.

    Science.gov (United States)

    Asinobi, Adanze O; Ademola, Adebowale D; Okolo, Clement A; Adepoju, Adedayo A; Samuel, Susan M; Hoy, Wendy E

    2018-02-01

    Kidney disease is an important extra-hepatic manifestation of hepatitis B virus (HBV) infection. However, there is paucity of recent literature on kidney disease in children and adolescents with HBV infection from several parts of sub-Saharan Africa including Nigeria. To review the pattern of kidney disease in hepatitis B surface antigen (HBsAg)-positive children and adolescents seen at a tertiary hospital in south-west Nigeria. A retrospective study was undertaken of HBsAg-seropositive children with kidney disease managed at University College Hospital, Ibadan, from January 2004 to December 2015. Patients were identified from the paediatric nephrology unit admissions and the renal histology registers. 24 children and adolescents were studied, 17 of whom were male (70.8%), and the median age was 10.0 years (range 3-15). Ten (41.7%) had nephrotic syndrome, five (20.8%) had non-nephrotic glomerulonephritis, five (20.8%) were in end-stage renal disease (ESRD), including a patient with posterior urethral valves, and four had acute kidney injury secondary to acute tubular necrosis. Renal histology was available for 10 patients: nine had nephrotic syndrome associated with minimal change disease in six, focal segmental glomerulosclerosis in two and one had membanoproliferative glomerulonephritis. The patient with non-nephrotic glomerulonephritis had diffuse global sclerosis. The pattern of kidney disease in HBV-positive children demonstrated a predominance of nephrotic syndrome, followed by non-nephrotic glomerulonephritis, ESRD and acute kidney injury. Better diagnostic facilities and treatment are required. Prevention of HBV infection by universal childhood immunisation is the ultimate goal.

  6. Screening of pregnant women for hepatitis B virus surface antigen (HBsAg) and subsequent management, Qiandongnan prefecture, Guizhou, China, 2010.

    Science.gov (United States)

    Huang, Yan; Li, Leilei; Sun, Xiaohong; Lu, Mei; Liu, Huihui; Tang, Guangpeng; Wang, Dingming; Hutin, Yvan J

    2013-12-27

    In China, in 2010, a high proportion of pregnant women were tested for hepatitis B surface antigen (HBsAg). However, the preventive actions taken following screening were unclear. We followed up infants who were born to HBsAg positive mothers to describe the management that took place after screening. We selected eight counties with a probability proportional to population size in the Qiandongnan prefecture, Guizhou province. In each county, we selected a hospital at random. In each hospital, we (a) reviewed records to estimate the proportion of pregnant women who had been screened for HBsAg in 2010 and (b) sampled 10 screened women at random to assess management after one year in 2011. We calculated proportions and confidence intervals (CI) using standard formulae. Among the 7232 women who delivered in 2010 in the 8 hospitals, 98% (95% CI: 97%-99%) had been tested for HBsAg. Among 82 HBsAg women sampled for follow-up, 45 (55%; 95% CI: 44%-65%) knew they had been tested during pregnancy and 60 (73%; 95% CI: 63%-82%) knew they were HBsAg positive. The 82 infants had received three doses of hepatitis B vaccines and 79 (96%; 95% CI: 90%-99%) had received the first dose within 24h. However, only 11 infants (13%; 95% CI: 9%-25%) had received HBIG in addition to hepatitis B vaccine and 16 (20%; 95% CI: 12%-29%) had been tested for HBsAg upon completion of the vaccine series as part of their routine management. HBsAg testing of pregnant women was common in Qiandongnan, Guizhou, but post-screening management was limited. There is a need to ensure continuity of care through engaging women in HBsAg testing and following up infants with comprehensive management, including immunoprophylaxis and serological testing. Copyright © 2013 Elsevier Ltd. All rights reserved.

  7. Quantification of hepatitis B surface antigen with the novel DiaSorin LIAISON XL Murex HBsAg Quant: correlation with the ARCHITECT quantitative assays.

    Science.gov (United States)

    Burdino, Elisa; Ruggiero, Tina; Proietti, Alex; Milia, Maria Grazia; Olivero, Antonella; Caviglia, Gian Paolo; Marietti, Milena; Rizzetto, Mario; Smedile, Antonina; Ghisetti, Valeria

    2014-08-01

    Recent technologic innovations allow for quantitative assessment of hepatitis B surface antigen (HBsAg) levels in serum; this has been used to monitor the course of chronic HBV hepatitis (CHB) and predict treatment response. LIAISON-XL Murex HBsAg Quant assay (DiaSorin, Saluggia, I) is the newest immunoassay CE approved to quantify HBsAg. To compare LIAISON-XL performances with ARCHITECT-QT HBsAg (Abbott Diagnostics, IL, USA), as reference test. Sequential serum samples (n=152) from 14 HBe-negative patients with CHB, the majority of them infected by HBV genotype D undergoing antiviral treatment, were retrospectively tested with both assays. The 2nd WHO Standard 00/588 for HBsAg was used as reference. LIAISON-XL and ARCHITECT-QT correlated by r=0.95, pARCHITECT-QT. Median baseline HBsAg level was similar with the two methods before antiviral treatment, throughout fluctuations of HBsAg level in treatment non-responders and during the decrease of HBsAg titer in treatment responders. Correlation between HBsAg levels and HBV DNA was statistically significant for both the two immunoassays (LIAISON-XL: r=0.4988, 95% CI: 0.3452-0.6264, pARCHITECT-QT: r=0.480, 95% CI: 0.3233-0.6111, pARCHITECT-QT was high. LIAISON-XL accurately quantified HBsAg in clinical samples at baseline or during antiviral therapy; it can be applied for HBsAg quantification in clinical practice and decision making in CHB. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. Comparison of the elecsys HBsAg II assay and the architect assay for quantification of hepatitis B surface antigen in chronic hepatitis B patients.

    Science.gov (United States)

    Liao, Chen-Chih; Hsu, Chao-Wei; Gu, Po-Wen; Yeh, Chau-Ting; Lin, Shi-Ming; Chiu, Cheng-Tang

    2015-01-01

    Hepatitis B virus (HBV) infection is one of the infections with a highest prevalence in Taiwan. The most important marker is hepatitis B surface antigen (HBsAg). Using the new generation of HBsAg quantitative assay, HBsAg level may have good correlation with viral activity during different phases of chronic hepatitis B virus infection. This study was conducted to compare two assays of HBsAg level to find if the same results are obtained in HBsAg quantification in treatment-naïve and on-treatment chronic hepatitis B patients. Between March 2012 and June 2012, 90 patients with chronic hepatitis B (68 males and 22 females) were assessed using Abbott Architect HBsAg QT and Roche Elecsys HBsAg II assay. HBV DNA was detected by Roche COBAS TaqMan instrument. HBsAg level measured with Elecsys and Architect assays correlated well in untreated patients (n = 53, γs = 0.997) and on-treatment patients (n = 37, γs = 0.988). Bland-Altman analyses of the discrepancies in HBsAg levels showed a bias of -4.2% in untreated patients and -6.2% in on-treatment patients. Patients with HBeAg-postive chronic hepatitis B had higher HBsAg level than the ones who were HBeAg negative, and both showed statistical differences. Further, HBV DNA concentration analysis also showed higher viral concentration in HBeAg-positive patients, but it revealed no statistical difference. There is a significant correlation between Abbott Architect HBsAg QT assay and Roche Elecsys HBsAg II assay. Moreover, HBsAg quantification may potentially provide complementary information about the deduction of the natural course in chronic hepatitis B infection.

  9. Anisotropic In Situ-Coated AuNPs on Screen-Printed Carbon Surface for Enhanced Prostate-Specific Antigen Impedimetric Aptasensor

    Science.gov (United States)

    Do, Tram T. N.; Van Phi, Toan; Nguy, Tin Phan; Wagner, Patrick; Eersels, Kasper; Vestergaard, Mun'delanji C.; Truong, Lien T. N.

    2017-06-01

    An impedimetric aptasensor has been used to study the effect of charge transfer on the binding of prostate-specific antigen (PSA) to its aptamer. Full understanding of this mechanism will be beneficial to further improve its sensitivity for PSA detection in human semen at physiologically relevant concentrations. Bare gold electrodes (SPAuEs) and gold nanoparticles (AuNPs)-coated screen-printed carbon ink electrodes (AuNPs/SPCEs) were coated with aptamer solution at various concentrations and the sensor response to increasing PSA concentration in buffer solution examined. AuNPs were deposited onto carbon electrodes in 10 cycles. AuNPs/SPCEs were then coated with a self-assembled monolayer (SAM) of 16-mercaptohexadecanoic acid prior to aptamer immobilization at dose of 5 μg mL-1. The results indicate that anisotropic AuNPs/SPCEs outperform bare gold electrodes in terms of decreased amount of aptamer bunches as well as the number of intermediate PSA-aptamer complexes formed on the electrode surface. The key finding is that the fabricated aptasensor is sensitive enough [limit of detection (LoD) 1.95 ng mL-1] for early diagnosis of prostate cancer and displays linear response in the physiologically relevant concentration range (0 ng mL-1 to 10 ng mL-1), as shown by the calibration curve of the relative change in electron transfer resistance (Δ R CT) versus PSA concentration when aptamer/SAM/AuNPs/SPCEs were exposed to buffer containing PSA at different concentrations.

  10. Comparison of the binding properties of the mushroom Marasmius oreades lectin and Griffonia simplicifolia I-B isolectin to alphagalactosyl carbohydrate antigens in the surface phase.

    Science.gov (United States)

    Kirkeby, Svend; Winter, Harry C; Goldstein, Irwin J

    2004-05-01

    The binding of two alpha-galactophilic lectins, Marasmius oreades agglutinin (MOA), and Griffonia simplicifolia I isolectin B(4) (GS I-B(4)) to neoglycoproteins and natural glycoproteins were compared in a surface phase assay. Neoglycoproteins carrying various alpha-galactosylated glycans and laminin from basement membrane of mouse sarcoma that contains the xenogenic Galalpha1-3Gal1-4GlcNAc epitope were immobilized in microtiter plate wells and lectin binding determined with an enzyme-linked assay. After 24 h of incubation, MOA had higher affinity for the xenogenic pentasaccharide (Galalpha1-3Gal1-4GlcNAcbeta1-3Galbeta1-4Glc) than for the Galalpha-monosaccharide. The binding properties of MOA and GS I-B(4) to the xenogenic disaccharide (Galalpha1-3Galbeta1) were comparable while the binding of MOA to the xenogenic pentasaccharide was much stronger than the binding of GS I-B(4) to the same epitope. Non-xenogenic disaccharide-coupled neoglycoproteins having galactose end groups linked alpha1-2 or alpha1-4 to Gal or linked alpha1-3 to GalNAc bound very weakly to MOA, whereas GS I-B(4) recognized all of these disaccharides with similarly high affinity. MOA also showed high affinity for laminin. The results indicate that the Marasmius oreades lectin has nearly the same affinities as does GS I-B(4) for the simple xenogenic carbohydrate antigens, but MOA has greater affinity for the pentasaccharide and is far more specific in its binding preferences than the Griffonia lectin. Copyright Blackwell Munksgaard, 2004

  11. Impact of "a" determinant mutations on detection of hepatitis B surface antigen (HBsAg) in HBV strains from Chinese patients with occult hepatitis B.

