Eight Nucleotide Substitutions Inhibit Splicing to HPV-16 3′-Splice Site SA3358 and Reduce the Efficiency by which HPV-16 Increases the Life Span of Primary Human Keratinocytes

PubMed Central

Li, Xiaoze; Johansson, Cecilia; Cardoso Palacios, Carlos; Mossberg, Anki; Dhanjal, Soniya; Bergvall, Monika; Schwartz, Stefan

2013-01-01

The most commonly used 3′-splice site on the human papillomavirus type 16 (HPV-16) genome named SA3358 is used to produce HPV-16 early mRNAs encoding E4, E5, E6 and E7, and late mRNAs encoding L1 and L2. We have previously shown that SA3358 is suboptimal and is totally dependent on a downstream splicing enhancer containingmultiple potential ASF/SF2 binding sites. Here weshow that only one of the predicted ASF/SF2 sites accounts for the majority of the enhancer activity. We demonstrate that single nucleotide substitutions in this predicted ASF/SF2 site impair enhancer function and that this correlates with less efficient binding to ASF/SF2 in vitro. We provide evidence that HPV-16 mRNAs that arespliced to SA3358 interact with ASF/SF2 in living cells. In addition,mutational inactivation of the ASF/SF2 site weakened the enhancer at SA3358 in episomal forms of the HPV-16 genome, indicating that the enhancer is active in the context of the full HPV-16 genome.This resulted in induction of HPV-16 late gene expression as a result of competition from late splice site SA5639. Furthermore, inactivation of the ASF/SF2 site of the SA3358 splicing enhancer reduced the ability of E6- and E7-encoding HPV-16 plasmids to increase the life span of primary keratinocytes in vitro, demonstrating arequirement for an intact splicing enhancer of SA3358 forefficient production of the E6 and E7 mRNAs. These results link the strength of the HPV-16 SA3358 splicing enhancer to expression of E6 and E7 and to the pathogenic properties of HPV-16. PMID:24039800

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Analysis of Host Responses to Mycobacterium tuberculosis Antigens in a Multi-Site Study of Subjects with Different TB and HIV Infection States in Sub-Saharan Africa

PubMed Central

Sutherland, Jayne S.; Lalor, Maeve K.; Black, Gillian F.; Ambrose, Lyn R.; Loxton, Andre G.; Chegou, Novel N.; Kassa, Desta; Mihret, Adane; Howe, Rawleigh; Mayanja-Kizza, Harriet; Gomez, Marie P.; Donkor, Simon; Franken, Kees; Hanekom, Willem; Klein, Michel R.; Parida, Shreemanta K.; Boom, W. Henry; Thiel, Bonnie A.; Crampin, Amelia C.; Ota, Martin; Walzl, Gerhard; Ottenhoff, Tom H. M.; Dockrell, Hazel M.; Kaufmann, Stefan H. E.

2013-01-01

Background Tuberculosis (TB) remains a global health threat with 9 million new cases and 1.4 million deaths per year. In order to develop a protective vaccine, we need to define the antigens expressed by Mycobacterium tuberculosis (Mtb), which are relevant to protective immunity in high-endemic areas. Methods We analysed responses to 23 Mtb antigens in a total of 1247 subjects with different HIV and TB status across 5 geographically diverse sites in Africa (South Africa, The Gambia, Ethiopia, Malawi and Uganda). We used a 7-day whole blood assay followed by IFN-γ ELISA on the supernatants. Antigens included PPD, ESAT-6 and Ag85B (dominant antigens) together with novel resuscitation-promoting factors (rpf), reactivation proteins, latency (Mtb DosR regulon-encoded) antigens, starvation-induced antigens and secreted antigens. Results There was variation between sites in responses to the antigens, presumably due to underlying genetic and environmental differences. When results from all sites were combined, HIV- subjects with active TB showed significantly lower responses compared to both TST- and TST+ contacts to latency antigens (Rv0569, Rv1733, Rv1735, Rv1737) and the rpf Rv0867; whilst responses to ESAT-6/CFP-10 fusion protein (EC), PPD, Rv2029, TB10.3, and TB10.4 were significantly higher in TST+ contacts (LTBI) compared to TB and TST- contacts fewer differences were seen in subjects with HIV co-infection, with responses to the mitogen PHA significantly lower in subjects with active TB compared to those with LTBI and no difference with any antigen. Conclusions Our multi-site study design for testing novel Mtb antigens revealed promising antigens for future vaccine development. The IFN-γ ELISA is a cheap and useful tool for screening potential antigenicity in subjects with different ethnic backgrounds and across a spectrum of TB and HIV infection states. Analysis of cytokines other than IFN-γ is currently on-going to determine correlates of protection, which may

Analysis of host responses to Mycobacterium tuberculosis antigens in a multi-site study of subjects with different TB and HIV infection states in sub-Saharan Africa.

PubMed

Sutherland, Jayne S; Lalor, Maeve K; Black, Gillian F; Ambrose, Lyn R; Loxton, Andre G; Chegou, Novel N; Kassa, Desta; Mihret, Adane; Howe, Rawleigh; Mayanja-Kizza, Harriet; Gomez, Marie P; Donkor, Simon; Franken, Kees; Hanekom, Willem; Klein, Michel R; Parida, Shreemanta K; Boom, W Henry; Thiel, Bonnie A; Crampin, Amelia C; Ota, Martin; Walzl, Gerhard; Ottenhoff, Tom H M; Dockrell, Hazel M; Kaufmann, Stefan H E

2013-01-01

Tuberculosis (TB) remains a global health threat with 9 million new cases and 1.4 million deaths per year. In order to develop a protective vaccine, we need to define the antigens expressed by Mycobacterium tuberculosis (Mtb), which are relevant to protective immunity in high-endemic areas. We analysed responses to 23 Mtb antigens in a total of 1247 subjects with different HIV and TB status across 5 geographically diverse sites in Africa (South Africa, The Gambia, Ethiopia, Malawi and Uganda). We used a 7-day whole blood assay followed by IFN-γ ELISA on the supernatants. Antigens included PPD, ESAT-6 and Ag85B (dominant antigens) together with novel resuscitation-promoting factors (rpf), reactivation proteins, latency (Mtb DosR regulon-encoded) antigens, starvation-induced antigens and secreted antigens. There was variation between sites in responses to the antigens, presumably due to underlying genetic and environmental differences. When results from all sites were combined, HIV- subjects with active TB showed significantly lower responses compared to both TST(-) and TST(+) contacts to latency antigens (Rv0569, Rv1733, Rv1735, Rv1737) and the rpf Rv0867; whilst responses to ESAT-6/CFP-10 fusion protein (EC), PPD, Rv2029, TB10.3, and TB10.4 were significantly higher in TST(+) contacts (LTBI) compared to TB and TST(-) contacts fewer differences were seen in subjects with HIV co-infection, with responses to the mitogen PHA significantly lower in subjects with active TB compared to those with LTBI and no difference with any antigen. Our multi-site study design for testing novel Mtb antigens revealed promising antigens for future vaccine development. The IFN-γ ELISA is a cheap and useful tool for screening potential antigenicity in subjects with different ethnic backgrounds and across a spectrum of TB and HIV infection states. Analysis of cytokines other than IFN-γ is currently on-going to determine correlates of protection, which may be useful for vaccine efficacy

Identification and analysis of cytochrome P450IID6 antigenic sites recognized by anti-liver-kidney microsome type-1 antibodies (LKM1).

PubMed

Yamamoto, A M; Cresteil, D; Boniface, O; Clerc, F F; Alvarez, F

1993-05-01

Anti-liver-kidney microsome type-1 antibodies (LKM1), present in sera from a group of patients with autoimmune hepatitis, are directed against P450IID6. Previous work, using cDNA constructions spanning most of the P450IID6 protein defined the main immunogenic site between the amino acids (aa), 254-271 and predicted the presence of other putative immunogenic sites in the molecule. Fusion proteins from new cDNA constructions, spanning so-far-untested regions between aa 1-125 and 431-522, were not recognized by LKM1-positive sera. Synthetic peptides, representing sequences from putative immunogenic regions or previously untested regions, allowed a precise definition of four antigenic sites located between peptides 257-269, 321-351, 373-389 and 410-429, which were recognized, respectively, by 14, 8, 1 and 2 out of 15 LKM1-positive sera tested. The minimal sequence of the main antigenic site (peptide 257-269) recognized by the autoantibody was established to be WDPAQPPRD (peptide 262-270). In addition, deletion and replacement experiments showed that aa 263 (Asp) was essential for the binding of the autoantibody to peptide 262-270. Analysis of the second most frequently recognized peptide between aa 321-351, was performed using peptides 321-339 and 340-351 in competitive inhibition studies. Complete elimination of antibody binding to peptide 321-351 obtained by absorption of both shorter peptides indicated that peptide 321-351 is a discontinuous antigenic site. LKM1-positive sera reacting against peptide 321-351 recognized either both the shorter peptides or just one of them preferentially. Results of the present study suggest that the production of LKM1 antibodies is an antigen-driven, poly- or oligoclonal B cell response. The identification of antigenic sites will allow: (i) the development of specific diagnostic tests and (ii) further studies on the pathogenic value of LKM1 antibodies in autoimmune hepatitis.

Location of a major antigenic site involved in Ross River virus neutralization.

PubMed

Vrati, S; Fernon, C A; Dalgarno, L; Weir, R C

1988-02-01

The location of a major antigenic domain involved in the neutralization of an alphavirus, Ross River virus, has been defined in terms of its position in the amino acid sequence of the E2 glycoprotein. The domain encompasses three topographically close epitopes which were identified using three E2-specific neutralizing monoclonal antibodies in competitive binding assays. Nucleotide sequencing of the structural protein genes of monoclonal antibody-selected antigenic variants showed that for each variant there was a single nucleotide change in the E2 gene leading to a nonconservative amino acid substitution in E2. Changes were at positions 216, 234, and 246-251 in the amino acid sequence. The epitopes are in a region of E2 which, though not strongly conserved as to sequence among Ross River virus, Semliki Forest virus, and Sindbis virus, is conserved in its hydropathy profile among the three alphaviruses. The epitopes lie between two asparagine-linked glycosylation sites (residues 200 and 262) in E2. They are conserved as to position between the mouse virulent T48 strain and the mouse avirulent NB5092 strain.

Longitudinal Antigenic Sequences and Sites from Intra-Host Evolution (LASSIE) identifies immune-selected HIV variants

DOE PAGES

Hraber, Peter; Korber, Bette; Wagh, Kshitij; ...

2015-10-21

Within-host genetic sequencing from samples collected over time provides a dynamic view of how viruses evade host immunity. Immune-driven mutations might stimulate neutralization breadth by selecting antibodies adapted to cycles of immune escape that generate within-subject epitope diversity. Comprehensive identification of immune-escape mutations is experimentally and computationally challenging. With current technology, many more viral sequences can readily be obtained than can be tested for binding and neutralization, making down-selection necessary. Typically, this is done manually, by picking variants that represent different time-points and branches on a phylogenetic tree. Such strategies are likely to miss many relevant mutations and combinations ofmore » mutations, and to be redundant for other mutations. Longitudinal Antigenic Sequences and Sites from Intrahost Evolution (LASSIE) uses transmitted founder loss to identify virus “hot-spots” under putative immune selection and chooses sequences that represent recurrent mutations in selected sites. LASSIE favors earliest sequences in which mutations arise. Here, with well-characterized longitudinal Env sequences, we confirmed selected sites were concentrated in antibody contacts and selected sequences represented diverse antigenic phenotypes. Finally, practical applications include rapidly identifying immune targets under selective pressure within a subject, selecting minimal sets of reagents for immunological assays that characterize evolving antibody responses, and for immunogens in polyvalent “cocktail” vaccines.« less

Antigenic Drift in H5N1 Avian Influenza Virus in Poultry Is Driven by Mutations in Major Antigenic Sites of the Hemagglutinin Molecule Analogous to Those for Human Influenza Virus▿†

PubMed Central

Cattoli, Giovanni; Milani, Adelaide; Temperton, Nigel; Zecchin, Bianca; Buratin, Alessandra; Molesti, Eleonora; Aly, Mona Meherez; Arafa, Abdel; Capua, Ilaria

2011-01-01

H5N1 highly pathogenic avian influenza virus has been endemic in poultry in Egypt since 2008, notwithstanding the implementation of mass vaccination and culling of infected birds. Extensive circulation of the virus has resulted in a progressive genetic evolution and an antigenic drift. In poultry, the occurrence of antigenic drift in avian influenza viruses is less well documented and the mechanisms remain to be clarified. To test the hypothesis that H5N1 antigenic drift is driven by mechanisms similar to type A influenza viruses in humans, we generated reassortant viruses, by reverse genetics, that harbored molecular changes identified in genetically divergent viruses circulating in the vaccinated population. Parental and reassortant phenotype viruses were antigenically analyzed by hemagglutination inhibition (HI) test and microneutralization (MN) assay. The results of the study indicate that the antigenic drift of H5N1 in poultry is driven by multiple mutations primarily occurring in major antigenic sites at the receptor binding subdomain, similarly to what has been described for human influenza H1 and H3 subtype viruses. PMID:21734057

Modification of a deoxynivalenol-antigen-mimicking nanobody to improve immunoassay sensitivity by site-saturation mutagenesis.

PubMed

Qiu, Yu-Lou; He, Qing-Hua; Xu, Yang; Wang, Wei; Liu, Yuan-Yuan

2016-01-01

A nanobody (N-28) which can act as a deoxynivalenol (DON) antigen has been generated, and its residues Thr102-Ser106 were identified to bind with anti-DON monoclonal antibody by alanine-scanning mutagenesis. Site-saturation mutagenesis was used to analyze the plasticity of five residues and to improve the sensitivity of the N-28-based immunoassay. After mutagenesis, three mutants were selected by phage immunoassay and were sequenced. The half-maximal inhibitory concentrations of the immunoassay based on mutants N-28-T102Y, N-28-V103L, and N-28-Y105F were 24.49 ± 1.0, 51.83 ± 2.5, and 35.65 ± 1.6 ng/mL, respectively, showing the assay was, respectively, 3.2, 1.5, and 2.2 times more sensitive than the wild-type-based assay. The best mutant, N-28-T102Y, was used to develop a competitive phage ELISA to detect DON in cereals with high specificity and accuracy. In addition, the structural properties of N-28-T102Y and N-28 were investigated, revealing that the affinity of N-28-T102Y decreased because of increased steric hindrance with the large side chain. The lower-binding-affinity antigen mimetic may contribute to the improvement of the sensitivity of competitive immunoassays. These results demonstrate that nanobodies would be a favorable tool for engineering. Moreover, our results have laid a solid foundation for site-saturation mutagenesis of antigen-mimicking nanobodies to improve immunoassay sensitivity for small molecules.

Cohesin-SA1 deficiency drives aneuploidy and tumourigenesis in mice due to impaired replication of telomeres

PubMed Central

Remeseiro, Silvia; Cuadrado, Ana; Carretero, María; Martínez, Paula; Drosopoulos, William C; Cañamero, Marta; Schildkraut, Carl L; Blasco, María A; Losada, Ana

2012-01-01

Cohesin is a protein complex originally identified for its role in sister chromatid cohesion, although increasing evidence portrays it also as a major organizer of interphase chromatin. Vertebrate cohesin consists of Smc1, Smc3, Rad21/Scc1 and either stromal antigen 1 (SA1) or SA2. To explore the functional specificity of these two versions of cohesin and their relevance for embryonic development and cancer, we generated a mouse model deficient for SA1. Complete ablation of SA1 results in embryonic lethality, while heterozygous animals have shorter lifespan and earlier onset of tumourigenesis. SA1-null mouse embryonic fibroblasts show decreased proliferation and increased aneuploidy as a result of chromosome segregation defects. These defects are not caused by impaired centromeric cohesion, which depends on cohesin-SA2. Instead, they arise from defective telomere replication, which requires cohesion mediated specifically by cohesin-SA1. We propose a novel mechanism for aneuploidy generation that involves impaired telomere replication upon loss of cohesin-SA1, with clear implications in tumourigenesis. PMID:22415365

Evaluation of formalin-fixed paraffin-embedded tissues from vaccine site-associated sarcomas of cats for polyomavirus DNA and antigen.

PubMed

Kidney, B A; Haines, D M; Ellis, J A; Burnham, M; Jackson, M L

2001-06-01

To determine whether vaccine site-associated sarcomas (VSS) from cats contain polyomavirus antigen or DNA. 50 formalin-fixed paraffin-embedded tissue blocks of VSS from cats. Sections from each tissue block were evaluated for polyomavirus antigen by use of an avidin-biotin-complex immunohistochemical staining method, using rabbit anti-murine polyomavirus polyclonal antiserum as the primary antibody. The DNA was extracted from sections of each tissue block, and a polymerase chain reaction assay was performed, using primers designed to amplify regions of the bovine polyomavirus genome and consensus polyomavirus primers designed to detect unknown polyomaviruses. Polyomavirus antigen and DNA were not detected in any of the VSS. Results suggest that polyomaviruses likely do not have any direct involvement in the pathogenesis of VSS in cats.

A Modular Vaccine Development Platform Based on Sortase-Mediated Site-Specific Tagging of Antigens onto Virus-Like Particles

PubMed Central

Tang, Shubing; Xuan, Baoqin; Ye, Xiaohua; Huang, Zhong; Qian, Zhikang

2016-01-01

Virus-like particles (VLPs) can be used as powerful nanoscale weapons to fight against virus infection. In addition to direct use as vaccines, VLPs have been extensively exploited as platforms on which to display foreign antigens for prophylactic vaccination and immunotherapeutic treatment. Unfortunately, fabrication of new chimeric VLP vaccines in a versatile, site-specific and highly efficient manner is beyond the capability of traditional VLP vaccine design approaches, genetic insertion and chemical conjugation. In this study, we described a greatly improved VLP display strategy by chemoenzymatic site-specific tailoring antigens on VLPs surface with high efficiency. Through the transpeptidation mediated by sortase A, one protein and two epitopes containing N-terminal oligoglycine were conjugated to the LPET motif on the surface of hepatitis B virus core protein (HBc) VLPs with high density. All of the new chimeric VLPs induced strong specific IgG responses. Furthermore, the chimeric VLPs with sortase A tagged enterovirus 71 (EV71) SP70 epitope could elicit effective antibodies against EV71 lethal challenging as well as the genetic insertion chimeric VLPs. The sortase A mediated chemoenzymatic site-specific tailoring of the HBc VLP approach shows great potential in new VLP vaccine design for its simplicity, site specificity, high efficiency, and versatility. PMID:27170066

Identification of a common antigenic site in the nucleocapsid protein of European and North American isolates of porcine reproductive and respiratory syndrome virus.

PubMed

Casal, J I; Rodriguez, M J; Sarraseca, J; Garcia, J; Plana-Duran, J; Sanz, A

1998-01-01

Porcine reproductive and respiratory syndrome virus (PRRSV) nucleocapsid (N) protein has been identified as the most immunodominant viral protein. The N protein genes from two PRRSV isolates Olot/91 (European) and Quebec 807/94 (North American) were cloned and expressed in Escherichia coli using the pET3x system. The antigenic structure of the PRRSV N protein was dissected using seven monoclonal antibodies (MAbs) and overlapping fragments of the protein expressed in E.coli. Three antigenic sites were found. Four MAbs recognized two discontinuous epitopes that were present in the partially folded protein or at least a large fragment comprising the first 78 residues, respectively. The other three MAbs revealed the presence of a common antigenic site localized in the central region of the protein (amino acids 50 to 66). This hydrophillic region is well conserved among different isolates of European and North American origin. However, since this epitope is not recognized by many pig sera, it is not adequate for diagnostic purposes. Moreover, none of the N protein fragments were able to mimic the antigenicity of the entire N protein.

Antigenic Variation of Clade 2.1 H5N1 Virus Is Determined by a Few Amino Acid Substitutions Immediately Adjacent to the Receptor Binding Site

PubMed Central

Koel, Björn F.; van der Vliet, Stefan; Burke, David F.; Bestebroer, Theo M.; Bharoto, Eny E.; Yasa, I. Wayan W.; Herliana, Inna; Laksono, Brigitta M.; Xu, Kemin; Skepner, Eugene; Russell, Colin A.; Rimmelzwaan, Guus F.; Perez, Daniel R.; Osterhaus, Albert D. M. E.; Smith, Derek J.; Prajitno, Teguh Y.

2014-01-01

ABSTRACT Highly pathogenic avian influenza (HPAI) viruses of the H5N1 subtype are genetically highly variable and have diversified into multiple phylogenetic clades over the past decade. Antigenic drift is a well-studied phenomenon for seasonal human influenza viruses, but much less is known about the antigenic evolution of HPAI H5N1 viruses that circulate in poultry. In this study, we focused on HPAI H5N1 viruses that are enzootic to Indonesia. We selected representative viruses from genetically distinct lineages that are currently circulating and determined their antigenic properties by hemagglutination inhibition assays. At least six antigenic variants have circulated between 2003, when H5N1 clade 2.1 viruses were first detected in Indonesia, and 2011. During this period, multiple antigenic variants cocirculated in the same geographic regions. Mutant viruses were constructed by site-directed mutagenesis to represent each of the circulating antigenic variants, revealing that antigenic differences between clade 2.1 viruses were due to only one or very few amino acid substitutions immediately adjacent to the receptor binding site. Antigenic variants of H5N1 virus evaded recognition by both ferret and chicken antibodies. The molecular basis for antigenic change in clade 2.1 viruses closely resembled that of seasonal human influenza viruses, indicating that the hemagglutinin of influenza viruses from different hosts and subtypes may be similarly restricted to evade antibody recognition. PMID:24917596

Superposition-free comparison and clustering of antibody binding sites: implications for the prediction of the nature of their antigen

PubMed Central

Di Rienzo, Lorenzo; Milanetti, Edoardo; Lepore, Rosalba; Olimpieri, Pier Paolo; Tramontano, Anna

2017-01-01

We describe here a superposition free method for comparing the surfaces of antibody binding sites based on the Zernike moments and show that they can be used to quickly compare and cluster sets of antibodies. The clusters provide information about the nature of the bound antigen that, when combined with a method for predicting the number of direct antibody antigen contacts, allows the discrimination between protein and non-protein binding antibodies with an accuracy of 76%. This is of relevance in several aspects of antibody science, for example to select the framework to be used for a combinatorial antibody library. PMID:28338016

Alanine scanning of the rabies virus glycoprotein antigenic site III using recombinant rabies virus: implication for post-exposure treatment.

PubMed

Papaneri, Amy B; Wirblich, Christoph; Marissen, Wilfred E; Schnell, Matthias J

2013-12-02

The safety and availability of the human polyclonal sera that is currently utilized for post-exposure treatment (PET) of rabies virus (RABV) infection remain a concern. Recombinant monoclonal antibodies have been postulated as suitable alternatives by WHO. To this extent, CL184, the RABV human antibody combination comprising monoclonal antibodies (mAbs) CR57 and CR4098, has been developed and has delivered promising clinical data to support its use for RABV PET. For this fully human IgG1 cocktail, mAbs CR57 and CR4098 are produced in the PER.C6 human cell line and combined in equal amounts in the final product. During preclinical evaluation, CR57 was shown to bind to antigenic site I whereas CR4098 neutralization was influenced by a mutation of position 336 (N336) located within antigenic site III. Here, alanine scanning was used to analyze the influence of mutations within the potential binding site for CR4098, antigenic site III, in order to evaluate the possibility of mutated rabies viruses escaping neutralization. For this approach, twenty flanking amino acids (10 upstream and 10 downstream) of the RABV glycoprotein (G) asparagine (N336) were exchanged to alanine (or serine, if already alanine) by site-directed mutagenesis. Analysis of G expression revealed four of the twenty mutant Gs to be non-functional, as shown by their lack of cell surface expression, which is a requirement for the production of infectious RABV. Therefore, these mutants were excluded from further study. The remaining sixteen mutants were introduced in an infectious clone of RABV, and recombinant RABVs (rRABVs) were recovered and utilized for in vitro neutralization assays. All of the viruses were effectively neutralized by CR4098 as well as by CR57, indicating that single amino acid exchanges in this region does not affect the broad neutralizing capability of the CL184 mAb combination. Copyright © 2013 Elsevier Ltd. All rights reserved.