    Science.gov (United States)

    Huang, Xiangyan; Ma, Chenyun; Zhang, Qiang; Shi, Qingfen; Huang, Tao; Liu, Chao; Li, Jie; Hollinger, F Blaine

    2017-10-01

    This study was designed to detect mutations that occur within the "a" determinant in the S gene of the hepatitis B virus (HBV) in patients with occult hepatitis B (OHB), and to analyze the influence of these mutations on expression and reactivity of the hepatitis B surface antigen (HBsAg). Twenty-three certified OHB samples were compared to 32 HBsAg positive samples from patients with chronic hepatitis B. The median HBV DNA levels in the OHB group were significantly lower than those in the control group (P < 0.0001). Mutations within the "a" determinant were analyzed by gene amplification and sequencing. This revealed mixed infections in which clones within a sample displayed either different mutations or mutations in association with clones that exhibited wild type amino acid patterns. Sequencing analysis also showed a significant difference between the proportions of amino acid mutations observed in the OHB and control groups. Seven recombinant S (rS) proteins with corresponding OHB mutations and three wild type alleles were expressed and purified in the Pichia pastoris expression system to preserve conformational attributes, and their reactivity analyzed using six commercial HBsAg assays. The OHB sera were HBsAg nonreactive while the rS proteins with corresponding OHB mutations were universally reactive. Thus, we postulate that the reduced binding affinity between mutated HBsAg and its antibody may not be as important in defining OHB as is the effect of specific mutations in the preS/S region of the genome that affect the synthesis and secretion of the S protein and/or the virion. © 2017 Wiley Periodicals, Inc.

  12. Enhancing recovery of recombinant hepatitis B surface antigen in lab-scale and large-scale anion-exchange chromatography by optimizing the conductivity of buffers.

    Science.gov (United States)

    Mojarrad Moghanloo, Gol Mohammad; Khatami, Maryam; Javidanbardan, Amin; Hosseini, Seyed Nezamedin

    2018-01-01

    In biopharmaceutical science, ion-exchange chromatography (IEC) is a well-known purification technique to separate the impurities such as host cell proteins from recombinant proteins. However, IEC is one of the limiting steps in the purification process of recombinant hepatitis B surface antigen (rHBsAg), due to its low recovery rate (buffers are easy-to-control parameters which can play a major role in optimizing the process and increasing the recovery. Thus, we investigated the effects of ionic strengths of buffers on rHBsAg recovery via adjusting Tris-HCl and NaCl concentrations. Increasing the conductivity of equilibration (Eq.), washing (Wash.) and elution (Elut.) buffers from their initial values of 1.6 mS/cm, 1.6 mS/cm, and 7.0 mS/cm to 1.6 mS/cm, 7 mS/cm and 50 mS/cm, respectively yielded an average recovery rate of 82% in both lab-scale and large-scale weak anion-exchange chromatography without any harsh effect on the purity percentage of rHBsAg. The recovery enhancement via increasing the conductivity of Eq. and Wash. buffers can be explained by their roles in reducing the binding strength and aggregation of retained particles in the column. Moreover, further increase in the salt concentration of Elut. Buffer could substantially promote the ion exchange process and the elution of retained rHBsAg. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. The Plasmodium falciparum STEVOR multigene family mediates antigenic variation of the infected erythrocyte.

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    Makhtar Niang

    2009-02-01

    Full Text Available Modifications of the Plasmodium falciparum-infected red blood cell (iRBC surface have been linked to parasite-associated pathology. Such modifications enable the parasite to establish long-lasting chronic infection by evading antibody mediate immune recognition and splenic clearance. With the exception of the well-demonstrated roles of var-encoded PfEMP1 in virulence and immune evasion, the biological significance of other variant surface antigens (rif and stevor is largely unknown. While PfEMP1 and RIFIN have been located on the iRBC surface, recent studies have located STEVOR at the iRBC membrane where it may be exposed on the erythrocyte surface. To investigate the role of STEVOR in more detail, we have developed antibodies against two putative STEVOR proteins and used a combination of indirect immunofluorescence assays (IFA, live IFA, flow cytometry, as well as agglutination assays, which enable us to demonstrate that STEVOR is clonally variant at the surface of schizont stage parasites. Crucially, expression of different STEVOR on the surface of the iRBC changes the antigenic property of the parasite. Taken together, our data for the first time demonstrate that STEVOR plays a role in creating antigenic diversity of schizont stage parasites, thereby adding additional complexity to the immunogenic properties of the iRBC. Furthermore, it clearly demonstrates that to obtain a complete understanding of how parasite-induced pathology is linked to variation on the surface of the iRBC, focusing the interactions of multiple multigene families needs to be considered.

  14. Co-delivery of polyinosinic:polycytidylic acid and flagellin by poly(lactic-co-glycolic acid) MPs synergistically enhances immune response elicited by intranasally delivered hepatitis B surface antigen

    OpenAIRE

    Dai, Xiaojing; He, Jintian; Zhang, Ruxia; Wu, Guanghao; Xiong, Fangfang; Zhao, Baohua

    2017-01-01

    Xiaojing Dai, Jintian He, Ruxia Zhang, Guanghao Wu, Fangfang Xiong, Baohua Zhao College of Life Science, Hebei Normal University, Shijiazhuang City, Hebei Province, People’s Republic of China Abstract: The aim of the present work was to investigate the synergistic effect between toll-like receptor (TLR) 3 ligand polyinosinic:polycytidylic acid (pI:C) and TLR5 ligand flagellin (FLN) on immune responses induced by nasally delivered hepatitis B virus surface antigen (HBsAg). Mannan a...

  15. 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.

  16. Natural selection promotes antigenic evolvability.

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    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

  17. Low-dose adenovirus vaccine encoding chimeric hepatitis B virus surface antigen-human papillomavirus type 16 E7 proteins induces enhanced E7-specific antibody and cytotoxic T-cell responses.

    Science.gov (United States)

    Báez-Astúa, Andrés; Herráez-Hernández, Elsa; Garbi, Natalio; Pasolli, Hilda A; Juárez, Victoria; Zur Hausen, Harald; Cid-Arregui, Angel

    2005-10-01

    Induction of effective immune responses may help prevent cancer progression. Tumor-specific antigens, such as those of human papillomaviruses involved in cervical cancer, are targets with limited intrinsic immunogenicity. Here we show that immunization with low doses (10(6) infectious units/dose) of a recombinant human adenovirus type 5 encoding a fusion of the E7 oncoprotein of human papillomavirus type 16 to the carboxyl terminus of the surface antigen of hepatitis B virus (HBsAg) induces remarkable E7-specific humoral and cellular immune responses. The HBsAg/E7 fusion protein assembled efficiently into virus-like particles, which stimulated antibody responses against both carrier and foreign antigens, and evoked antigen-specific kill of an indicator cell population in vivo. Antibody and T-cell responses were significantly higher than those induced by a control adenovirus vector expressing wild-type E7. Such responses were not affected by preexisting immunity against either HBsAg or adenovirus. These data demonstrate that the presence of E7 on HBsAg particles does not interfere with particle secretion, as it occurs with bigger proteins fused to the C terminus of HBsAg, and results in enhancement of CD8(+)-mediated T-cell responses to E7. Thus, fusion to HBsAg is a convenient strategy for developing cervical cancer therapeutic vaccines, since it enhances the immunogenicity of E7 while turning it into an innocuous secreted fusion protein.

  18. Dynamic interaction of /sup 111/indium-labeled monoclonal antibodies with surface antigens of solid tumors visualized in vivo by external scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, K.M.; Keenan, A.M.; Frincke, J.; David, G.; Pearson, J.; Oldham, R.K.; Morgan, A.C. Jr.

    1986-05-01

    Two /sup 111/indium-labeled murine monoclonal antibodies (MoAb), D3 and 9.2.27, directed to tumor antigens of L-10 hepatocarcinoma and human melanoma, respectively, selectively localized antigen-positive target cells in guinea pigs and nude mice. The fate of MoAb differed in the two antigen-antibody systems after reacting with their corresponding tumor antigens in vivo as reflected by patterns of distribution and turnover in vivo. The 9.2.27 localized in melanoma xenograft in nude mice after intravenous administration with slow loss from tumor but more rapid loss from normal tissues and thus demonstrated optimal imaging of small tumors (approximately equal to 5 mm) between 3 and 6 days after injection of the radiolabeled antibody. In contrast, D3 demonstrated a biphasic localization in guinea pig L-10 hepatocarcinoma with a maximal activity on the 2d day after administration and showed rapid loss from both tumor and normal tissues. Nonspecific localization of antibodies in liver and in kidney was found both in syngeneic (nude mice) and xenogeneic (guinea pig) hosts but was more pronounced in the xenogeneic species. These results indicate that the nature of the antigen-antibody interaction may be of importance in selecting MoAb for both diagnosis and therapy of malignant diseases.

  19. The microRNAs in an ancient protist repress the variant-specific surface protein expression by targeting the entire coding sequence.

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    Ashesh A Saraiya

    2014-02-01

    Full Text Available microRNAs (miRNA have been detected in the deeply branched protist, Giardia lamblia, and shown to repress expression of the family of variant-specific surface proteins (VSPs, only one of which is expressed in Giardia trophozoite at a given time. Three next-generation sequencing libraries of Giardia Argonaute-associated small RNAs were constructed and analyzed. Analysis of the libraries identified a total of 99 new putative miRNAs with a size primarily in the 26 nt range similar to the size previously predicted by the Giardia Dicer crystal structure and identified by our own studies. Bioinformatic analysis identified multiple putative miRNA target sites in the mRNAs of all 73 VSPs. The effect of miRNA target sites within a defined 3'-region were tested on two vsp mRNAs. All the miRNAs showed partial repression of the corresponding vsp expression and were additive when the targeting sites were separately located. But the combined repression still falls short of 100%. Two other relatively short vsp mRNAs with 15 and 11 putative miRNA target sites identified throughout their ORFs were tested with their corresponding miRNAs. The results indicate that; (1 near 100% repression of vsp mRNA expression can be achieved through the combined action of multiple miRNAs on target sites located throughout the ORF; (2 the miRNA machinery could be instrumental in repressing the expression of vsp genes in Giardia; (3 this is the first time that all the miRNA target sites in the entire ORF of a mRNA have been tested and shown to be functional.