Evaluation of formalin-fixed paraffin-embedded tissues from vaccine site-associated sarcomas of cats for papillomavirus DNA and antigen.

PubMed

Kidney, B A; Haines, D M; Ellis, J A; Burnham, M L; Teifke, J P; Czerwinski, G; Jackson, M L

2001-06-01

To determine whether vaccine site-associated sarcomas (VSS) from cats contain papillomavirus antigen or DNA. 50 formalin-fixed paraffin-embedded tissue blocks of VSS from cats. Sections from each tissue block were evaluated for papillomavirus antigen by use of an avidin-biotin-complex immunohistochemical staining method, using rabbit anti-bovine papillomavirus type-1 antibody. The DNA was extracted from sections of each tissue block, and polymerase chain reaction assays were performed, using primers designed to amplify regions of the E5 gene of bovine papillomavirus and consensus primers designed to amplify a region of the L1 gene of animal papillomaviruses. Sections from 20 of the tissue blocks were evaluated by use of nonradioactive in situ hybridization for bovine papillomavirus DNA. Papillomavirus antigen and DNA were not detected in any of the VSS. Results suggest that papillomaviruses likely do not have any direct involvement in the pathogenesis of VSS in cats.

Antigen injection (image)

MedlinePlus

Leprosy is caused by the organism Mycobacterium leprae . The leprosy test involves injection of an antigen just under ... if your body has a current or recent leprosy infection. The injection site is labeled and examined ...

Suppression or knockout of SaF/SaM overcomes the Sa-mediated hybrid male sterility in rice.

PubMed

Xie, Yongyao; Niu, Baixiao; Long, Yunming; Li, Gousi; Tang, Jintao; Zhang, Yaling; Ren, Ding; Liu, Yao-Guang; Chen, Letian

2017-09-01

Hybrids between the indica and japonica subspecies of rice (Oryza sativa) are usually sterile, which hinders utilization of heterosis in the inter-subspecific hybrid breeding. The complex locus Sa comprises two adjacently located genes, SaF and SaM, which interact to cause abortion of pollen grains carrying the japonica allele in japonica-indica hybrids. Here we showed that silencing of SaF or SaM by RNA interference restored male fertility in indica-japonica hybrids with heterozygous Sa. We further used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-based genome editing to knockout the SaF and SaM alleles, respectively, of an indica rice line to create hybrid-compatible lines. The resultant artificial neutral alleles did not affect pollen viability and other agricultural traits, but did break down the reproductive barrier in the hybrids. We found that some rice lines have natural neutral allele Sa-n, which was compatible with the typical japonica or indica Sa alleles in hybrids. Our results demonstrate that SaF and SaM are required for hybrid male sterility, but are not essential for pollen development. This study provides effective approaches for the generation of hybrid-compatible lines by knocking out the Sa locus or using the natural Sa-n allele to overcome hybrid male sterility in rice breeding. © 2017 The Authors. Bioelectromagnetics published by Wiley Periodicals, Inc. © 2017 Institute of Botany, Chinese Academy of Sciences.

Recent progress of the JT-60SA project

NASA Astrophysics Data System (ADS)

Shirai, H.; Barabaschi, P.; Kamada, Y.; the JT-60SA Team

2017-10-01

The JT-60SA project has been implemented for the purpose of an early realization of fusion energy. With a powerful and versatile NBI and ECRF system, a flexible plasma-shaping capability, and various kinds of in-vessel coils to suppress MHD instabilities, JT-60SA plays an essential role in addressing the key physics and engineering issues of ITER and DEMO. It aims to achieve the long sustainment of high integrated performance plasmas under the high β N condition required in DEMO. The fabrication and installation of components and systems of JT-60SA procured by the EU and Japan are steadily progressing. The installation of toroidal field (TF) coils around the vacuum vessel started in December 2016. The commissioning of the cryogenic system and power supply system has been implemented in the Naka site, and JT-60SA will start operation in 2019. The JT-60SA research plan covers a wide area of issues in ITER and DEMO relevant operation regimes, and has been regularly updated on the basis of intensive discussion among European and Japanese researchers.

Isolation of a High Affinity Neutralizing Monoclonal Antibody against 2009 Pandemic H1N1 Virus That Binds at the ‘Sa’ Antigenic Site

PubMed Central

Mishra, Arpita; Yeolekar, Leena; Dhere, Rajeev; Kapre, Subhash; Varadarajan, Raghavan; Gupta, Satish Kumar

2013-01-01

Influenza virus evades host immunity through antigenic drift and shift, and continues to circulate in the human population causing periodic outbreaks including the recent 2009 pandemic. A large segment of the population was potentially susceptible to this novel strain of virus. Historically, monoclonal antibodies (MAbs) have been fundamental tools for diagnosis and epitope mapping of influenza viruses and their importance as an alternate treatment option is also being realized. The current study describes isolation of a high affinity (K D = 2.1±0.4 pM) murine MAb, MA2077 that binds specifically to the hemagglutinin (HA) surface glycoprotein of the pandemic virus. The antibody neutralized the 2009 pandemic H1N1 virus in an in vitro microneutralization assay (IC50 = 0.08 µg/ml). MA2077 also showed hemagglutination inhibition activity (HI titre of 0.50 µg/ml) against the pandemic virus. In a competition ELISA, MA2077 competed with the binding site of the human MAb, 2D1 (isolated from a survivor of the 1918 Spanish flu pandemic) on pandemic H1N1 HA. Epitope mapping studies using yeast cell-surface display of a stable HA1 fragment, wherein ‘Sa’ and ‘Sb’ sites were independently mutated, localized the binding site of MA2077 within the ‘Sa’ antigenic site. These studies will facilitate our understanding of antigen antibody interaction in the context of neutralization of the pandemic influenza virus. PMID:23383214

Case report: diffuse splenic metastasis of occult breast cancer with incompatible blood group antigenic determinants.

PubMed

Baranyay, Ferenc

2009-01-01

Cancer cells with immunogenic properties having altered protein glycosilation, modified blood group substances have been widely studied [Kannagi R, Miyake M, Zenita KM, Itai S, Hiraiwa N, Shigeta K, et al. Cancer-associated carbohydrate antigens: modified blood group substances and oncodevelopmental antigens on tumor cells. Gann Monogr Cancer Res 1988; 34: p. 15-28; Hakomori S. Antigen structure and genetic basis of histo-blood groups A, B and O their changes associated with human cancer. Biochem Biophys Acta 1999; 1473: p. 247-266; Brooks SA, Carter TM, Royle L, Harvey DJ, Fry SA, Kinch C, et al. Altered glycosilation of proteins in cancer: what is the potential for new anti-tumour strategies. Anticancer Agents Med Chem 2008; 8: p. 2-21]. In the study reported here, a 78-year-old female patient was admitted to the hospital with circulatory failure. At autopsy, the spleen (weight: 420 g) was extremely firm with a diffusely blackberry-colored cut surface. There were no signs of carcinomatous process at autopsy. By histology, the spleen showed diffuse metastatic carcinomatous infiltration. Using immunohistochemistry, an antibody to breast carcinoma antigen (BioGenex) labelled metastatic cells of the spleen and bone marrow. The patient was blood group O. Labelling for binding of lectins with and without blood group antigen specificity and monoclonal antibodies was carried out. The B blood group specific Banderiaea simplicifolia agglutinin I and an anti-B blood group monoclonal antibody labelled all the metastatic cells of spleen and bone marrow intensely. There was no detection of blood group A antigen by either binding of Dolichos biflorus agglutinin or anti-blood group A monoclonal antibodies. These observations raise the possibility that the detected incompatible B blood group antigen determinants on the metastatic cells were immunogenic. The surviving carcinoma cells may have found a place of refuge from immune surveillance in the spleen and in the bone marrow

Targeting of plant-derived vaccine antigens to immunoresponsive mucosal sites.

PubMed

Rigano, M Manuela; Sala, Francesco; Arntzen, Charles J; Walmsley, Amanda M

2003-01-30

Most pathogenic microorganisms enter their host via the mucosal surfaces lining the digestive, respiratory and urino-reproductive tracts of the body. The most efficient means of protecting these surfaces is through mucosal immunization. Transgenic plants are safe and inexpensive vehicles to produce and mucosally deliver protective antigens. However, the application of this technology is limited by the poor response of the immune system to non-particulate, subunit vaccines. Co-delivery of therapeutic proteins with targeting proteins, such as the B subunit of the Escherichia coli heat labile enterotoxin (LTB), could increase the effectiveness of such antigens.

Expression of simian virus 40 T antigen in Escherichia coli: localization of T-antigen origin DNA-binding domain to within 129 amino acids.

PubMed Central

Arthur, A K; Höss, A; Fanning, E

1988-01-01

The genomic coding sequence of the large T antigen of simian virus 40 (SV40) was cloned into an Escherichia coli expression vector by joining new restriction sites, BglII and BamHI, introduced at the intron boundaries of the gene. Full-length large T antigen, as well as deletion and amino acid substitution mutants, were inducibly expressed from the lac promoter of pUC9, albeit with different efficiencies and protein stabilities. Specific interaction with SV40 origin DNA was detected for full-length T antigen and certain mutants. Deletion mutants lacking T-antigen residues 1 to 130 and 260 to 708 retained specific origin-binding activity, demonstrating that the region between residues 131 and 259 must carry the essential binding domain for DNA-binding sites I and II. A sequence between residues 302 and 320 homologous to a metal-binding "finger" motif is therefore not required for origin-specific binding. However, substitution of serine for either of two cysteine residues in this motif caused a dramatic decrease in origin DNA-binding activity. This region, as well as other regions of the full-length protein, may thus be involved in stabilizing the DNA-binding domain and altering its preference for binding to site I or site II DNA. Images PMID:2835505

The genetic origin of minor histocompatibility antigens.

PubMed

Roopenian, D C; Christianson, G J; Davis, A P; Zuberi, A R; Mobraaten, L E

1993-01-01

The purpose of this study was to elucidate the genetic origin of minor histocompatibility (H) antigens. Toward this end common inbred mouse strains, distinct subspecies, and species of the subgenus Mus were examined for expression of various minor H antigens. These antigens were encoded by the classical minor H loci H-3 and H-4 or by newly identified minor H antigens detected as a consequence of mutation. Both minor H antigens that stimulate MHC class I-restricted cytotoxic T cells (Tc) and antigens that stimulate MHC class II-restricted helper T cells (Th) were monitored. The results suggested that strains of distinct ancestry commonly express identical or cross-reactive antigens. Moreover, a correlation between the lack of expression of minor H antigens and ancestral heritage was observed. To address whether the antigens found on unrelated strains were allelic with the sensitizing minor H antigens or a consequence of antigen cross-reactivity, classical genetic segregation analysis was carried out. Even in distinct subspecies and species, the minor H antigens always mapped to the site of the appropriate minor H locus. Together the results suggest: 1) minor H antigen sequences are evolutionarily stable in that their pace of antigenic change is slow enough to predate subspeciation and speciation; 2) the minor H antigens originated in the inbred strains as a consequence of a rare polymorphism or loss mutation carried in a founder mouse stock that caused the mouse to perceive the wild-type protein as foreign; 3) there is a remarkable lack of antigenic cross-reactivity between the defined minor H antigens and other gene products.