  20. Antigenicity and immunogenicity of a novel Plasmodium vivax circumsporozoite derived synthetic vaccine construct

    DEFF Research Database (Denmark)

    Céspedes, Nora; Jiménez, Eliécer; Lopez-Perez, Mary

    2014-01-01

    BACKGROUND: The circumsporozoite (CS) protein is a major malaria sporozoite surface antigen currently being considered as vaccine candidate. Plasmodium vivax CS (PvCS) protein comprises a dimorphic central repeat fragment flanked by conserved regions that contain functional domains involved...... and the two natural repeat variant regions known as VK210 and VK247. We studied the natural immune response to this region in human sera from different malaria-endemic areas and its immunogenicity in mice. RESULTS: PvNR1R2 was more frequently recognized by sera from Papua New Guinea (PNG) (83%) than...

  1. Enrichment and purging of human embryonic stem cells by detection of cell surface antigens using the monoclonal antibodies TG30 and GCTM-2.

    Science.gov (United States)

    Polanco, Juan Carlos; Wang, Bei; Zhou, Qi; Chy, Hun; O'Brien, Carmel; Laslett, Andrew L

    2013-12-06

    Human embryonic stem cells (hESC) can self-renew indefinitely in vitro, and with the appropriate cues can be induced to differentiate into potentially all somatic cell lineages. Differentiated hESC derivatives can potentially be used in transplantation therapies to treat a variety of cell-degenerative diseases. However, hESC differentiation protocols usually yield a mixture of differentiated target and off-target cell types as well as residual undifferentiated cells. For the translation of differentiated hESC-derivatives from the laboratory to the clinic, it is important to be able to discriminate between undifferentiated (pluripotent) and differentiated cells, and generate methods to separate these populations. Safe application of hESC-derived somatic cell types can only be accomplished with pluripotent stem cell-free populations, as residual hESCs could induce tumors known as teratomas following transplantation. Towards this end, here we describe a methodology to detect pluripotency associated cell surface antigens with the monoclonal antibodies TG30 (CD9) and GCTM-2 via fluorescence activated cell sorting (FACS) for the identification of pluripotent TG30(Hi)-GCTM-2(Hi) hESCs using positive selection. Using negative selection with our TG30/GCTM-2 FACS methodology, we were able to detect and purge undifferentiated hESCs in populations undergoing very early-stage differentiation (TG30(Neg)-GCTM-2(Neg)). In a further study, pluripotent stem cell-free samples of differentiated TG30(Neg)-GCTM-2(Neg) cells selected using our TG30/GCTM-2 FACS protocol did not form teratomas once transplanted into immune-compromised mice, supporting the robustness of our protocol. On the other hand, TG30/GCTM-2 FACS-mediated consecutive passaging of enriched pluripotent TG30(Hi)-GCTM-2(Hi) hESCs did not affect their ability to self-renew in vitro or their intrinsic pluripotency. Therefore, the characteristics of our TG30/GCTM-2 FACS methodology provide a sensitive assay to obtain highly

  2. A new trivalent SnSAG surface antigen chimera for efficient detection of antibodies against Sarcocystis neurona and diagnosis of equine protozoal myeloencephalitis.

    Science.gov (United States)

    Yeargan, Michelle; de Assis Rocha, Izabela; Morrow, Jennifer; Graves, Amy; Reed, Stephen M; Howe, Daniel K

    2015-05-01

    Enzyme-linked immunosorbent assays (ELISAs) based on the SnSAG surface antigens of Sarcocystis neurona provide reliable detection of infection by the parasite. Moreover, accurate serodiagnosis of equine protozoal myeloencephalitis (EPM) is achieved with the SnSAG ELISAs by measuring antibodies in serum and cerebrospinal fluid (CSF) to reveal active infection in the central nervous system. Two independent ELISAs based on recombinant (r)SnSAG2 or a chimeric fusion of SnSAG3 and SnSAG4 (rSnSAG4/3) are currently used together for EPM serodiagnosis to overcome varied antibody responses in different horses. To achieve reliable antibody detection with a single ELISA instead of 2 separate ELISAs, rSnSAG2 was fused with rSnSAG4/3 into a single trivalent protein, designated rSnSAG2/4/3. Paired serum and CSF from 163 horses were tested with all 3 ELISAs. When the consensus antibody titers obtained with the rSnSAG2 and rSnSAG4/3 ELISAs were compared to the single SAG2/4/3 ELISA titers, Spearman rank correlation coefficients of ρ = 0.74 and ρ = 0.90 were obtained for serum and CSF, respectively, indicating strong agreement between the tests. When the rSnSAG2 and rSnSAG4/3 consensus serum-to-CSF titer ratio was compared to the rSnSAG2/4/3 serum-to-CSF titer ratio, the Spearman correlation coefficient was ρ = 0.87, again signifying strong agreement. Importantly, comparing the diagnostic interpretation of the serum-to-CSF titer ratios yielded a Cohen kappa value of 0.77. These findings suggest that the single ELISA based on the trivalent rSnSAG2/4/3 will provide serologic and diagnostic results that are highly comparable to the consensus of the 2 independent ELISAs based on rSnSAG2 and rSnSAG4/3. © 2015 The Author(s).

  3. Long-term effect of interferon plus ribavirin on hepatitis B surface antigen seroclearance in patients dually infected with hepatitis B and C viruses.

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    Ming-Lun Yeh

    Full Text Available BACKGROUND: Interferon-α/ribavirin combination therapy might promote hepatitis B surface antigen (HBsAg seroclearance in patients dually infected with hepatitis B and C viruses (HBV/HCV, but the long-term effect remains unclear. We aimed to investigate the rate of and the factors associated with HBsAg seroclearance during long-term follow-up after interferon-α/ribavirin combination therapy in HBV/HCV dually-infected patients. METHODOLOGY/PRINCIPAL FINDINGS: Eighty-one patients who received interferon-α/ribavirin combination therapy for 24 weeks with a follow-up period of >24 weeks were enrolled. HBV serological markers and HBV DNA were determined every 6 months. Early and late HBsAg seroclearance were defined as HBsAg loss in less or more than 6 months after end-of-treatment, respectively. Fifteen (18.5% patients had HBsAg seroclearance during a mean follow-up period of 3.4 (0.5-5.1 years. The 5-year cumulative incidence was 25.6%. Baseline cirrhosis and HBV DNA negativity 1 year after end-of-treatment were independently predictive of HBsAg seroclearance with an odds ratio (OR, 95% confidence intervals (CI of 16.6, 1.8-153 and 9.2, 1.4-62.1, respectively, by Cox regression hazard analysis. Four patients developed early and 11 developed late HBsAg seroclearance, respectively. Cox regression hazard analysis showed no factor was associated with early HBsAg seroclearance, whilst HBV DNA negativity 1 year after end-of-treatment was the only significant factor predicting late HBsAg loss (OR, 43.0; CI, 2.5-745. Five patients had HBsAg seroconversion with a 5-year cumulative incidence of 8.3%. HBV DNA negativity at baseline and one year after EOT had a trend for HBsAg seroconversion. HCV response did not correlate to HBsAg loss. CONCLUSIONS: We demonstrated that interferon-α/ribavirin had long-term effect on HBsAg seroclearance in dually HBV/HCV-infected patients. Baseline cirrhosis and seroclearance of HBV DNA 1 year after end-of-treatment were

  4. Prenatal Screening for and Prevalence of Hepatitis B Surface Antigen in Pregnant Women and Prevention of Transmission to Infants Born to Infected Mothers-Guam, 2014.

    Science.gov (United States)

    Abara, Winston E; Cha, Susan; Malik, Tasneem; DeSimone, Mia S; Schillie, Sarah; Collier, Melissa; Schumann, Bernadette; Klemme, Michael; Kamb, Mary

    2017-09-05

    Perinatal transmission is the major mode of hepatitis B virus (HBV) transmission and drives HBV endemicity in the US territory of Guam. We assessed correlates of prenatal hepatitis B surface antigen (HBsAg) screening and HBsAg positivity among pregnant women and evaluated the care of infants of HBsAg-positive women. Demographic and clinical data were abstracted from the maternal medical records of 966 randomly selected live infants born in 2014. Frequencies were calculated, and prevalence ratios (PRs) and 95% confidence intervals (CIs) were estimated using Poisson regression. Among the mothers of the 966 infants, 78.2% were Pacific Islanders, 56.9% were >25 years old (born before universal infant hepatitis B vaccination in Guam), 89.0% received prenatal care (PNC), 96.7% underwent prenatal HBsAg screening, and 2.0% were HBsAg positive. Approximately 15% of the women who did not have PNC were not screened for HBsAg. Receipt of PNC was associated with HBsAg screening (adjusted PR, 1.13 [95% CI, 1.04-1.23]), and HBsAg positivity was associated with a maternal age of >25 years (adjusted PR, 6.80 [95% CI, 1.32-35.08]). All 18 infants of the HBsAg-positive mothers received hepatitis B vaccine, and 17 (94.4%) received hepatitis B immunoglobulin. Although the prenatal HBsAg screening prevalence in this sample was high, the maternal HBsAg prevalence among women in this sample was more than 14 times and 2 times the prevalence among US-born Pacific Islander/Asian women and all women in the continental United States, respectively. Improving access to PNC, ensuring that all pregnant women in Guam (especially those born before universal hepatitis B vaccination) are screened for HBsAg, and adopting postexposure prophylaxis for infants of HBsAg-positive mothers as standard clinical practice are important for preventing perinatal HBV transmission and reducing HBV endemicity.

  5. Hepatitis B surface antigen quantity positively correlates with plasma levels of microRNAs differentially expressed in immunological phases of chronic hepatitis B in children.

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    Thilde Nordmann Winther

    Full Text Available BACKGROUND AND AIM: Children with chronic hepatitis B (CHB are at high risk of progressive liver disease. It is suggested that a newly-identified panel of 16 microRNAs is important in the pathogenesis of CHB in children. Subviral hepatitis B surface antigen (HBsAg particles are produced in large excess over infectious virions. Interestingly, circulating HBsAg particles have been shown to carry microRNAs. A thorough characterisation of the identified microRNAs and HBsAg over time in plasma from children with CHB may provide useful information about the natural course of childhood CHB. PATIENTS AND METHODS: A cohort of 42 children with CHB was followed over time. Three to five blood samples were obtained from each child at minimum intervals of half a year; in total 180 blood samples. Plasma levels of the 16 microRNAs previously identified were analysed by quantitative real-time polymerase-chain-reaction. Plasma HBsAg was quantified using ARCHITECT® HBsAg assay. RESULTS: The presence of 14/16 plasma microRNAs in children with CHB was confirmed. All 14 microRNAs were significantly differentially expressed in different immunological phases of the disease. MicroRNA plasma levels were highest in immune-tolerant children, lower in immune-active children, and reached the lowest values in immune-inactive children, p<0.001. Plasma levels of four microRNAs decreased significantly over time in immune-tolerant and immune-active children whereas the microRNA plasma levels were stable in immune-inactive children, p<0.004. HBsAg quantity was positively correlated with plasma levels of 11/14 microRNAs, p<0.004. CONCLUSION: This is the first study to characterise plasma microRNAs and HBsAg over time in children with CHB. Our data suggest that plasma levels of selected microRNAs and HBsAg are inversely correlated with immunological control of CHB in children. Further studies are, however, needed to advance the understanding of microRNAs and HBsAg in the

  6. Transplacentally acquired maternal antibody against hepatitis B surface antigen in infants and its influence on the response to hepatitis B vaccine.

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    Zhiqun Wang

    Full Text Available BACKGROUND: Passively acquired maternal antibodies in infants may inhibit active immune responses to vaccines. Whether maternal antibody against hepatitis B surface antigen (anti-HBs in infants may influence the long-term immunogenicity of hepatitis B vaccine remains unknown. METHODOLOGY/PRINCIPAL FINDINGS: Totally 338 pairs of mothers and children were enrolled. All infants were routinely vaccinated against hepatitis B based on 0-, 1- and 6-month schedule. We characterized the transplacental transfer of maternal anti-HBs, and compared anti-HBs response in children of mothers with or without anti-HBs. In a prospective observation, all 63 anti-HBs positive mothers transferred anti-HBs to their infants; 84.1% of the infants had higher anti-HBs concentrations than their mothers. One and half years after vaccination with three doses of hepatitis B vaccine, the positive rate and geometric mean concentration (GMC of anti-HBs in 32 infants with maternal anti-HBs were comparable with those in 32 infants without maternal antibody (90.6% vs 87.5%, P = 0.688, and 74.5 vs 73.5 mIU/ml, P = 0.742, respectively. In a retrospective analysis, five and half years after vaccination with three doses vaccine, the positive rates of anti-HBs in 88 children of mothers with anti-HBs ≥1000 mIU/ml, 94 children of mothers with anti-HBs 10-999 mIU/ml, and 61 children of mothers with anti-HBs <10 mIU/ml were 72.7%, 69.2%, and 63.9% (P = 0.521, respectively; anti-HBs GMC in these three groups were 38.9, 43.9, and 31.7 mIU/ml (P = 0.726, respectively. CONCLUSIONS/SIGNIFICANCE: The data demonstrate that maternal anti-HBs in infants, even at high concentrations, does not inhibit the long-term immunogenicity of hepatitis B vaccine. Thus, current hepatitis B vaccination schedule for infants will be still effective in the future when most infants are positive for maternal anti-HBs due to the massive vaccination against hepatitis B.

  7. Association of oral lichen planus with hepatitis C virus, surface antigen of hepatitis B virus, and diabetes: A clinical and biochemical study

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    Pavani Donempudi

    2016-01-01

    Full Text Available Background: Oral lichen planus is an inflammatory mucocutaneous disease commonly encountered in the dental clinic. The etiology of oral lichen planus is still unknown. The probable factors associated with oral lichen planus include anxiety, trauma, malnutrition, infection, and autoimmunity. Objectives: The aim of the study was to evaluate the serum levels of fasting and postprandial blood sugar levels, surface antigen of hepatitis B virus (HBsAg, and hepatitis C virus (HCV autoantibodies in oral lichen planus patients and controls, and to assess the association of oral lichen planus with diabetes, HCV, and HBsAg. Study Design: The study included a total of 25 oral lichen planus patients. Twenty-five individuals who did not have the above clinical conditions were also included in the study and comprised the control group. Both the study and control groups consisted of both sexes in the age group of 20–70 years. Patients were clinically diagnosed as oral lichen planus after obtaining a detailed history and was later confirmed by histopathologic examination. Materials and Methods: Blood samples were collected from both the oral lichen planus patients and controls. Biochemical analysis of blood glucose levels, i.e., fasting and postprandial blood glucose levels, HBsAg, hepatitis C autoantibodies were done by enzyme-linked immunosorbent assay. Results: Blood glucose levels, i.e., fasting and postprandial blood glucose levels, were found to be statistically increased in oral lichen planus patients when compared with controls. There was no significant change in HCV autoantibodies and HBsAg. The results of the present study also showed a significant association of oral lichen planus with diabetes. However, there was no significant association of oral lichen planus with HCV and HBsAg in patients when compared with controls. Conclusion: There was a direct relation of oral lichen planus with diabetes whereas there was no significant association with HCV

  8. Inhibition of Viability, Proliferation, Cytokines Secretion, Surface Antigen Expression, and Adipogenic and Osteogenic Differentiation of Adipose-Derived Stem Cells by Seven-Day Exposure to 0.5 T Static Magnetic Fields

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    Jian Wang

    2016-01-01

    Full Text Available After seven-day exposure to 0.5-Tesla Static Magnetic Field (SMF, Adipose-derived Stem Cells (ASCs and those labeled by superparamagnetic iron oxide (SPIO nanoparticles were examined for viability by methyl thiazol tetrazolium (MTT assay, proliferation by cell counting and bromodeoxyuridine (BrdU incorporation, DNA integrity by single cell gel electrophoresis, surface antigen by flow cytometry analysis, and the expression of cytokines and genetic markers by reverse transcription-PCR and underwent adipogenic and osteogenic differentiation assessed by quantifying related specific genes expression. The SMF slightly reduced cell viability and proliferation and inhibited the expression of CD49d, CD54, and CD73 but did not damage DNA integrity. The SMF slightly downregulated the expression of cytokines including Vascular Endothelial Growth Factor (VEGF, Insulin-like Growth Factor-1 (IGF-1, Transforming Growth Factor Beta 1 (TGF-β1, genetic markers comprising Stem Cell Antigen-1 (Sca1, Octamer-4 (Oct-4, ATP-binding Cassette Subfamily B Member 1 (ABCB1, adipogenic marker genes containing Lipoprotein Lipase (LPL, Peroxisome Proliferator-Activated Receptor Gamma (PPAR-γ, and osteogenic marker genes including Secreted Phosphor-protein 1 (SPP1 and Osterix (OSX. Exposure to 0.5 T SMF for seven days inhibited viability, proliferation, surface antigen expression, cytokine secretion, stem cell genetic marker expression, and adipogenic and osteogenic differentiation but did not affect the DNA integrity in ASCs with or without SPIO labeling.

  9. Sequence-based antigenic change prediction by a sparse learning method incorporating co-evolutionary information.

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    Jialiang Yang

    Full Text Available Rapid identification of influenza antigenic variants will be critical in selecting optimal vaccine candidates and thus a key to developing an effective vaccination program. Recent studies suggest that multiple simultaneous mutations at antigenic sites accumulatively enhance antigenic drift of influenza A viruses. However, pre-existing methods on antigenic variant identification are based on analyses from individual sites. Because the impacts of these co-evolved sites on influenza antigenicity may not be additive, it will be critical to quantify the impact of not only those single mutations but also multiple simultaneous mutations or co-evolved sites. Here, we developed and applied a computational method, AntigenCO, to identify and quantify both single and co-evolutionary sites driving the historical antigenic drifts. AntigenCO achieved an accuracy of up to 90.05% for antigenic variant prediction, significantly outperforming methods based on single sites. AntigenCO can be useful in antigenic variant identification in influenza surveillance.

  10. Improved cell mediated immune responses after successful re-vaccination of non-responders to the hepatitis B virus surface antigen (HBsAg) vaccine using the combined hepatitis A and B vaccine.

    Science.gov (United States)

    Nyström, Jessica; Cardell, Kristina; Björnsdottir, Thora Björg; Fryden, Aril; Hultgren, Catharina; Sällberg, Matti

    2008-11-05

    We successfully re-vaccinated hepatitis B virus (HBV) vaccine non-responders using a double dose of the combined hepatitis A virus (HAV) and HBV vaccine. The hope was to improve priming of hepatitis B surface antigen (HBsAg)-specific cell mediated immune response (CMI) by an increased antigen dose and a theoretical adjuvant-effect from the local presence of a HAV-specific CMI. A few non-responders had a detectable HBsAg-specific CMI before re-vaccination. An in vitro detectable HBsAg-specific CMI was primed equally effective in non-responders (58%) as in first time vaccine recipients (68%). After the third dose a weak, albeit significant, association was observed between the magnitude of HBsAg-specific proliferation and anti-HBs levels. This regimen improves the priming of HBsAg-specific CMIs and antibodies.

  11. A mucosal IgA response, but no systemic antibody response, is evoked by intranasal immunisation of dogs with Echinococcus granulosus surface antigens iscoms.

    Science.gov (United States)

    Carol, H; Nieto, A

    1998-09-16

    The search for protective antigens of intestinal parasites is conditioned by the methodology used to induce a relevant local immune response against them. The present work describes the use of immuno stimulating complexes (iscoms) from tegumental antigens from protoscoleces (PSC) of the cestode Echinococcus granulosus as immunogens in dogs by the intranasal route. It also describes the evaluation of the immune response evoked at the antibody level (systemically and at a distant mucosal location) as well as at the level of antibody secreting cells in peripheral blood. Iscoms from both E. granulosus tegumental antigens and hen ovalbumin (OVA), given at 50 microg doses by intranasal route, evoked significant secretory IgA antibody responses detected in saliva. Specific IgA secreting cells in peripheral blood also increased 10-20-fold, although transiently, after primary and secondary stimulation, whereas specific IgG secreting cells in peripheral blood were only detected in some individuals after the second antigenic exposure. Generation of immune responses at a related mucosal site provides evidence of localised immunity. No significant increase in systemic antibody titers of either IgM, IgG or IgA isotype was detected in plasma as a result of the immunisation. This fact could reflect that the nasopharyngeal mucosal associated lymphoid tissue of dogs is more strictly compartmentalised than that of other mammals.

  12. Safety of currently licensed hepatitis B surface antigen vaccines in the United States, Vaccine Adverse Event Reporting System (VAERS), 2005-2015.

    Science.gov (United States)

    Haber, Penina; Moro, Pedro L; Ng, Carmen; Lewis, Paige W; Hibbs, Beth; Schillie, Sarah F; Nelson, Noele P; Li, Rongxia; Stewart, Brock; Cano, Maria V

    2018-01-25

    Currently four recombinant hepatitis B (HepB) vaccines are in use in the United States. HepB vaccines are recommended for infants, children and adults. We assessed adverse events (AEs) following HepB vaccines reported to the Vaccine Adverse Event Reporting System (VAERS), a national spontaneous reporting system. We searched VAERS for reports of AEs following single antigen HepB vaccine and HepB-containing vaccines (either given alone or with other vaccines), from January 2005 - December 2015. We conducted descriptive analyses and performed empirical Bayesian data mining to assess disproportionate reporting. We reviewed serious reports including reports of special interest. VAERS received 20,231 reports following HepB or HepB-containing vaccines: 10,291 (51%) in persons 18 years; for 1485 (7.3%) age was missing. Dizziness and nausea (8.4% each) were the most frequently reported AEs following a single antigen HepB vaccine: fever (23%) and injection site erythema (11%) were most frequent following Hep-containing vaccines. Of the 4444 (22%) reports after single antigen HepB vaccine, 303 (6.8%) were serious, including 45 deaths. Most commonly reported cause of death was Sudden Infant Death Syndrome (197). Most common non-death serious reports following single antigen HepB vaccines among infants aged 18 years. Most common vaccination error following single antigen HepB was incorrect product storage. Review current U.S.-licensed HepB vaccines administered alone or in combination with other vaccines did not reveal new or unexpected safety concerns. Vaccination errors were identified which indicate the need for training and education of providers on HepB vaccine indications and recommendations. Published by Elsevier Ltd.