Glycosylations in the globular head of the hemagglutinin protein modulate the virulence and antigenic properties of the H1N1 influenza viruses

PubMed Central

Medina, Rafael A.; Stertz, Silke; Manicassamy, Balaji; Zimmermann, Petra; Sun, Xiangjie; Albrecht, Randy A.; Uusi-Kerttula, Hanni; Zagordi, Osvaldo; Belshe, Robert B.; Frey, Sharon E.; Eggink, Dirk; Tumpey, Terrence M.; García-Sastre, Adolfo

2014-01-01

The global spread of the 2009 pandemic H1N1 (pH1N1) virus in humans increases the likelihood that this influenza virus strain could undergo antigenic drift in the coming years. Previous seasonal H1N1 and H3N2 influenza strains acquired additional glycosylations in the globular head of their hemagglutinin (HA) proteins as they evolved over time; these are believed to shield antigenically relevant regions. We used influenza A/Netherlands/602/2009 recombinant (rpH1N1) viruses to which we added additional HA glycosylation sites reflecting their temporal appearance in previous seasonal H1N1 viruses. Additional glycosylations resulted in substantial attenuation in mice and ferrets, while deleting HA glycosylation sites from a pre-pandemic 1991 seasonal H1N1 influenza virus resulted in increased pathogenicity in mice. Sera from mice infected with wild type (WT) rpH1N1 virus showed a considerable loss of HA inhibitory (HI) activity against rpH1N1 viruses glycosylated at sites 144 or 144-172, indicating that the polyclonal antibody response elicited by WT rpH1N1 HA seems to be directed against an immunodominant region, likely site Sa, shielded by glycosylation at 144. Sera from humans vaccinated with the pH1N1 inactivated vaccine also showed reduced activity against the 144 and 144-172 mutant viruses. Remarkably, the HI activity of sera from virus-infected mice demonstrated that glycosylation at position 144 resulted in the induction of a broader polyclonal response able to cross-neutralize all WT and glycosylation mutant pH1N1 viruses. Mice infected with a recent seasonal virus in which glycosylation sites 71, 142 and 177 were removed, elicited antibodies that protected against challenge with the antigenically distant pH1N1 virus. Thus, acquisition of glycosylation sites in the HA of H1N1 human influenza viruses not only affects their pathogenicity and ability to escape from polyclonal antibodies elicited by previous influenza virus strains, but also their ability to

Application of the dengue virus NS1 antigen rapid test for on-site detection of imported dengue cases at airports.

PubMed

Shu, Pei-Yun; Yang, Cheng-Fen; Kao, Jeng-Fong; Su, Chien-Ling; Chang, Shu-Fen; Lin, Chien-Chou; Yang, Wen-Chih; Shih, Hsiu; Yang, Shih-Yan; Wu, Ping-Fuai; Wu, Ho-Sheng; Huang, Jyh-Hsiung

2009-04-01

We used the dengue virus NS1 antigen (Ag) rapid test for on-site detection of imported dengue cases at airports. Among 22 positive cases of dengue identified from 850 patients with a fever suspected to have dengue, 17 were NS1 Ag test positive. These findings demonstrate the usefulness of the NS1 Ag rapid test in screening imported dengue cases at airports.

Application of the Dengue Virus NS1 Antigen Rapid Test for On-Site Detection of Imported Dengue Cases at Airports▿

PubMed Central

Shu, Pei-Yun; Yang, Cheng-Fen; Kao, Jeng-Fong; Su, Chien-Ling; Chang, Shu-Fen; Lin, Chien-Chou; Yang, Wen-Chih; Shih, Hsiu; Yang, Shih-Yan; Wu, Ping-Fuai; Wu, Ho-Sheng; Huang, Jyh-Hsiung

2009-01-01

We used the dengue virus NS1 antigen (Ag) rapid test for on-site detection of imported dengue cases at airports. Among 22 positive cases of dengue identified from 850 patients with a fever suspected to have dengue, 17 were NS1 Ag test positive. These findings demonstrate the usefulness of the NS1 Ag rapid test in screening imported dengue cases at airports. PMID:19193828

Mimicry of the immunodominant conformation-dependent antigenic site of hepatitis A virus by motifs selected from synthetic peptide libraries.

PubMed

Mattioli, S; Imberti, L; Stellini, R; Primi, D

1995-09-01

Hepatitis A virus (HAV) is a positive-strand RNA virus with a genome length of approximately 7,480 nucleotides. Although HAV morphogenesis is thought to be similar to that of poliovirus, the prototype picornavirus, the complete characterization of the antigenic structure of this virus remains elusive. All the available evidences, however, support the existence, on HAV virions and empty capsids, of an immunodominant neutralization antigenic site which is conformation dependent and whose structure involves residues of both VP1 and VP3 capsid proteins. This particular feature and the difficulty of obtaining high virus yield in tissue cultures make HAV an ideal target for developing synthetic peptides that simulate the structure of its main antigenic determinant. To this end we utilized, in the present work, the divide-couple-recombine approach to generate a random library composed of millions of different hexapeptides. This vast library was screened with a well-characterized anti-HAV monoclonal antibody. By this strategy we identified a peptide that reacted specifically with monoclonal and polyclonal anti-HAV antibodies and, in mice, induced a specific anti-virus immune response. Furthermore, the peptide could also be used in an enzyme-linked immunosorbent assay for revealing a primary immunoglobulin M immune response in sera of acutely infected human patients. Interestingly, no sequence homology was found between the identified peptide and the HAV capsid proteins VP1 and VP3. Collectively, these data represent an additional important paradigm of a mimotope capable of mimicking an antigenic determinant with unknown tertiary structure.

Editing Citrus Genome via SaCas9/sgRNA System

PubMed Central

Jia, Hongge; Xu, Jin; Orbović, Vladimir; Zhang, Yunzeng; Wang, Nian

2017-01-01

SaCas9/sgRNA, derived from Staphylococcus aureus, is an alternative system for genome editing to Streptococcus pyogenes SpCas9/sgRNA. The smaller SaCas9 recognizes a different protospacer adjacent motif (PAM) sequence from SpCas9. SaCas9/sgRNA has been employed to edit the genomes of Arabidopsis, tobacco and rice. In this study, we aimed to test its potential in genome editing of citrus. Transient expression of SaCas9/sgRNA in Duncan grapefruit via Xcc-facilitated agroinfiltration showed it can successfully modify CsPDS and Cs2g12470. Subsequently, binary vector GFP-p1380N-SaCas9/35S-sgRNA1:AtU6-sgRNA2 was developed to edit two target sites of Cs7g03360 in transgenic Carrizo citrange. Twelve GFP-positive Carrizo transformants were successfully established, designated as #Cz1 to #Cz12. Based on targeted next generation sequencing results, the mutation rates for the two targets ranged from 15.55 to 39.13% for sgRNA1 and 49.01 to 79.67% for sgRNA2. Therefore, SaCas9/sgRNA can be used as an alternative tool to SpCas9/sgRNA for citrus genome editing. PMID:29312390

Masked Chimeric Antigen Receptor for Tumor-Specific Activation.

PubMed

Han, Xiaolu; Bryson, Paul D; Zhao, Yifan; Cinay, Gunce E; Li, Si; Guo, Yunfei; Siriwon, Natnaree; Wang, Pin

2017-01-04

Adoptive cellular therapy based on chimeric antigen receptor (CAR)-engineered T (CAR-T) cells is a powerful form of cancer immunotherapy. CAR-T cells can be redirected to specifically recognize tumor-associated antigens (TAAs) and induce high levels of antitumor activity. However, they may also display "on-target off-tumor" toxicities, resulting from low-level expression of TAAs in healthy tissues. These adverse effects have raised considerable safety concerns and limited the clinical application of this otherwise promising therapeutic modality. To minimize such side effects, we have designed an epidermal growth factor receptor (EGFR)-specific masked CAR (mCAR), which consists of a masking peptide that blocks the antigen-binding site and a protease-sensitive linker. Proteases commonly active in the tumor microenvironment can cleave the linker and disengage the masking peptide, thereby enabling CAR-T cells to recognize target antigens only at the tumor site. In vitro mCAR showed dramatically reduced antigen binding and antigen-specific activation in the absence of proteases, but normal levels of binding and activity upon treatment with certain proteases. Masked CAR-T cells also showed antitumor efficacy in vivo comparable to that of unmasked CAR. Our study demonstrates the feasibility of improving the safety profile of conventional CARs and may also inspire future design of CAR molecules targeting broadly expressed TAAs. Copyright © 2017 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

Tuberculous Lymphadenitis Is Associated with Enhanced Baseline and Antigen-Specific Induction of Type 1 and Type 17 Cytokines and Reduced Interleukin-1β (IL-1β) and IL-18 at the Site of Infection.

PubMed

Kathamuthu, Gokul Raj; Moideen, Kadar; Baskaran, Dhanaraj; Banurekha, Vaithilingam V; Nair, Dina; Sekar, Gomathi; Sridhar, Rathinam; Vidyajayanthi, Bharathi; Gajendraraj, Ganeshan; Parandhaman, Dinesh Kumar; Srinivasan, Alena; Babu, Subash

2017-05-01

Tuberculous lymphadenitis (TBL) is characterized by an expansion of Th1 and Th17 cells with altered serum levels of proinflammatory cytokines. However, the cytokine profile at the site of infection, i.e., the affected lymph nodes, has not been examined in detail. To estimate the baseline and mycobacterial antigen-stimulated concentrations of type 1, type 17, and other proinflammatory cytokines in patients with TBL ( n = 14), we examined both the baseline and the antigen-specific concentrations of these cytokines before and after chemotherapy and compared them with those in individuals with pulmonary tuberculosis (PTB) ( n = 14). In addition, we also compared the cytokine responses in whole blood and those in the lymph nodes of TBL individuals. We observed significantly enhanced baseline and antigen-specific levels of type 1 cytokines (gamma interferon [IFN-γ] and tumor necrosis factor alpha [TNF-α]) and a type 17 cytokine (interleukin-17 [IL-17]) and significantly diminished baseline and antigen-specific levels of proinflammatory cytokines (IL-1β and IL-18) in the whole blood of TBL individuals compared to those in the whole blood of PTB individuals. Moreover, we also observed a pattern of baseline and antigen-specific cytokine production at the site of infection (lymph node) similar to that in the whole blood of TBL individuals. Following standard antituberculosis (anti-TB) treatment, we observed alterations in the baseline and/or antigen-specific levels of IFN-γ, TNF-α, IL-1β, and IL-18. TBL is therefore characterized by enhanced baseline and antigen-specific production of type 1 and type 17 cytokines and reduced baseline and antigen-specific production of IL-1β and IL-18 at the site of infection. Copyright © 2017 American Society for Microbiology.

Lipopolysaccharide Antigens of Pseudomonas aeruginosa and Design of Novel Vaccines.

DTIC Science & Technology

1987-09-01

Pseudomonas aeruginosa, OA 1-C LChemical structure, Fisher immunotypes, M; ig0-Chain polysaccharide , and Synthetic antigens 19. ABSTRACT (Conu on rftvm if...have been characterized in our laboratories. Partial structures for the remaining two types have been elucidated. The O-chain polysaccharides of the... polysaccharide antigens for native structure, and (5) binding-site xa[lJ11:, of the antibodies using the synthetic antigens. b% B.. Sirmificance: General

Simian virus 40 T-antigen-related cell surface antigen: serological demonstration on simian virus 40-transformed monolayer cells in situ.