  13. A splice variant of the two-pore domain potassium channel TREK-1 with only one pore domain reduces the surface expression of full-length TREK-1 channels.

    Science.gov (United States)

    Rinné, Susanne; Renigunta, Vijay; Schlichthörl, Günter; Zuzarte, Marylou; Bittner, Stefan; Meuth, Sven G; Decher, Niels; Daut, Jürgen; Preisig-Müller, Regina

    2014-08-01

    We have identified a novel splice variant of the human and rat two-pore domain potassium (K2P) channel TREK-1. The splice variant TREK-1e results from skipping of exon 5, which causes a frame shift in exon 6. The frame shift produces a novel C-terminal amino acid sequence and a premature termination of translation, which leads to a loss of transmembrane domains M3 and M4 and of the second pore domain. RT-PCR experiments revealed a preferential expression of TREK-1e in kidney, adrenal gland, and amygdala. TREK-1e was nonfunctional when expressed in Xenopus oocytes. However, both the surface expression and the current density of full-length TREK-1 were reduced by co-expression of TREK-1e. Live cell imaging in COS-7 cells transfected with GFP-tagged TREK-1e showed that this splice variant was retained in the endoplasmic reticulum (ER). Attachment of the C-terminus of TREK-1e to two different reporter proteins (Kir2.1 and CD8) led to a strong reduction in the surface expression of these fusion proteins. Progressive truncation of the C-terminus of TREK-1e in these reporter constructs revealed a critical region (amino acids 198 to 205) responsible for the intracellular retention. Mutagenesis experiments indicated that amino acids I204 and W205 are key residues mediating the ER retention of TREK-1e. Our results suggest that the TREK-1e splice variant may interfere with the vesicular traffic of full-length TREK-1 channels from the ER to the plasma membrane. Thus, TREK-1e might modulate the copy number of functional TREK-1 channels at the cell surface, providing a novel mechanism for fine tuning of TREK-1 currents.

  14. Expression and secretion of the Giardia duodenalis variant surface protein 9B10A by transfected trophozoites causes damage to epithelial cell monolayers mediated by protease activity.

    Science.gov (United States)

    Cabrera-Licona, Ariana; Solano-González, Eduardo; Fonseca-Liñán, Rocío; Bazán-Tejeda, Ma Luisa; Raúl Argüello-García; Bermúdez-Cruz, Rosa Ma; Ortega-Pierres, Guadalupe

    2017-08-01

    Giardia duodenalis is the protozoan parasite responsible for most cases of parasitic diarrhea worldwide. The pathogenic mechanisms of giardiasis have not yet been fully characterized. In this context parasite's excretory/secretory products have been related to the damage induced by the parasite on enterocytes. Among these is the Variable Surface Proteins (VSPs) family involved in antigenic variation and in the induction of protective response. In proteomic analyses carried out to identify the proteases with high molecular weight secreted by Giardia trophozoites during the initial phase of interaction with IEC-6 cell monolayers we identified the VSP9B10A protein. In silico bioinformatics analyses predicted a central region in residues 324-684 displaying the catalytic triad and the substrate binding pocket of cysteine proteases. The analysis of the effect of the VSP9B10A protein on epithelial cell monolayers using trophozoites that were transfected with a plasmid carrying the vsp9b10a gene sequence under the control of a constitutive promoter showed that transfected trophozoites expressing the VSP9B10A protein caused cytotoxic damages on IEC-6 and MDCK cell monolayers. This was characterized by loss of cell-cell contacts and cell detachment from the substrate while no damage was observed with trophozoites that did not express the VSP9B10A protein. The same cytotoxic effect was detected when IEC-6 cell monolayers were incubated only with supernatants from co-cultures of IEC-6 cell monolayers with VSP9B10A transfected trophozoites and this effect was not observed when transfected trophozoites were incubated with a monospecific polyclonal antibody anti-VSP9B10A previous to interaction with IEC-6 monolayers. These results demonstrate that the VSP9B10A protein secreted upon interaction with epithelial cells caused damage in these cells. Thus this protein might be considered as a conditional virulence factor candidate. To our knowledge this is the first report on the

  15. Antibodies against Lewis antigens inhibit the binding of human norovirus GII.4 virus-like particles to saliva but not to intestinal Caco-2 cells.

    Science.gov (United States)

    Carmona-Vicente, Noelia; Allen, David J; Rodríguez-Díaz, Jesús; Iturriza-Gómara, Miren; Buesa, Javier

    2016-05-21

    Human noroviruses (NoVs) are the main cause of gastroenteritis worldwide. The most commonly detected NoV strains belong to the genetically diverse GII.4 genotype, with new pandemic variants emerging periodically. Despite extensive efforts, NoV investigation has been hampered by the lack of an effective in vitro cell culture system. However, NoV-derived recombinant virus-like particles (VLPs) resembling empty capsids are good surrogates for analysing NoV antigenicity and virus-ligand interactions. NoV VLPs have been reported to bind to histo-blood group antigens (HBGAs). We have analysed the ability of NoV VLPs derived from GI.1 genotype and from three GII.4 genotype variants, GII.4-1999, GII.4-2004 and GII.4-2006b, to bind to porcine gastric mucin (PGM), human saliva and differentiated human intestinal Caco-2 cells (D-Caco-2 cells). Distinct patterns of saliva binding with the NoV GII.4 variant VLPs were observed, although they bound to D-Caco-2 cells independently of the expression of HBGAs. Monoclonal antibodies against Lewis antigens were able to block the binding of NoV VLPs to saliva, but not to D-Caco-2 cells. Blocking HBGAs on the surface of D-Caco-2 cells with specific monoclonal antibodies did not affect NoV VLP binding to cellular membranes. Co-localisation of Lewis y (Le(y)) and H-type 2 antigens with NoV VLPs was not observed by immunofluorescence assays. Although the binding of NoV VLPs of GII.4 genotype variants to human saliva samples occur with distinct HBGA binding patterns and can be blocked by antibodies against Lewis antigens, their attachment to D-Caco-2 cells can be mediated by other receptors, which still need further investigation.

  16. Sequence-based in silico analysis of well studied Hepatitis C Virus epitopes and their variants in other genotypes (particularly genotype 5a against South African human leukocyte antigen backgrounds

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    Prabdial-Sing Nishi

    2012-12-01

    Full Text Available Abstract Background Host genetics influence the outcome of HCV disease. HCV is also highly mutable and escapes host immunity. HCV genotypes are geographically distributed and HCV subtypes have been shown to have distinct repertoires of HLA-restricted viral epitopes which explains the lack of cross protection across genotypes observed in some studies. Despite this, immune databases and putative epitope vaccines concentrate almost exclusively on HCV genotype 1 class I-epitopes restricted by the HLA-A*02 allele. While both genotype and allele predominate in developed countries, we hypothesise that HCV variation and population genetics will affect the efficacy of proposed epitope vaccines in South Africa. This in silico study investigates HCV viral variability within well-studied epitopes identified in genotype 1 and uses algorithms to predict the immunogenicity of their variants from other less studied genotypes and thus rate the most promising vaccine candidates for the South African population. Six class I- and seven class II- restricted epitope sequences within the core, NS3, NS4B and NS5B regions were compared across the six HCV genotypes using local genotype 5a sequence data together with global data. Common HLA alleles in the South African population are A30:01, A02:01, B58:02, B07:02; DRB1*13:01 and DRB1*03:01. Epitope binding to 13 class I- and 8 class –II alleles were described using web-based prediction servers, Immune Epitope Database, (IEDB and Propred. Online population coverage tools were used to assess vaccine efficacy. Results Despite the homogeneity of genotype 1 and genotype 5 over the epitopes, there was limited promiscuity to local HLA-alleles.Host differences will make a putative vaccine less effective in South Africa. Of the 6 well-characterized class I- epitopes, only 2 class I- epitopes were promiscuous and 3 of the 7 class-II epitopes were better conserved and promiscuous. By fine tuning the putative vaccine using an

  17. Physiological response of Pichia pastoris GS115 to methanol-induced high level production of the Hepatitis B surface antigen: catabolic adaptation, stress responses, and autophagic processes.

    Science.gov (United States)

    Vanz, Ana Leticia; Lünsdorf, Heinrich; Adnan, Ahmad; Nimtz, Manfred; Gurramkonda, Chandrasekhar; Khanna, Navin; Rinas, Ursula

    2012-08-08

    Pichia pastoris is an established eukaryotic host for the production of recombinant proteins. Most often, protein production is under the control of the strong methanol-inducible aox1 promoter. However, detailed information about the physiological alterations in P. pastoris accompanying the shift from growth on glycerol to methanol-induced protein production under industrial relevant conditions is missing. Here, we provide an analysis of the physiological response of P. pastoris GS115 to methanol-induced high-level production of the Hepatitis B virus surface antigen (HBsAg). High product titers and the retention of the protein in the endoplasmic reticulum (ER) are supposedly of major impact on the host physiology. For a more detailed understanding of the cellular response to methanol-induced HBsAg production, the time-dependent changes in the yeast proteome and ultrastructural cell morphology were analyzed during the production process. The shift from growth on glycerol to growth and HBsAg production on methanol was accompanied by a drastic change in the yeast proteome. In particular, enzymes from the methanol dissimilation pathway started to dominate the proteome while enzymes from the methanol assimilation pathway, e.g. the transketolase DAS1, increased only moderately. The majority of methanol was metabolized via the energy generating dissimilatory pathway leading to a corresponding increase in mitochondrial size and numbers. The methanol-metabolism related generation of reactive oxygen species induced a pronounced oxidative stress response (e.g. strong increase of the peroxiredoxin PMP20). Moreover, the accumulation of HBsAg in the ER resulted in the induction of the unfolded protein response (e.g. strong increase of the ER-resident disulfide isomerase, PDI) and the ER associated degradation (ERAD) pathway (e.g. increase of two cytosolic chaperones and members of the AAA ATPase superfamily) indicating that potential degradation of HBsAg could proceed via the

  18. Impact of the thimerosal controversy on hepatitis B vaccine coverage of infants born to women of unknown hepatitis B surface antigen status in Michigan.