PubMed Central

Deppert, W; Hanke, K; Henning, R

1980-01-01

Simian virus 40 (SV40)-transformed monolayer cells were analyzed in situ by indirect immunofluorescence microscopy for the postulated cell surface location of SV40 T-antigen-related molecules. With antisera prepared against purified, sodium dodecyl sulfate-denatured SV40 T-antigen, positive surface staining was obtained when the cells had been treated with formaldehyde before immunofluorescence analysis. In contrast, living SV40-transformed cells analyzed in monolayer were surface fluorescence negative. The fixation procedure developed in this study combined with a double staining immunofluorescence technique allowed the simultaneous analysis of the same cells for the expression of both SV40 T-antigen-related surface antigen and nuclear T-antigen. The localization of SV40 T-antigen-related surface antigen on the outer surface of the plasma membrane of formaldehyde-fixed SV40-transformed cells was demonstrated directly by the protein A-mediated binding of Staphylococcus aureus bacteria on formaldehyde-fixed SV40-transformed cells precoated with antiserum against sodium dodecyl sulfate-denatured T-antigen. Both cell surface staining and S. aureus binding were found to be highly specific for SV40 T-antigen-related binding sites. These results indicate that T-antigen-related molecules in a cryptic form are located on the surface of SV40-transformed monolayer cells and can be detected in situ after modification of the cell surface architecture. Images PMID:6255189

Polyoma virus small tumor antigen pre-mRNA splicing requires cooperation between two 3' splice sites.

PubMed Central

Ge, H; Noble, J; Colgan, J; Manley, J L

1990-01-01

We have studied splicing of the polyoma virus early region pre-mRNA in vitro. This RNA is alternatively spliced in vivo to produce mRNA encoding the large, middle-sized (MTAg), and small (StAg) tumor antigens. Our primary interest was to learn how the 48-nucleotide StAg intron is excised, because the length of this intron is significantly less than the apparent minimum established for mammalian introns. Although the products of all three splices are detected in vitro, characterization of the pathway and sequence requirements of StAg splicing suggests that splicing factors interact with the precursor RNA in an unexpected way to catalyze removal of this intron. Specifically, StAg splicing uses either of two lariat branch points, one of which is located only 4 nucleotides from the 3' splice site. Furthermore, the StAg splice absolutely requires that the alternative MTAg 3' splice site, located 14 nucleotides downstream of the StAg 3' splice site, be intact. Insertion mutations that increase or decrease the quality of the MTAg pyrimidine stretch enhance or repress StAg as well as MTAg splicing, and a single-base change in the MTAg AG splice acceptor totally blocks both splices. These results demonstrate the ability of two 3' splice sites to cooperate with each other to bring about removal of a single intron. Images PMID:2159146

CD34 Antigen: Determination of Specific Sites of Phosphorylation In Vitro and In Vivo¶

PubMed Central

Deterding, Leesa J.; Williams, Jason G.; Humble, Margaret M.; Petrovich, Robert M.; Wei, Sung-Jen; Trempus, Carol S.; Gates, Matthew B.; Zhu, Feng; Smart, Robert C.; Tennant, Raymond W.; Tomer, Kenneth B.

2010-01-01

CD34, a type I transmembrane glycoprotein, is a surface antigen which is expressed on several cell types, including hematopoietic progenitors, endothelial cells, as well as mast cells. Recently, CD34 has been described as a marker for epidermal stem cells in mouse hair follicles, and is expressed in outer root sheath cells of the human hair follicle. Although the biological function and regulation of CD34 is not well understood, it is thought to be involved in cell adhesion as well as possibly having a role in signal transduction. In addition, CD34 was shown to be critical for skin tumor development in mice, although the exact mechanism remains unknown. Many proteins' functions and biological activities are regulated through post-translational modifications. The extracellular domain of CD34 is heavily glycosylated but the role of these glycans in CD34 function is unknown. Additionally, two sites of tyrosine phosphorylation have been reported on human CD34 and it is known that CD34 is phosphorylated, at least in part, by protein kinase C; however, the precise location of the sites of phosphorylation has not been reported. In an effort to identify specific phosphorylation sites in CD34 and delineate the possible role of protein kinase C, we undertook the identification of the in vitro sites of phosphorylation on the intracellular domain of mouse CD34 (aa 309–382) following PKC treatment. For this work, we are using a combination of enzymatic proteolysis and peptide sequencing by mass spectrometry. After which the in vivo sites of phosphorylation of full-length mouse CD34 expressed from HEK293F cells were determined. The observed in vivo sites of phosphorylation, however, are not consensus PKC sites, but our data indicate that one of these sites may possibly be phosphorylated by AKT2. These results suggest that other kinases, as well as PKC, may have important signaling functions in CD34. PMID:21499536

Present status and future prospects of the JT-60SA project

NASA Astrophysics Data System (ADS)

Ishida, S.; Barabaschi, P.; Kamada, Y.

2014-10-01

The JT-60SA project has been implemented jointly by Europe and Japan since June 2007. After the disassembly of JT-60 from the torus hall had been completed in October 2012, the project achieved the major milestone of starting the tokamak's assembly at the JAEA Naka site in January 2013 following the completion of the cryostat base in Europe and its transport to Japan. Procurement and assembly activities for components such as the superconducting magnet, cryogenic system, power supply, vacuum vessel, divertor and cryostat are progressing on track towards the start of operation in March 2019. In preparation for exploitation, the JT-60SA Research Plan was issued in December 2011, and the research integration activities are addressing JT-60SA data management, validation and analysis tools. This paper overviews the latest evolution of the project in terms of construction and exploitation for JT-60SA.

Structural Insights into the Protease-like Antigen Plasmodium falciparum SERA5 and Its Noncanonical Active-Site Serine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hodder, Anthony N.; Malby, Robyn L.; Clarke, Oliver B.

The sera genes of the malaria-causing parasite Plasmodium encode a family of unique proteins that are maximally expressed at the time of egress of parasites from infected red blood cells. These multi-domain proteins are unique, containing a central papain-like cysteine-protease fragment enclosed between the disulfide-linked N- and C-terminal domains. However, the central fragment of several members of this family, including serine repeat antigen 5 (SERA5), contains a serine (S596) in place of the active-site cysteine. Here we report the crystal structure of the central protease-like domain of Plasmodium falciparum SERA5, revealing a number of anomalies in addition to the putativemore » nucleophilic serine: (1) the structure of the putative active site is not conducive to binding substrate in the canonical cysteine-protease manner; (2) the side chain of D594 restricts access of substrate to the putative active site; and (3) the S{sub 2} specificity pocket is occupied by the side chain of Y735, reducing this site to a small depression on the protein surface. Attempts to determine the structure in complex with known inhibitors were not successful. Thus, despite having revealed its structure, the function of the catalytic domain of SERA5 remains an enigma.« less

Natural selection promotes antigenic evolvability.

PubMed

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.

Natural Selection Promotes Antigenic Evolvability

PubMed Central

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

Heterogeneous distribution of antigens on human platelets demonstrated by fluorescence flow cytometry.

PubMed

Dunstan, R A; Simpson, M B

1985-12-01

We have used fluorescence flow cytometry to analyse cell-to-cell variability in the density of platelet ABH, Ii, Lewis, P, P1A1, Bak,a and HLA class I antigens. Human IgG and IgM antibodies were used in a two-stage assay with goat FITC-conjugated antihuman IgG (H&L) antibody as the label, followed by single cell analysis of 10 000 platelets per sample using a 256-channel fluorescence flow cytometer (Becton-Dickinson FACS Analyser). Computer analysis of fluorescence intensity histograms for mean and peak channel and coefficient of variation shows that the degree of heterogeneity in platelet antigen density varies with each particular blood group. The broad fluorescence distribution curves with oligosaccharide antigens (CVs: A = 53, B = 40, I = 44, Lea = 40, P = 40) indicate that these antigens possess a greater variability in the number of sites per cell compared to the more homogeneous distribution of P1,A1 BaK,a and HLA (CVs: P1A1 = 24, HLA = 30). These findings may partly account for the mechanism by which transfusion of ABO-incompatible platelets results in a biphasic survival curve, with a period of early rapid removal of those platelets with a high density of antigen sites, followed by a relatively normal survival curve for those platelets that possess only a few or no antigen sites. In contrast, P1A1 and HLA sites are less variable in number from one platelet to another in a given donor, and immune-mediated removal would be more likely to approximate a single exponential curve.

Antigenic characterization of a formalin-inactivated poliovirus vaccine derived from live-attenuated Sabin strains.

PubMed

Tano, Yoshio; Shimizu, Hiroyuki; Martin, Javier; Nishimura, Yorihiro; Simizu, Bunsiti; Miyamura, Tatsuo

2007-10-10

A candidate inactivated poliovirus vaccine derived from live-attenuated Sabin strains (sIPV), which are used in the oral poliovirus vaccine (OPV), was prepared in a large-production scale. The modification of viral antigenic epitopes during the formalin inactivation process was investigated by capture ELISA assays using type-specific and antigenic site-specific monoclonal antibodies (MoAbs). The major antigenic site 1 was modified during the formalin inactivation of Sabin 1. Antigenic sites 1-3 were slightly modified during the formalin inactivation of Sabin 2 strain. Sites 1 and 3 were altered on inactivated Sabin 3 virus. These alterations were different to those shown by wild-type Saukett strain, used in conventional IPV (cIPV). It has been previously reported that type 1 sIPV showed higher immunogenicity to type 1 cIPV whereas types 2 and 3 sIPV induced lower level of immunogenicity than their cIPV counterparts. Our results suggest that the differences in epitope structure after formalin inactivation may account, at least in part, for the observed differences in immunogenicity between Sabin and wild-type inactivated poliovaccines.

AntigenMap 3D: an online antigenic cartography resource.

PubMed

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

Humoral immunity targeting site I of antigenic domain 2 of glycoprotein B upon immunization with different cytomegalovirus candidate vaccines.

PubMed

Axelsson, Fredrika; Adler, Stuart P; Lamarre, Alain; Ohlin, Mats

2007-12-21

Glycoprotein B (gB) is a major component in several vaccines that are under development for prevention of disease by cytomegalovirus. It contains multiple determinants that are targets for neutralizing antibodies. One of them is site I of antigenic domain 2 (AD-2). The epitope, defined by short peptides, is quite conserved between different isolates. However, it is poorly immunogenic in natural infection. In this study we investigated the extent to which different vaccines, attenuated live Towne vaccine with or without priming with a canarypox virus coding for gB, or a recombinant gB vaccine adjuvanted with MF59, induced antibodies to this epitope. As in natural infection only a fraction of all subjects developed antibody responses against site I of AD-2 following vaccination. We suggest that strategies that enhance immunogenicity of this epitope will improve vaccine efficacy.

Cancer vaccine--Antigenics.

PubMed

2002-01-01

Antigenics is developing a therapeutic cancer vaccine based on heat-shock proteins (HSPs). The vaccine [HSPPC-96, Oncophage] is in a pivotal phase III clinical trial for renal cancer at 80 clinical sites worldwide. The trial is enrolling at least 500 patients who are randomised to receive surgical removal of the primary tumour followed by out-patient treatment with Oncophage((R)) or surgery only. This study was initiated on the basis of results from a pilot phase I/II study and preliminary results from a phase II study in patients with renal cell cancer. In October 2001, Oncophage was designated as a fast-track product by the Food and Drug Administration in the US for the treatment of renal cell carcinoma. Oncophage is in phase I/II trials in Italy for colorectal cancer (30 patients) and melanoma. The trials in Italy are being conducted at the Istituto dei Tumouri, Milan (in association with Sigma-Tau). Preliminary data from the phase II trial for melanoma was presented at the AACR-NCI-EORTC International Conference in Florida, USA, in October 2001. Oncophage is also in a phase I/II (42 patients) and a phase II trial (84 patients) in the US for renal cell cancer, a phase II trial in the US for non-Hodgkin's lymphoma (35 patients), a phase II trial in the US for sarcoma (20-35 patients), a phase I/II trial in the US for melanoma (36 patients), and phase I/II trials in Germany for gastric (30 patients) and pancreatic cancers. A pilot phase I trial in patients with pancreatic cancer began in the US in 1997 with 5 patients enrolled. In November 2000, Antigenics announced that this trial had been expanded to a phase I/II study which would now include survival as an endpoint and would enroll 5 additional patients. The US trials are being performed at Memorial Sloan-Kettering Cancer Center and the M.D. Anderson Cancer Center. The trials in Germany are being carried out at Johannes Gutenberg-University Hospital, Mainz. Oncophage is an autologous vaccine consisting of

Local and global anatomy of antibody-protein antigen recognition.