    Science.gov (United States)

    Biroscak, Brian J; Fiore, Anthony E; Fasano, Nancy; Fineis, Patrick; Collins, Michael P; Stoltman, Gillian

    2003-06-01

    Hepatitis B vaccine is recommended for all infants, and the series may be started during the delivery admission. For infants who are born either to women who are positive for hepatitis B surface antigen (HBsAg) or to women whose HBsAg status is unknown, vaccination should be started within 12 hours of birth to prevent perinatal and early childhood hepatitis B virus infection. Because of concerns about mercury exposures from vaccines that contain thimerosal, the United States Public Health Service (USPHS) and the American Academy of Pediatrics (AAP) recommended in July 1999 that the first dose of hepatitis B vaccine be deferred until 2-6 months of age but only for infants who are born to HBsAg-negative women. To assess the impact on birth-dose vaccine coverage for infants who are born to women with unknown HBsAg status, we measured coverage before and after July 1999. A sample of Michigan infants who were born to women whose HBsAg status was either unknown or missing were identified by reviewing newborn screening cards for infants who were born during 1) March-April 1999 (before recommendation changes [T1]); 2) July 15-September 15, 1999 (immediately after recommendation changes [T2]); and 3) March-April 2000 (6 months after resumption of pre-1999 practices were recommended [T3]). We verified maternal HBsAg screening and newborn hepatitis B vaccination by reviewing infant and maternal hospital records. Of 1201 infants who were born to women whose HBsAg status was indicated as unknown or missing on the newborn screening card during the 3 time periods, 216 (18%) were born to women whose status was truly unknown at the time of delivery, as determined by medical record review. During T1, 53% of these 216 infants received hepatitis B vaccine before hospital discharge, compared with 7% of infants who were born during T2 and 57% of infants who were born during T3. During T1, 19% of these infants received hepatitis B vaccine within 12 hours of birth compared with 1% of

  19. Physiological response of Pichia pastoris GS115 to methanol-induced high level production of the Hepatitis B surface antigen: catabolic adaptation, stress responses, and autophagic processes

    Science.gov (United States)

    2012-01-01

    Background Pichia pastoris is an established eukaryotic host for the production of recombinant proteins. Most often, protein production is under the control of the strong methanol-inducible aox1 promoter. However, detailed information about the physiological alterations in P. pastoris accompanying the shift from growth on glycerol to methanol-induced protein production under industrial relevant conditions is missing. Here, we provide an analysis of the physiological response of P. pastoris GS115 to methanol-induced high-level production of the Hepatitis B virus surface antigen (HBsAg). High product titers and the retention of the protein in the endoplasmic reticulum (ER) are supposedly of major impact on the host physiology. For a more detailed understanding of the cellular response to methanol-induced HBsAg production, the time-dependent changes in the yeast proteome and ultrastructural cell morphology were analyzed during the production process. Results The shift from growth on glycerol to growth and HBsAg production on methanol was accompanied by a drastic change in the yeast proteome. In particular, enzymes from the methanol dissimilation pathway started to dominate the proteome while enzymes from the methanol assimilation pathway, e.g. the transketolase DAS1, increased only moderately. The majority of methanol was metabolized via the energy generating dissimilatory pathway leading to a corresponding increase in mitochondrial size and numbers. The methanol-metabolism related generation of reactive oxygen species induced a pronounced oxidative stress response (e.g. strong increase of the peroxiredoxin PMP20). Moreover, the accumulation of HBsAg in the ER resulted in the induction of the unfolded protein response (e.g. strong increase of the ER-resident disulfide isomerase, PDI) and the ER associated degradation (ERAD) pathway (e.g. increase of two cytosolic chaperones and members of the AAA ATPase superfamily) indicating that potential degradation of HBsAg could

  20. Physiological response of Pichia pastoris GS115 to methanol-induced high level production of the Hepatitis B surface antigen: catabolic adaptation, stress responses, and autophagic processes

    Directory of Open Access Journals (Sweden)

    Vanz Ana Leticia

    2012-08-01

    Full Text Available Abstract Background Pichia pastoris is an established eukaryotic host for the production of recombinant proteins. Most often, protein production is under the control of the strong methanol-inducible aox1 promoter. However, detailed information about the physiological alterations in P. pastoris accompanying the shift from growth on glycerol to methanol-induced protein production under industrial relevant conditions is missing. Here, we provide an analysis of the physiological response of P. pastoris GS115 to methanol-induced high-level production of the Hepatitis B virus surface antigen (HBsAg. High product titers and the retention of the protein in the endoplasmic reticulum (ER are supposedly of major impact on the host physiology. For a more detailed understanding of the cellular response to methanol-induced HBsAg production, the time-dependent changes in the yeast proteome and ultrastructural cell morphology were analyzed during the production process. Results The shift from growth on glycerol to growth and HBsAg production on methanol was accompanied by a drastic change in the yeast proteome. In particular, enzymes from the methanol dissimilation pathway started to dominate the proteome while enzymes from the methanol assimilation pathway, e.g. the transketolase DAS1, increased only moderately. The majority of methanol was metabolized via the energy generating dissimilatory pathway leading to a corresponding increase in mitochondrial size and numbers. The methanol-metabolism related generation of reactive oxygen species induced a pronounced oxidative stress response (e.g. strong increase of the peroxiredoxin PMP20. Moreover, the accumulation of HBsAg in the ER resulted in the induction of the unfolded protein response (e.g. strong increase of the ER-resident disulfide isomerase, PDI and the ER associated degradation (ERAD pathway (e.g. increase of two cytosolic chaperones and members of the AAA ATPase superfamily indicating that potential

  1. Protection of hamsters from amebic liver abscess formation by immunization with the 150- and 170-kDa surface antigens of Entamoeba histolytica.

    Science.gov (United States)

    Cheng, X J; Tachibana, H

    2001-02-01

    A monoclonal antibody, EH3015, prevents in vitro adherence of Entamoeba histolytica trophozoites to mammalian cells and inhibits amebic liver abscess formation in hamsters. By immunoaffinity chromatography with the monoclonal antibody, purified E. histolytica antigens with molecular masses of 150 and 170 kDa under non-reduced conditions were obtained. Hamsters were immunized with these antigens (group I) or with fractions further purified by polyacrylamide gel electrophoresis (group II). Pooled immune sera from the two groups inhibited in vitro amebic adherence to Chinese hamster ovary cells by 98% at 1:10 dilutions. The immunized hamsters were challenged by the intrahepatic injection of E. histolytica trophozoites. Complete protection from abscess formation was observed in 38% of hamsters in group I and 67% in group II, whereas all control hamsters inoculated only with adjuvant developed amebic liver abscesses. In the immunized hamsters, the abscesses in the two groups were significantly smaller than in the controls. These results demonstrate that the E. histolytica antigens are possible vaccine candidates for amebiasis.

  2. Amino Acids in Hemagglutinin Antigenic Site B Determine Antigenic and Receptor Binding Differences between A(H3N2)v and Ancestral Seasonal H3N2 Influenza Viruses

    Science.gov (United States)

    Wang, Xiaoquan; Ilyushina, Natalia A.; Lugovtsev, Vladimir Y.; Bovin, Nicolai V.; Couzens, Laura K.; Gao, Jin

    2016-01-01

    .S. swine population in the mid-1990s, but they are different from both these ancestral viruses and current circulating human seasonal H3N2 strains in terms of their antigenic characteristics as measured by hemagglutination inhibition (HI) assay. In this study, we identified amino acids in antigenic site B of the surface glycoprotein hemagglutinin (HA) that explain the antigenic difference between A(H3N2)v and the ancestral H3N2 strains. These amino acid mutations also alter binding to minor human-type glycans, suggesting that host adaptation may contribute to the selection of antigenically distinct H3N2 variants which pose a threat to public health. PMID:27807224

  3. Single hemagglutinin mutations that alter both antigenicity and receptor binding avidity influence influenza virus antigenic clustering.

    Science.gov (United States)

    Li, Yang; Bostick, David L; Sullivan, Colleen B; Myers, Jaclyn L; Griesemer, Sara B; Stgeorge, Kirsten; Plotkin, Joshua B; Hensley, Scott E

    2013-09-01

    The hemagglutination inhibition (HAI) assay is the primary measurement used for identifying antigenically novel influenza virus strains. HAI assays measure the amount of reference sera required to prevent virus binding to red blood cells. Receptor binding avidities of viral strains are not usually taken into account when interpreting these assays. Here, we created antigenic maps of human H3N2 viruses that computationally account for variation in viral receptor binding avidities. These new antigenic maps differ qualitatively from conventional antigenic maps based on HAI measurements alone. We experimentally focused on an antigenic cluster associated with a single N145K hemagglutinin (HA) substitution that occurred between 1992 and 1995. Reverse-genetics experiments demonstrated that the N145K HA mutation increases viral receptor binding avidity. Enzyme-linked immunosorbent assays (ELISA) revealed that the N145K HA mutation does not prevent antibody binding; rather, viruses possessing this mutation escape antisera in HAI assays simply by attaching to cells more efficiently. Unexpectedly, we found an asymmetric antigenic effect of the N145K HA mutation. Once H3N2 viruses acquired K145, an epitope involving amino acid 145 became antigenically dominant. Antisera raised against an H3N2 strain possessing K145 had reduced reactivity to H3N2 strains possessing N145. Thus, individual mutations in HA can influence antigenic groupings of strains by altering receptor binding avidity and by changing the dominance of antibody responses. Our results indicate that it will be important to account for variation in viral receptor binding avidity when performing antigenic analyses in order to identify genuine antigenic differences among influenza virus variants.

  4. Increased expression of the type IV secretion system in piliated Neisseria gonorrhoeae variants.

    Science.gov (United States)

    Salgado-Pabón, Wilmara; Du, Ying; Hackett, Kathleen T; Lyons, Katelynn M; Arvidson, Cindy Grove; Dillard, Joseph P

    2010-04-01

    Neisseria gonorrhoeae produces a type IV secretion system that secretes chromosomal DNA. The secreted DNA is active in the transformation of other gonococci in the population and may act to transfer antibiotic resistance genes and variant alleles for surface antigens, as well as other genes. We observed that gonococcal variants that produced type IV pili secreted more DNA than variants that were nonpiliated, suggesting that the process may be regulated. Using microarray analysis, we found that a piliated strain showed increased expression of the gene for the putative type IV secretion coupling protein TraD, whereas a nonpiliated variant showed increased expression of genes for transcriptional and translational machinery, consistent with its higher growth rate compared to that of the piliated strain. These results suggested that type IV secretion might be controlled by either traD expression or growth rate. A mutant with a deletion in traD was found to be deficient in DNA secretion. Further mutation and complementation analysis indicated that traD is transcriptionally and translationally coupled to traI, which encodes the type IV secretion relaxase. We were able to increase DNA secretion in a nonpiliated strain by inserting a gene cassette with a strong promoter to drive the expression of the putative operon containing traI and traD. Together, these data suggest a model in which the type IV secretion system apparatus is made constitutively, while its activity is controlled through regulation of traD and traI.

  5. Effective adoptive immunotherapy of triple-negative breast cancer by folate receptor-alpha redirected CAR T cells is influenced by surface antigen expression level

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    De-Gang Song

    2016-07-01

    Full Text Available Abstract Background The poor prognosis and the limited efficacy of targeted therapy in patients with triple-negative breast cancer (TNBC have raised the need for alternative therapies. Recent studies have demonstrated that folate receptor-alpha (FRα may represent an ideal tumor-associated marker for immunotherapy for TNBC. Methods The aim of the present study was to apply a chimeric antigen receptor (CAR approach for the targeting of FRα expressed on TNBC cells and evaluate the antitumor activity of CAR-engineered T cells in vitro and in vivo. Results We found that human T cells expressing a FRα-specific CAR were potent and specific killers of TNBC cells that express moderate levels of FRα in vitro and significantly inhibited tumor outgrowth following infusion into immunodeficient mice bearing an MDA-MB-231 tumor xenograft. However, the antitumor activity of the FRα CAR T cells was modest when compared to the same CAR T cells applied in an ovarian tumor xenograft model where FRα expression is more abundant. Notably, FRα CAR T cells induced superior tumor regression in vivo against MDA-MB-231 that was engineered for overexpression of FRα. Conclusions Taken together, our results show that FRα CAR T cells can mediate antitumor activity against established TNBC tumor, particularly when FRα is expressed at higher levels. These results have significant implications for the pre-selection of patients with high antigen expression levels when utilizing CAR-based adoptive T cell therapies of cancer in future clinical trials.

  6. Genetic diversity of Plasmodium falciparum merozoite surface protein-1 (block 2), glutamate-rich protein and sexual stage antigen Pfs25 from Chandigarh, North India.

    Science.gov (United States)

    Kaur, Hargobinder; Sehgal, Rakesh; Goyal, Kapil; Makkar, Nikita; Yadav, Richa; Bharti, Praveen K; Singh, Neeru; Sarmah, Nilanju P; Mohapatra, Pradyumna K; Mahanta, Jagadish; Bansal, Devendra; Sultan, Ali A; Kanwar, Jagat R

    2017-12-01

    To elucidate the genetic diversity of Plasmodium falciparum in residual transmission foci of northern India. Clinically suspected patients with malaria were screened for malaria infection by microscopy. 48 P. falciparum-infected patients were enrolled from tertiary care hospital in Chandigarh, India. Blood samples were collected from enrolled patients, genomic DNA extraction and nested PCR was performed for further species confirmation. Sanger sequencing was carried out using block 2 region of msp1, R2 region of glurp and pfs25-specific primers. Extensive diversity was found in msp1 alleles with predominantly RO33 alleles. Overall allelic prevalence was 55.8% for RO33, 39.5% for MAD20 and 4.7% for K1. Six variants were observed in MAD20, whereas no variant was found in RO33 and K1 alleles. A phylogenetic analysis of RO33 alleles indicated more similarity to South African isolates, whereas MAD20 alleles showed similarity with South-East Asian isolates. In glurp, extensive variation was observed with eleven different alleles based on the AAU repeats. However, pfs25 showed less diversity and was the most stable among the targeted genes. Our findings document the genetic diversity among circulating strains of P. falciparum in an area of India with low malaria transmission and could have implications for control strategies to reach the national goal of malaria elimination. © 2017 John Wiley & Sons Ltd.

  7. Identification of sVSG117 as an immunodiagnostic antigen and evaluation of a dual-antigen lateral flow test for the diagnosis of human African trypanosomiasis.

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    Lauren Sullivan

    2014-07-01

    Full Text Available The diagnosis of human African trypanosomiasis (HAT caused by Trypanosoma brucei gambiense relies mainly on the Card Agglutination Test for Trypanosomiasis (CATT. There is no immunodiagnostic for HAT caused by T. b. rhodesiense. Our principle aim was to develop a prototype lateral flow test that might be an improvement on CATT.Pools of infection and control sera were screened against four different soluble form variant surface glycoproteins (sVSGs by ELISA and one, sVSG117, showed particularly strong immunoreactivity to pooled infection sera. Using individual sera, sVSG117 was shown to be able to discriminate between T. b. gambiense infection and control sera by both ELISA and lateral flow test. The sVSG117 antigen was subsequently used with a previously described recombinant diagnostic antigen, rISG65, to create a dual-antigen lateral flow test prototype. The latter was used blind in a virtual field trial of 431 randomized infection and control sera from the WHO HAT Specimen Biobank.In the virtual field trial, using two positive antigen bands as the criterion for infection, the sVSG117 and rISG65 dual-antigen lateral flow test prototype showed a sensitivity of 97.3% (95% CI: 93.3 to 99.2 and a specificity of 83.3% (95% CI: 76.4 to 88.9 for the detection of T. b. gambiense infections. The device was not as good for detecting T. b. rhodesiense infections using two positive antigen bands as the criterion for infection, with a sensitivity of 58.9% (95% CI: 44.9 to 71.9 and specificity of 97.3% (95% CI: 90.7 to 99.7. However, using one or both positive antigen band(s as the criterion for T. b. rhodesiense infection improved the sensitivity to 83.9% (95% CI: 71.7 to 92.4 with a specificity of 85.3% (95% CI: 75.3 to 92.4. These results encourage further development of the dual-antigen device for clinical use.

  8. A rabies virus vampire bat variant shows increased neuroinvasiveness in mice when compared to a carnivore variant.

    Science.gov (United States)

    Mesquita, Leonardo Pereira; Gamon, Thais Helena Martins; Cuevas, Silvia Elena Campusano; Asano, Karen Miyuki; Fahl, Willian de Oliveira; Iamamoto, Keila; Scheffer, Karin Correa; Achkar, Samira Maria; Zanatto, Dennis Albert; Mori, Cláudia Madalena Cabrera; Maiorka, Paulo César; Mori, Enio

    2017-12-01

    Rabies is one of the most important zoonotic diseases and is caused by several rabies virus (RABV) variants. These variants can exhibit differences in neurovirulence, and few studies have attempted to evaluate the neuroinvasiveness of variants derived from vampire bats and wild carnivores. The aim of this study was to evaluate the neuropathogenesis of infection with two Brazilian RABV street variants (variant 3 and crab-eating fox) in mice. BALB/c mice were inoculated with RABV through the footpad, with the 50% mouse lethal dose (LD50) determined by intracranial inoculation. The morbidity of rabies in mice infected with variant 3 and the crab-eating fox strain was 100% and 50%, respectively, with an incubation period of 7 and 6 days post-inoculation (dpi), respectively. The clinical disease in mice was similar with both strains, and it was characterized initially by weight loss, ruffled fur, hunched posture, and hind limb paralysis progressing to quadriplegia and recumbency at 9 to 12 dpi. Histological lesions within the central nervous system (CNS) characterized by nonsuppurative encephalomyelitis with neuronal degeneration and necrosis were observed in mice infected with variant 3 and those infected with the crab-eating fox variant. However, lesions and the presence of RABV antigen, were more widespread within the CNS of variant-3-infected mice, whereas in crab-eating fox-variant-infected mice, RABV antigens were more restricted to caudal areas of the CNS, such as the spinal cord and brainstem. In conclusion, the results shown here demonstrate that the RABV vampire bat strain (variant 3) has a higher potential for neuroinvasiveness than the carnivore variant.

  9. A Rapid Method to Characterize Mouse IgG Antibodies and Isolate Native Antigen Binding IgG B Cell Hybridomas.

    Directory of Open Access Journals (Sweden)

    Haolin Liu

    Full Text Available B cell hybridomas are an important source of monoclonal antibodies. In this paper, we developed a high-throughput method to characterize mouse IgG antibodies using surface plasmon resonance technology. This assay rapidly determines their sub-isotypes, whether they bind native antigen and their approximate affinities for the antigen using only 50 μl of hybridoma cell culture supernatant. Moreover, we found that mouse hybridomas secreting IgG antibodies also have membrane form IgG expression without Igα. Based on this surface IgG, we used flow cytometry to isolate rare γ2a isotype switched variants from a γ2b antibody secreting hybridoma cell line. Also, we used fluorescent antigen to single cell sort antigen binding hybridoma cells from bulk mixture of fused hybridoma cells instead of the traditional multi-microwell plate screening and limiting dilution sub-cloning thus saving time and labor. The IgG monoclonal antibodies specific for the native antigen identified with these methods are suitable for in vivo therapeutic uses, but also for sandwich ELISA assays, histology, flow cytometry, immune precipitation and x-ray crystallography.

  10. Sperm antigens in fertilization.

    Science.gov (United States)

    Saling, P M

    1990-02-01

    A review of sperm antigens involved in fertilization includes a description of sperm differentiation, seminal fluid components that coat sperm, sperm antigens involved in binding to the zona pellucida (ZP), antigens involved in the acrosome reaction, in zona pellucida penetration, and those active in fusion with the ova membrane. Sperm antigens are located in certain domains of the cell, and they are altered during capacitation and passage through the female tract. Caltrin and acrosome-stabilizing factor are applied by seminal fluid. At least 2 antigens have been studied that occur in sterile women, although one cross reacts with milk proteins. Some antigens active in ZP binding are trypsin, proacrosin, acrosin, PH-20 from guinea pigs, and rabbit sperm autoantigen I. Antigens involved in the acrosome reaction, such as M42, are likely to cross react with other body proteins that also entail exocytosis. A mouse antigen involved in ZP penetration, MS 207 is well characterized. PH-30 from guinea pigs and M29 from mouse participate in sperm-egg membrane fusion, as does fertilization antigen I from human and mouse sperm which is know to cause infertility. Oddly, patients' sera react with polymers but not monomers of this antigen. Studies with antisperm antibodies suggest that it will not be necessary to agglutinate all sperm to block fertility, only to inhibit a single sperm epitope and function. It will probably be feasible to inhibit multiple successive events, and possibly to induce temporary immunity.

  11. Characterization and allelic variation of the transporters associated with antigen processing (TAP) genes in the domestic dog (Canis lupus familiaris)

    Science.gov (United States)

    Gojanovich, Gregory S.; Ross, Peter; Holmer, Savannah R.; Holmes, Jennifer C.; Hess, Paul R.