PubMed

Wang, Meryl; Zhu, David; Zhu, Jianwei; Nussinov, Ruth; Ma, Buyong

2018-05-01

Deciphering antibody-protein antigen recognition is of fundamental and practical significance. We constructed an antibody structural dataset, partitioned it into human and murine subgroups, and compared it with nonantibody protein-protein complexes. We investigated the physicochemical properties of regions on and away from the antibody-antigen interfaces, including net charge, overall antibody charge distributions, and their potential role in antigen interaction. We observed that amino acid preference in antibody-protein antigen recognition is entropy driven, with residues having low side-chain entropy appearing to compensate for the high backbone entropy in interaction with protein antigens. Antibodies prefer charged and polar antigen residues and bridging water molecules. They also prefer positive net charge, presumably to promote interaction with negatively charged protein antigens, which are common in proteomes. Antibody-antigen interfaces have large percentages of Tyr, Ser, and Asp, but little Lys. Electrostatic and hydrophobic interactions in the Ag binding sites might be coupled with Fab domains through organized charge and residue distributions away from the binding interfaces. Here we describe some features of antibody-antigen interfaces and of Fab domains as compared with nonantibody protein-protein interactions. The distributions of interface residues in human and murine antibodies do not differ significantly. Overall, our results provide not only a local but also a global anatomy of antibody structures. Copyright © 2017 John Wiley & Sons, Ltd.

Antigenic Determinants of Alpha-Like Proteins of Streptococcus agalactiae

PubMed Central

Maeland, Johan A.; Bevanger, Lars; Lyng, Randi Valsoe

2004-01-01

The majority of group B streptococcus (GBS) isolates express one or more of a family of surface-anchored proteins that vary by strain and that form ladder-like patterns on Western blotting due to large repeat units. These proteins, which are important as GBS serotype markers and as inducers of protective antibodies, include the alpha C (Cα) and R4 proteins and the recently described alpha-like protein 2 (Alp2), encoded by alp2, and Alp3, encoded by alp3. In this study, we examined antigenic determinants possessed by Alp2 and Alp3 by testing of antibodies raised in rabbits, mainly by using enzyme-linked immunosorbent assays (ELISA) and an ELISA absorption test. The results showed that Alp2 and Alp3 shared an antigenic determinant, which may be a unique immunological marker of the Alp variants of GBS proteins. Alp2, in addition, possessed an antigenic determinant which showed specificity for Alp2 and a third determinant which showed serological cross-reactivity with Cα. Alp3, in addition to the determinant common to Alp2 and Alp3, harbored an antigenic site which also was present in the R4 protein, whereas no Alp3-specific antigenic site was detected. These ELISA-based results were confirmed by Western blotting and a fluorescent-antibody test. The results are consistent with highly complex antigenic structures of the alpha-like proteins in a fashion which is in agreement with the recently described structural mosaicism of the alp2 and alp3 genes. The results are expected to influence GBS serotyping, immunoprotection studies, and GBS vaccine developments. PMID:15539502

Sa-Lrp from Sulfolobus acidocaldarius is a versatile, glutamine-responsive, and architectural transcriptional regulator

PubMed Central

Vassart, Amelia; Wolferen, Marleen; Orell, Alvaro; Hong, Ye; Peeters, Eveline; Albers, Sonja-Verena; Charlier, Daniel

2013-01-01

Sa-Lrp is a member of the leucine-responsive regulatory protein (Lrp)-like family of transcriptional regulators in Sulfolobus acidocaldarius. Previously, we demonstrated the binding of Sa-Lrp to the control region of its own gene in vitro. However, the function and cofactor of Sa-Lrp remained an enigma. In this work, we demonstrate that glutamine is the cofactor of Sa-Lrp by inducing the formation of octamers and increasing the DNA-binding affinity and sequence specificity. In vitro protein-DNA interaction assays indicate that Sa-Lrp binds to promoter regions of genes with a variety of functions including ammonia assimilation, transcriptional control, and UV-induced pili synthesis. DNA binding occurs with a specific affinity for AT-rich binding sites, and the protein induces DNA bending and wrapping upon binding, indicating an architectural role of the regulator. Furthermore, by analyzing an Sa-lrp deletion mutant, we demonstrate that the protein affects transcription of some of the genes of which the promoter region is targeted and that it is an important determinant of the cellular aggregation phenotype. Taking all these results into account, we conclude that Sa-Lrp is a glutamine-responsive global transcriptional regulator with an additional architectural role. PMID:23255531

The antigenic surface of staphylococcal nuclease. II. Analysis of the N-1 epitope by site-directed mutagenesis.

PubMed

Smith, A M; Benjamin, D C

1991-02-15

Previous studies in our laboratory on the production and isolation of a panel of mAb to staphylococcal nuclease allowed us to define a series of eight overlapping epitopes. Using site-directed mutagenesis of the nuclease coding sequences we were able to map the nonoverlapping epitopes recognized by two members of this panel. In the study reported here, we report the generation and analysis of a number of single amino acid substitutions for seven surface residues predicted to lie within one of these two epitopes. Immunochemical analysis showed that one or more substitutions at each of these seven positions had a major effect on mAb binding, whereas other substitutions had none. Based on the nature of these substitutions and the chemical and physical properties of the variant molecules, we believe that any structural effects induced by these substitutions are local and do not result in long-range structural alterations that indirectly influence antibody reactivity. Therefore, we conclude that disruption of mAb binding can be directly attributed to changes in amino acid side chains and that not only are all seven of the residues studied part of the epitope but all seven make contact with the antibody combining site. These studies demonstrate the advantages of using site-directed mutagenesis to study antigen structure and emphasize the importance of constructing the examining multiple substitutions for any given amino acid.

G‐LoSA: An efficient computational tool for local structure‐centric biological studies and drug design

PubMed Central

2016-01-01

Abstract Molecular recognition by protein mostly occurs in a local region on the protein surface. Thus, an efficient computational method for accurate characterization of protein local structural conservation is necessary to better understand biology and drug design. We present a novel local structure alignment tool, G‐LoSA. G‐LoSA aligns protein local structures in a sequence order independent way and provides a GA‐score, a chemical feature‐based and size‐independent structure similarity score. Our benchmark validation shows the robust performance of G‐LoSA to the local structures of diverse sizes and characteristics, demonstrating its universal applicability to local structure‐centric comparative biology studies. In particular, G‐LoSA is highly effective in detecting conserved local regions on the entire surface of a given protein. In addition, the applications of G‐LoSA to identifying template ligands and predicting ligand and protein binding sites illustrate its strong potential for computer‐aided drug design. We hope that G‐LoSA can be a useful computational method for exploring interesting biological problems through large‐scale comparison of protein local structures and facilitating drug discovery research and development. G‐LoSA is freely available to academic users at http://im.compbio.ku.edu/GLoSA/. PMID:26813336

G-LoSA: An efficient computational tool for local structure-centric biological studies and drug design.

PubMed

Lee, Hui Sun; Im, Wonpil

2016-04-01

Molecular recognition by protein mostly occurs in a local region on the protein surface. Thus, an efficient computational method for accurate characterization of protein local structural conservation is necessary to better understand biology and drug design. We present a novel local structure alignment tool, G-LoSA. G-LoSA aligns protein local structures in a sequence order independent way and provides a GA-score, a chemical feature-based and size-independent structure similarity score. Our benchmark validation shows the robust performance of G-LoSA to the local structures of diverse sizes and characteristics, demonstrating its universal applicability to local structure-centric comparative biology studies. In particular, G-LoSA is highly effective in detecting conserved local regions on the entire surface of a given protein. In addition, the applications of G-LoSA to identifying template ligands and predicting ligand and protein binding sites illustrate its strong potential for computer-aided drug design. We hope that G-LoSA can be a useful computational method for exploring interesting biological problems through large-scale comparison of protein local structures and facilitating drug discovery research and development. G-LoSA is freely available to academic users at http://im.compbio.ku.edu/GLoSA/. © 2016 The Protein Society.

Goodbye warts, hello vitiligo: Candida antigen-induced depigmentation.

PubMed

Wilmer, Erin N; Burkhart, Craig N; Morrell, Dean S

2013-01-01

Depigmentation after the use of topical immune modulators is a rare but reported event. Herein we present what is to our knowledge the first case of vitiligo at a site of Candida antigen injection. © 2012 Wiley Periodicals, Inc.

Selection of antigenically advanced variants of seasonal influenza viruses

PubMed Central

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

Selection of antigenically advanced variants of seasonal influenza viruses.

PubMed

Li, Chengjun; Hatta, Masato; Burke, David F; Ping, Jihui; Zhang, Ying; 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; Klimov, Alexander I; 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

2016-05-23

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. We also 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.

Mapping replication dynamics in Trypanosoma brucei reveals a link with telomere transcription and antigenic variation

PubMed Central

Devlin, Rebecca; Marques, Catarina A; Paape, Daniel; Prorocic, Marko; Zurita-Leal, Andrea C; Campbell, Samantha J; Lapsley, Craig; Dickens, Nicholas; McCulloch, Richard

2016-01-01

Survival of Trypanosoma brucei depends upon switches in its protective Variant Surface Glycoprotein (VSG) coat by antigenic variation. VSG switching occurs by frequent homologous recombination, which is thought to require locus-specific initiation. Here, we show that a RecQ helicase, RECQ2, acts to repair DNA breaks, including in the telomeric site of VSG expression. Despite this, RECQ2 loss does not impair antigenic variation, but causes increased VSG switching by recombination, arguing against models for VSG switch initiation through direct generation of a DNA double strand break (DSB). Indeed, we show DSBs inefficiently direct recombination in the VSG expression site. By mapping genome replication dynamics, we reveal that the transcribed VSG expression site is the only telomeric site that is early replicating – a differential timing only seen in mammal-infective parasites. Specific association between VSG transcription and replication timing reveals a model for antigenic variation based on replication-derived DNA fragility. DOI: http://dx.doi.org/10.7554/eLife.12765.001 PMID:27228154

Immunogenicity in humans of a recombinant bacterial antigen delivered in a transgenic potato.

PubMed

Tacket, C O; Mason, H S; Losonsky, G; Clements, J D; Levine, M M; Arntzen, C J

1998-05-01

Compared with vaccine delivery by injection, oral vaccines offer the hope of more convenient immunization strategies and a more practical means of implementing universal vaccination programs throughout the world. Oral vaccines act by stimulating the immune system at effector sites (lymphoid tissue) located in the gut. Genetic engineering has been used with variable success to design living and non-living systems as a means to deliver antigens to these sites and to stimulate a desired immune response. More recently, plant biotechnology techniques have been used to create plants which contain a gene derived from a human pathogen; the resultant plant tissues will accumulate an antigenic protein encoded by the foreign DNA. In pre-clinical trials, we found that antigenic proteins produced in transgenic plants retained immunogenic properties when purified; if injected into mice the antigen caused production of protein-specific antibodies. Moreover, in some experiments, if the plant tissues were simply fed to mice, a mucosal immune response occurred. The present study was conducted as a proof of principle to determine if humans would also develop a serum and/or mucosal immune response to an antigen delivered in an uncooked foodstuff.

Immunity to tumour antigens.

PubMed

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

2005-01-01

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

Antigen-dependent fluorescence response of anti-c-Myc Quenchbody studied by molecular dynamics simulations

NASA Astrophysics Data System (ADS)

Mori, Yoshiharu; Okumura, Hisashi; Watanabe, Takayoshi; Hohsaka, Takahiro

2018-04-01

We performed metadynamics molecular dynamics simulations to reveal mechanism of antigen-dependent fluorescence response observed for site-specifically fluorescent-labeled single-chain antibody against c-Myc peptide antigen. We found that VH and VL bind with each other only when the antigen exists and that the fluorophore labeled at the N-terminus of VH interacts with Trp103 most stably. These results support the mechanism proposed from previous experiments: In the absence of antigen, Trp residues are partially exposed at the interface of VH and quench the fluorophore. In the presence of antigen, the Trp residues are buried between VH and VL , and the quenching is eliminated.