    2013-01-01

    The function of the transporters associated with antigen processing (TAP) complex is to shuttle antigenic peptides from the cytosol to the endoplasmic reticulum to load MHC class I molecules for CD8+ T-cell immunosurveillance. Here we report the promoter and coding regions of the canine TAP1 and TAP2 genes, which encode the homologous subunits forming the TAP heterodimer. By sampling genetically divergent breeds, polymorphisms in both genes were identified, although there were few amino acid differences between alleles. Splice variants were also found. When aligned to TAP genes of other species, functional regions appeared conserved, and upon phylogenetic analysis, canine sequences segregated appropriately with their orthologs. Transfer of the canine TAP2 gene into a murine TAP2-defective cell line rescued surface MHC class I expression, confirming exporter function. This data should prove useful in investigating the association of specific TAP defects or alleles with immunity to intracellular pathogens and cancer in dogs. PMID:23892057

  12. Comparative Investigation of the Genomic Regions Involved in Antigenic Variation of the TprK Antigen among Treponemal Species, Subspecies, and Strains

    Science.gov (United States)

    Brandt, Stephanie L.; Puray-Chavez, Maritza; Reid, Tara Brinck; Godornes, Charmie; Molini, Barbara J.; Benzler, Martin; Hartig, Jörg S.; Lukehart, Sheila A.; Centurion-Lara, Arturo

    2012-01-01

    Although the three Treponema pallidum subspecies (T. pallidum subsp. pallidum, T. pallidum subsp. pertenue, and T. pallidum subsp. endemicum), Treponema paraluiscuniculi, and the unclassified Fribourg-Blanc treponeme cause clinically distinct diseases, these pathogens are genetically and antigenically highly related and are able to cause persistent infection. Recent evidence suggests that the putative surface-exposed variable antigen TprK plays an important role in both treponemal immune evasion and persistence. tprK heterogeneity is generated by nonreciprocal gene conversion between the tprK expression site and donor sites. Although each of the above-mentioned species and subspecies has a functional tprK antigenic variation system, it is still unclear why the level of expression and the rate at which tprK diversifies during infection can differ significantly among isolates. To identify genomic differences that might affect the generation and expression of TprK variants among these pathogens, we performed comparative sequence analysis of the donor sites, as well as the tprK expression sites, among eight T. pallidum subsp. pallidum isolates (Nichols Gen, Nichols Sea, Chicago, Sea81-4, Dal-1, Street14, UW104, and UW126), three T. pallidum subsp. pertenue isolates (Gauthier, CDC2, and Samoa D), one T. pallidum subsp. endemicum isolate (Iraq B), the unclassified Fribourg-Blanc isolate, and the Cuniculi A strain of T. paraluiscuniculi. Synteny and sequence conservation, as well as deletions and insertions, were found in the regions harboring the donor sites. These data suggest that the tprK recombination system is harbored within dynamic genomic regions and that genomic differences might be an important key to explain discrepancies in generation and expression of tprK variants among these Treponema isolates. PMID:22661689

  13. The persistence ofhepatitis B antigen in the bloodtneal of the ...

    African Journals Online (AJOL)

    Abstract The persistence of the hepatitis B virus surface antigen (HBsAg) was used as an index of the sur- vival tim.e ... or unknown origin in the alleviation of venous congestion in failing microsurgical procedures. ... sistence of hepatitis B surface antigen (HBsAg) in the bloodmeal of the leech, Asiancobdella bumonensis, ...

  14. Antigens of Streptococcus sanguis

    Science.gov (United States)

    Rosan, Burton

    1973-01-01

    An antigenic analysis of the alpha-hemolytic streptococci isolated from dental plaque was performed by use of antisera against a strain of Streptococcus sanguis (M-5) which was isolated from dental plaque. Immunoelectrophoretic and Ouchterlony tests of Rantz and Randall extracts of 45 strains gave positive reactions with the M-5 antisera. These strains represented 60% of the strains tested. The number of antigens which could be identified in these extracts varied from one to five and were designated a to e. The a antigen was found in 36 of the strains tested, including reference strains of S. sanguis and the group H streptococci. The strains reacting with the M-5 antisera were divided into two majors types: type I consisted of 23 strains in which the a antigen was found alone or with one or more of the c, d, and e antigens; type II consisted of 13 strains in which both the a and b antigens were found with or without one or more of the c, d, and e antigens. The remaining strains contained, either singly or in combination, the b, c, d, and e antigens but not the a antigen. Biochemical tests of representatives of each serotype and reference strains indicated that strains reacting with M-5 antisera were S. sanguis. These findings suggest that S. sanguis strains share common physiological and serological properties. Images PMID:4633291

  15. Antigenic presentation of heterologous epitopes engineered into the outer surface-exposed helix 4 loop region of human papillomavirus L1 capsomeres

    Directory of Open Access Journals (Sweden)

    Murata Yoshihiko

    2009-06-01

    Full Text Available Abstract Background Human papillomavirus (HPV L1 capsid proteins can self-assemble into pentamers (capsomeres that are immunogenic and can elicit neutralizing antibodies. Structural modelling of L1 inter-pentameric interactions predicts that helix 4 (h4 of each of the five L1 monomers project laterally and outwards from the pentamer. We sought to utilize HPV L1 capsomeres as a vaccine platform by engineering heterologous epitopes within L1 derivatives deleted for h4 domain. Results We used baculovirus – infected Trichoplusia ni cells and ultracentrifugation to synthesize and purify three 16L1 derivatives: one bearing a short deletion (amino acids 404–436 encompassing the h4 domain, and two others, each bearing a conserved neutralizing epitope of the human respiratory syncytial virus (RSV fusion (F protein (residues 255–278 and 423–436 that was substituted for the deleted L1 h4 domain residues. Each of the three capsomere derivatives was recognized by anti-L1 antibodies, while two bearing the RSV F-derived moieties were recognized by anti-RSV F antibodies. All three L1 derivatives formed ring-like structures that were similar in morphology and size to those described for native 16L1 capsomeres. When injected into mice, each of the capsomere derivatives was immunogenic with respect to L1 protein, and immunization with chimeric L1-RSV F pentamers resulted in RSV non-neutralizing antisera that recognized purified RSV F protein in immunoblots. Conclusion HPV L1 monomers bearing heterologous epitopes within the L1 h4 region can self-assemble into capsomeres that elicit antibody response against such non-HPV encoded epitopes. Thus, the L1 h4 region can function as a novel antigen display site within the L1 pentamer, which in turn may serve as a potential vaccine template.

  16. Neisseria meningitidis antigen NMB0088: sequence variability, protein topology and vaccine potential.

    Science.gov (United States)

    Sardiñas, Gretel; Yero, Daniel; Climent, Yanet; Caballero, Evelin; Cobas, Karem; Niebla, Olivia

    2009-02-01

    The significance of Neisseria meningitidis serogroup B membrane proteins as vaccine candidates is continually growing. Here, we studied different aspects of antigen NMB0088, a protein that is abundant in outer-membrane vesicle preparations and is thought to be a surface protein. The gene encoding protein NMB0088 was sequenced in a panel of 34 different meningococcal strains with clinical and epidemiological relevance. After this analysis, four variants of NMB0088 were identified; the variability was confined to three specific segments, designated VR1, VR2 and VR3. Secondary structure predictions, refined with alignment analysis and homology modelling using FadL of Escherichia coli, revealed that almost all the variable regions were located in extracellular loop domains. In addition, the NMB0088 antigen was expressed in E. coli and a procedure for obtaining purified recombinant NMB0088 is described. The humoral immune response elicited in BALB/c mice was measured by ELISA and Western blotting, while the functional activity of these antibodies was determined in a serum bactericidal assay and an animal protection model. After immunization in mice, the recombinant protein was capable of inducing a protective response when it was administered inserted into liposomes. According to our results, the recombinant NMB0088 protein may represent a novel antigen for a vaccine against meningococcal disease. However, results from the variability study should be considered for designing a cross-protective formulation in future studies.

  17. Comparison of the binding properties of the mushroom Marasmius oreades lectin and Griffonia simplicifolia I-B isolectin to alphagalactosyl carbohydrate antigens in the surface phase

    DEFF Research Database (Denmark)

    Kirkeby, Svend; Winter, Harry C; Goldstein, Irwin J

    2004-01-01

    The binding of two alpha-galactophilic lectins, Marasmius oreades agglutinin (MOA), and Griffonia simplicifolia I isolectin B(4) (GS I-B(4)) to neoglycoproteins and natural glycoproteins were compared in a surface phase assay. Neoglycoproteins carrying various alpha-galactosylated glycans and lam...

  18. DNA rearrangements and antigenic variation in Trypanosoma equiperdum: multiple expression-linked sites in independent isolates of trypanosomes expressing the same antigen.

    Science.gov (United States)

    Longacre, S; Hibner, U; Raibaud, A; Eisen, H; Baltz, T; Giroud, C; Baltz, D

    1983-03-01

    African trypanosomes resist the immune response of their mammalian hosts by varying the surface glycoprotein which constitutes their antigenic identity. The molecular mechanism of this antigenic variation involves the successive activation of a series of genes which code for different variant surface glycoproteins (VSGs). We have studied the expression of two VSG genes (those of VSG-1 and VSG-28) in Trypanosoma equiperdum, and we report the following findings. (i) The expression of both VSG genes is associated with the duplication and transposition of corresponding basic copy genes. (ii) The duplicated transposed copy appears to be the expressed copy. (iii) Although there are multiple genes which cross-hybridize with the VSG-1 cDNA probe, only one of these appears to be used as a template for the expression-linked copy in four independent BoTat-1 clones. (iv) Analysis of the genomic environments of the expressed VSG-1 genes from each of four independently derived BoTat-1 trypanosome clones revealed that there are at least three different sites into which the expression-linked copy can be inserted.

  19. Mini-review: Strategies for Variation and Evolution of Bacterial Antigens

    Directory of Open Access Journals (Sweden)

    Janet Foley

    2015-01-01

    Full Text Available 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.

  20. Induction of long-lasting protective immunity against Toxoplasma gondii in BALB/c mice by recombinant surface antigen 1 protein encapsulated in poly (lactide-co-glycolide microparticles

    Directory of Open Access Journals (Sweden)

    Chuang Shu-Chun

    2013-02-01

    Full Text Available Abstract Background Current development efforts of subunit vaccines against Toxoplasma gondii, the etiological agent of toxoplasmosis, have been focused mainly on tachyzoite surface antigen 1 (SAG1. Since microparticles made from poly (lactide-co-glycolide (PLG polymers have been developed as safe, potent adjuvants or delivery systems, we aimed to encapsulate recombinant SAG1 (rSAG1 with the PLG polymers to prepare PLG-encapsulated rSAG1 (PLG-rSAG1 microparticles that would sustain rSAG1 release and generate long-lasting protective immunity against T. gondii in BALB/c mice. Methods In the present study, rSAG1 was encapsulated into PLG microparticles by the double emulsion method. PLG-rSAG1 microparticles were then intraperitoneally injected twice at a 14-day interval into BALB/c mice. We examined the ability of PLG-rSAG1 microparticles to induce and prolong effective anti-Toxoplasma immune responses, in comparison with rSAG1 formulated with a Vet L-10 adjuvant (rSAG1 (Vet L-10. Eight weeks after the last immunization, protective activities were also evaluated after a lethal subcutaneous challenge of 1x104 live T. gondii tachyzoites. Results PLG-rSAG1 microparticles, 4.25~6