Prediction, dynamics, and visualization of antigenic phenotypes of seasonal influenza viruses

PubMed Central

Neher, Richard A.; Bedford, Trevor; Daniels, Rodney S.; Shraiman, Boris I.

2016-01-01

Human seasonal influenza viruses evolve rapidly, enabling the virus population to evade immunity and reinfect previously infected individuals. Antigenic properties are largely determined by the surface glycoprotein hemagglutinin (HA), and amino acid substitutions at exposed epitope sites in HA mediate loss of recognition by antibodies. Here, we show that antigenic differences measured through serological assay data are well described by a sum of antigenic changes along the path connecting viruses in a phylogenetic tree. This mapping onto the tree allows prediction of antigenicity from HA sequence data alone. The mapping can further be used to make predictions about the makeup of the future A(H3N2) seasonal influenza virus population, and we compare predictions between models with serological and sequence data. To make timely model output readily available, we developed a web browser-based application that visualizes antigenic data on a continuously updated phylogeny. PMID:26951657

Multiple signal amplification strategies for ultrasensitive label-free electrochemical immunoassay for carbohydrate antigen 24-2 based on redox hydrogel.

PubMed

Tang, Zhongxue; Fu, Yuanyuan; Ma, Zhanfang

2017-05-15

In this work, multiple signal amplification strategies for ultrasensitive label-free electrochemical immunoassay for carbohydrate antigen 24-2 (CA242) were developed using redox sodium alginate-Pb 2+ -graphene oxide (SA-Pb 2+ -GO) hydrogel. The SA-Pb 2+ -GO hydrogel was synthesised by simply mixing SA, GO, and Pb 2+ and then implemented as a novel redox species with a strong current signal at -0.46V (vs. Ag/AgCl). After the three-dimensional and porous SA-Pb 2+ -GO hydrogel was in situ generated on a glassy carbon electrode (GCE), chitosan was adsorbed on the obtained electrode to further enrich Pb 2+ . When chitosan-Pb 2+ /SA-Pb 2+ -GO/GCE was incubated with anti-CA242 using glutaraldehyde and blocked by bovine serum albumin, the immunoassay platform for CA242 was obtained. Owing to the addition of GO, the obtained conductive SA-GO/GCE was beneficial for signal amplification. After incubating SA-GO/GCE with excessive amounts of Pb 2+ , the resistance of SA-Pb 2+ -GO/GCE further decreased and a strong redox signal was obtained. The chitosan fixed by electrostatic adsorption resulted in further adsorption of Pb 2+ , behaving as further amplifying the signal and improving conductivity. In this case, multiple signal amplification strategies were involved in the proposed immunosensor for the ultrasensitive detection of CA242. Under the optimal conditions, the proposed immunosensor exhibited a wide linear range from 0.005UmL -1 to 500UmL -1 with an ultralow detection limit of 0.067mUmL -1 . In comparison to previous works, the sensitivity of this method was 32.98μA (log 10 C CA242 ) -1 , which was a five-fold increase from the previous works. Copyright © 2016 Elsevier B.V. All rights reserved.

The Role of FcRn in Antigen Presentation

PubMed Central

Baker, Kristi; Rath, Timo; Pyzik, Michal; Blumberg, Richard S.

2014-01-01

Immunoglobulins are unique molecules capable of simultaneously recognizing a diverse array of antigens and themselves being recognized by a broad array of receptors. The abundance specifically of the IgG subclass and the variety of signaling receptors to which it binds render this an important immunomodulatory molecule. In addition to the classical Fcγ receptors that bind IgG at the cell surface, the neonatal Fc receptor (FcRn) is a lifelong resident of the endolysosomal system of most hematopoietic cells where it determines the intracellular fate of both IgG and IgG-containing immune complexes (IgG IC). Cross-linking of FcRn by multivalent IgG IC within antigen presenting cells such as dendritic cells initiates specific mechanisms that result in trafficking of the antigen-bearing IgG IC into compartments from which the antigen can successfully be processed into peptide epitopes compatible with loading onto both major histocompatibility complex class I and II molecules. In turn, this enables the synchronous activation of both CD4+ and CD8+ T cell responses against the cognate antigen, thereby bridging the gap between the humoral and cellular branches of the adaptive immune response. Critically, FcRn-driven T cell priming is efficient at very low doses of antigen due to the exquisite sensitivity of the IgG-mediated antigen delivery system through which it operates. FcRn-mediated antigen presentation has important consequences in tissue compartments replete with IgG and serves not only to determine homeostatic immune activation at a variety of sites but also to induce inflammatory responses upon exposure to antigens perceived as foreign. Therapeutically targeting the pathway by which FcRn enables T cell activation in response to IgG IC is thus a highly attractive prospect not only for the treatment of diseases that are driven by immune complexes but also for manipulating local immune responses against defined antigens such as those present during infections and

Antigen Binding and Site-Directed Labeling of Biosilica-Immobilized Fusion Proteins Expressed in Diatoms.

PubMed

Ford, Nicole R; Hecht, Karen A; Hu, DeHong; Orr, Galya; Xiong, Yijia; Squier, Thomas C; Rorrer, Gregory L; Roesijadi, Guritno

2016-03-18

The diatom Thalassiosira pseudonana was genetically modified to express biosilica-targeted fusion proteins comprising either enhanced green fluorescent protein (EGFP) or single chain antibodies engineered with a tetracysteine tagging sequence. Of interest were the site-specific binding of (1) the fluorescent biarsenical probe AsCy3 and AsCy3e to the tetracysteine tagged fusion proteins and (2) high and low molecular mass antigens, the Bacillus anthracis surface layer protein EA1 or small molecule explosive trinitrotoluene (TNT), to biosilica-immobilized single chain antibodies. Analysis of biarsenical probe binding using fluorescence and structured illumination microscopy indicated differential colocalization with EGFP in nascent and mature biosilica, supporting the use of either EGFP or bound AsCy3 and AsCy3e in studying biosilica maturation. Large increases in the lifetime of a fluorescent analogue of TNT upon binding single chain antibodies provided a robust signal capable of discriminating binding to immobilized antibodies in the transformed frustule from nonspecific binding to the biosilica matrix. In conclusion, our results demonstrate an ability to engineer diatoms to create antibody-functionalized mesoporous silica able to selectively bind chemical and biological agents for the development of sensing platforms.

Site-specific fab fragment biotinylation at the conserved nucleotide binding site for enhanced Ebola detection.

PubMed

Mustafaoglu, Nur; Alves, Nathan J; Bilgicer, Basar

2015-07-01

The nucleotide binding site (NBS) is a highly conserved region between the variable light and heavy chains at the Fab domains of all antibodies, and a small molecule that we identified, indole-3-butyric acid (IBA), binds specifically to this site. Fab fragment, with its small size and simple production methods compared to intact antibody, is good candidate for use in miniaturized diagnostic devices and targeted therapeutic applications. However, commonly used modification techniques are not well suited for Fab fragments as they are often more delicate than intact antibodies. Fab fragments are of particular interest for sensor surface functionalization but immobilization results in damage to the antigen binding site and greatly reduced activity due to their truncated size that allows only a small area that can bind to surfaces without impeding antigen binding. In this study, we describe an NBS-UV photocrosslinking functionalization method (UV-NBS(Biotin) in which a Fab fragment is site-specifically biotinylated with an IBA-EG11-Biotin linker via UV energy exposure (1 J/cm(2)) without affecting its antigen binding activity. This study demonstrates successful immobilization of biotinylated Ebola detecting Fab fragment (KZ52 Fab fragment) via the UV-NBS(Biotin) method yielding 1031-fold and 2-fold better antigen detection sensitivity compared to commonly used immobilization methods: direct physical adsorption and NHS-Biotin functionalization, respectively. Utilization of the UV-NBS(Biotin) method for site-specific conjugation to Fab fragment represents a proof of concept use of Fab fragment for various diagnostic and therapeutic applications with numerous fluorescent probes, affinity molecules and peptides. © 2015 Wiley Periodicals, Inc.

Antigenic regions within the hepatitis C virus envelope 1 and non-structural proteins: identification of an IgG3-restricted recognition site with the envelope 1 protein.

PubMed Central

Sällberg, M; Rudén, U; Wahren, B; Magnius, L O

1993-01-01

Antibody binding to antigenic regions of hepatitis C virus (HCV) envelope 1 (E1; residues 183-380, E2/non-structural (NS) 1 (residues 380-437), NS1 (residues 643-690), and NS4 (1684-1751) proteins were assayed for 50 sera with antibodies to HCV (anti-HCV) and for 46 sera without anti-HCV. Thirty-four peptides, 18 residues long with an eight-amino acid overlap within each HCV region, were synthesized and tested with all 96 sera. Within the E region 183-380, the major binding site was located to residues 203-220, and was recognized by eight sera. Within the E2/NS1 region 380-437, the peptide covering residues 410-427 was recognized by two sera, and within the NS1 region 643-690, peptides covering residues 663-690 were recognized by four sera. Within the NS4 region 1684-1751, 27 sera were reactive to one or more of the NS4 peptides, and 21 out of these were reactive with peptide 1694-1711. One part of the major binding site could be located to residues 1701-1704, with the sequence Leu-Tyr-Arg-Glu. The IgG1, IgG3 and IgG4 subclasses were reactive with the five antigenic regions of HCV core, residues 1-18, 11-28, 21-38, 51-68 and 101-118. Reactivity to the major envelope site consisted almost exclusively of IgG3, and reactivity to the major site of NS4 consisted only of IgG1. Thus, a non-restricted IgG response to linear HCV-encoded binding sites was found to the core protein, whereas IgG subclass-restricted linear binding sites were found within the E1 protein, and within the NS4 protein. PMID:7680297

Ovarian tumor antigens.

PubMed

Bhattacharya, M; Barlow, J J

1978-09-01

Evidence has been reported for at least two common tumor-associated antigens, or antigenic determinants, in human cystadenocarcinomas of the ovary that are apparently absent in tissues of normal reproductive organs. These antigenic determinants are immunologically distinct from carcinoembryonic antigen, alpha-fetoprotein, ferritins and histocompatibility antigens. One of these two ovarian cystadenocarcinoma-associated antigens (OCAA) is not detectable in any ovarian carcinomas except serous or mucinous types, other gynecologic or nongynecologic malignancies thus far tested, while the second antigen is present in about 90% of all gynecologic tumors and occasionally in breast and colon tumors. OCAA has been purified and partially characterized. It is a high molecular weight glycoprotein which carries the unique ovarian tumor-specific antigenic determinant along with some normal cross-reacting determinants. High levels of this glycoprotein antigen have been detected in the sera of ovarian cancer patients with advanced disease by the radioimmunoassay inhibition technique. The serial determination of circulating OCAA appeared to correlate with tumor volume as well as the clinical status of the patients.

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens

PubMed Central

Kleiveland, Charlotte R.; Minic, Rajna; Moen, Lars F.; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Eijsink, Vincent G. H.

2016-01-01

ABSTRACT Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum. The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo. The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. IMPORTANCE This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and

Immunogenic Properties of Lactobacillus plantarum Producing Surface-Displayed Mycobacterium tuberculosis Antigens.

PubMed

Kuczkowska, Katarzyna; Kleiveland, Charlotte R; Minic, Rajna; Moen, Lars F; Øverland, Lise; Tjåland, Rannei; Carlsen, Harald; Lea, Tor; Mathiesen, Geir; Eijsink, Vincent G H

2017-01-15

Tuberculosis (TB) remains among the most deadly diseases in the world. The only available vaccine against tuberculosis is the bacille Calmette-Guérin (BCG) vaccine, which does not ensure full protection in adults. There is a global urgency for the development of an effective vaccine for preventing disease transmission, and it requires novel approaches. We are exploring the use of lactic acid bacteria (LAB) as a vector for antigen delivery to mucosal sites. Here, we demonstrate the successful expression and surface display of a Mycobacterium tuberculosis fusion antigen (comprising Ag85B and ESAT-6, referred to as AgE6) on Lactobacillus plantarum The AgE6 fusion antigen was targeted to the bacterial surface using two different anchors, a lipoprotein anchor directing the protein to the cell membrane and a covalent cell wall anchor. AgE6-producing L. plantarum strains using each of the two anchors induced antigen-specific proliferative responses in lymphocytes purified from TB-positive donors. Similarly, both strains induced immune responses in mice after nasal or oral immunization. The impact of the anchoring strategies was reflected in dissimilarities in the immune responses generated by the two L. plantarum strains in vivo The present study comprises an initial step toward the development of L. plantarum as a vector for M. tuberculosis antigen delivery. This work presents the development of Lactobacillus plantarum as a candidate mucosal vaccine against tuberculosis. Tuberculosis remains one of the top infectious diseases worldwide, and the only available vaccine, bacille Calmette-Guérin (BCG), fails to protect adults and adolescents. Direct antigen delivery to mucosal sites is a promising strategy in tuberculosis vaccine development, and lactic acid bacteria potentially provide easy, safe, and low-cost delivery vehicles for mucosal immunization. We have engineered L. plantarum strains to produce a Mycobacterium tuberculosis fusion antigen and to anchor this

Empirical Site Amplification Factors Incorporating Soil Nonlinearity in Taiwan

NASA Astrophysics Data System (ADS)

Kuo, C. H.; Chung, C. H.; Che-Min, L.; Huang, J. Y.; Wen, K. L.

2017-12-01

Characteristics of site amplifications caused by both crustal and subduction earthquakes are important in Taiwan. For example, seismic waves were amplified and led to significant building damages in the Taipei Basin by the 1986 Hualien offshore (subduction interface) and the 1999 Chi-Chi earthquakes (crustal), for which the epicentral distances were about 100 km. To understand local site amplifications in Taiwan, empirical site amplification factors for horizontal ground motions are studied using recently constructed strong motion and site databases for the free-field TSMIP stations in Taiwan. Records of large magnitude earthquakes of ML larger than six from 1994 to 2014 were selected for this study. Site amplification factors at site conditions with Vs30 of 120 m/s to 1500 m/s and base accelerations up to 0.7g were inferred from intensity ratios of station pairs within specific distances. The reference site condition is assumed as Vs30 of 760 m/s (B/C boundary). Preliminary results indicate: 1. Soil nonlinearity is more obviously at short periods (PGA, Sa0.3) than long periods (PGV, Sa1.0). 2. Soil nonlinearity is significant for stations belong to site classes of B, C, D, and E in Taiwan. 3. Effect of station-pair distance is seen at short periods (PGA and Sa0.3). 4. No significant different is found in site amplifications of crustal and subduction earthquakes. The result could be a reference for the Fa and Fv in Taiwan's building code.

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

PubMed

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

2000-03-01

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

Determination of antigenicity-altering patches on the major surface protein of human influenza A/H3N2 viruses

PubMed Central

Kratsch, Christina; Klingen, Thorsten R.; Mümken, Linda; Steinbrück, Lars; McHardy, Alice C.

2016-01-01

Human influenza viruses are rapidly evolving RNA viruses that cause short-term respiratory infections with substantial morbidity and mortality in annual epidemics. Uncovering the general principles of viral coevolution with human hosts is important for pathogen surveillance and vaccine design. Protein regions are an appropriate model for the interactions between two macromolecules, but the currently used epitope definition for the major antigen of influenza viruses, namely hemagglutinin, is very broad. Here, we combined genetic, evolutionary, antigenic, and structural information to determine the most relevant regions of the hemagglutinin of human influenza A/H3N2 viruses for interaction with human immunoglobulins. We estimated the antigenic weights of amino acid changes at individual sites from hemagglutination inhibition data using antigenic tree inference followed by spatial clustering of antigenicity-altering protein sites on the protein structure. This approach determined six relevant areas (patches) for antigenic variation that had a key role in the past antigenic evolution of the viruses. Previous transitions between successive predominating antigenic types of H3N2 viruses always included amino acid changes in either the first or second antigenic patch. Interestingly, there was only partial overlap between the antigenic patches and the patches under strong positive selection. Therefore, besides alterations of antigenicity, other interactions with the host may shape the evolution of human influenza A/H3N2 viruses. PMID:27774294

A Lipid Based Antigen Delivery System Efficiently Facilitates MHC Class-I Antigen Presentation in Dendritic Cells to Stimulate CD8+ T Cells

NASA Astrophysics Data System (ADS)

Maji, Mithun; Mazumder, Saumyabrata; Bhattacharya, Souparno; Choudhury, Somsubhra Thakur; Sabur, Abdus; Shadab, Md.; Bhattacharya, Pradyot; Ali, Nahid

2016-06-01

The most effective strategy for protection against intracellular infections such as Leishmania is vaccination with live parasites. Use of recombinant proteins avoids the risks associated with live vaccines. However, due to low immunogenicity, they fail to trigger T cell responses particularly of CD8+ cells requisite for persistent immunity. Previously we showed the importance of protein entrapment in cationic liposomes and MPL as adjuvant for elicitation of CD4+ and CD8+ T cell responses for long-term protection. In this study we investigated the role of cationic liposomes on maturation and antigen presentation capacity of dendritic cells (DCs). We observed that cationic liposomes were taken up very efficiently by DCs and transported to different cellular sites. DCs activated with liposomal rgp63 led to efficient presentation of antigen to specific CD4+ and CD8+ T cells. Furthermore, lymphoid CD8+ T cells from liposomal rgp63 immunized mice demonstrated better proliferative ability when co-cultured ex vivo with stimulated DCs. Addition of MPL to vaccine enhanced the antigen presentation by DCs and induced more efficient antigen specific CD8+ T cell responses when compared to free and liposomal antigen. These liposomal formulations presented to CD8+ T cells through TAP-dependent MHC-I pathway offer new possibilities for a safe subunit vaccine.

Identification of Caspase Cleavage Sites in KSHV Latency-Associated Nuclear Antigen and Their Effects on Caspase-Related Host Defense Responses.

PubMed

Davis, David A; Naiman, Nicole E; Wang, Victoria; Shrestha, Prabha; Haque, Muzammel; Hu, Duosha; Anagho, Holda A; Carey, Robert F; Davidoff, Katharine S; Yarchoan, Robert

2015-07-01

Kaposi's sarcoma-associated herpesvirus (KSHV), also known as human herpesvirus-8, is the causative agent of three hyperproliferative disorders: Kaposi's sarcoma, primary effusion lymphoma (PEL) and multicentric Castleman's disease. During viral latency a small subset of viral genes are produced, including KSHV latency-associated nuclear antigen (LANA), which help the virus thwart cellular defense responses. We found that exposure of KSHV-infected cells to oxidative stress, or other inducers of apoptosis and caspase activation, led to processing of LANA and that this processing could be inhibited with the pan-caspase inhibitor Z-VAD-FMK. Using sequence, peptide, and mutational analysis, two caspase cleavage sites within LANA were identified: a site for caspase-3 type caspases at the N-terminus and a site for caspase-1 and-3 type caspases at the C-terminus. Using LANA expression plasmids, we demonstrated that mutation of these cleavage sites prevents caspase-1 and caspase-3 processing of LANA. This indicates that these are the principal sites that are susceptible to caspase cleavage. Using peptides spanning the identified LANA cleavage sites, we show that caspase activity can be inhibited in vitro and that a cell-permeable peptide spanning the C-terminal cleavage site could inhibit cleavage of poly (ADP-ribose) polymerase and increase viability in cells undergoing etoposide-induced apoptosis. The C-terminal peptide of LANA also inhibited interleukin-1 beta (IL-1β) production from lipopolysaccharide-treated THP-1 cells by more than 50%. Furthermore, mutation of the two cleavage sites in LANA led to a significant increase in IL-1β production in transfected THP-1 cells; this provides evidence that these sites function to blunt the inflammasome, which is known to be activated in latently infected PEL cells. These results suggest that specific caspase cleavage sites in KSHV LANA function to blunt apoptosis as well as interfere with the caspase-1-mediated inflammasome

Glioma antigen.

PubMed

Toda, Masahiro

2012-01-01

Because several antigenic peptides of human tumors that are recognized by T-lymphocytes have been identified, immune responses against cancer can now be artificially manipulated. Furthermore, since T-lymphocytes have been found to play an important role in the rejection of tumors by the host and also to have antigen-specific proliferative potentials and memory mechanisms, T-lymphocytes are thought to play a central role in cancer vaccination. Although multidisciplinary therapies have been attempted for the treatment of gliomas, the results remain unsatisfactory. For the development of new therapies against gliomas, it is required to identify tumor antigens as targets for specific immunotherapy. In this chapter, recent progress in research on glioma antigens is described.

SaLEM (v1.0) - the Soil and Landscape Evolution Model (SaLEM) for simulation of regolith depth in periglacial environments

NASA Astrophysics Data System (ADS)

Bock, Michael; Conrad, Olaf; Günther, Andreas; Gehrt, Ernst; Baritz, Rainer; Böhner, Jürgen

2018-04-01

We propose the implementation of the Soil and Landscape Evolution Model (SaLEM) for the spatiotemporal investigation of soil parent material evolution following a lithologically differentiated approach. Relevant parts of the established Geomorphic/Orogenic Landscape Evolution Model (GOLEM) have been adapted for an operational Geographical Information System (GIS) tool within the open-source software framework System for Automated Geoscientific Analyses (SAGA), thus taking advantage of SAGA's capabilities for geomorphometric analyses. The model is driven by palaeoclimatic data (temperature, precipitation) representative of periglacial areas in northern Germany over the last 50 000 years. The initial conditions have been determined for a test site by a digital terrain model and a geological model. Weathering, erosion and transport functions are calibrated using extrinsic (climatic) and intrinsic (lithologic) parameter data. First results indicate that our differentiated SaLEM approach shows some evidence for the spatiotemporal prediction of important soil parental material properties (particularly its depth). Future research will focus on the validation of the results against field data, and the influence of discrete events (mass movements, floods) on soil parent material formation has to be evaluated.

Targeted delivery of antigen processing inhibitors to antigen presenting cells via mannose receptors.

PubMed

Raiber, Eun-Ang; Tulone, Calogero; Zhang, Yanjing; Martinez-Pomares, Luisa; Steed, Emily; Sponaas, Anna M; Langhorne, Jean; Noursadeghi, Mahdad; Chain, Benjamin M; Tabor, Alethea B

2010-05-21

Improved chemical inhibitors are required to dissect the role of specific antigen processing enzymes and to complement genetic models. In this study we explore the in vitro and in vivo properties of a novel class of targeted inhibitor of aspartic proteinases, in which pepstatin is coupled to mannosylated albumin (MPC6), creating an inhibitor with improved solubility and the potential for selective cell tropism. Using these compounds, we have demonstrated that MPC6 is taken up via mannose receptor facilitated endocytosis, leading to a slow but continuous accumulation of inhibitor within large endocytic vesicles within dendritic cells and a parallel inhibition of intracellular aspartic proteinase activity. Inhibition of intracellular proteinase activity is associated with reduction in antigen processing activity, but this is epitope-specific, preferentially inhibiting processing of T cell epitopes buried within compact proteinase-resistant protein domains. Unexpectedly, we have also demonstrated, using quenched fluorescent substrates, that little or no cleavage of the disulfide linker takes place within dendritic cells. This does not appear to affect the activity of MPC6 as an inhibitor of cathepsins D and E in vitro and in vivo. Finally, we have shown that MPC6 selectively targets dendritic cells and macrophages in spleen in vivo. Preliminary results suggest that access to nonlymphoid tissues is very limited in the steady state but is strongly enhanced at local sites of inflammation. The strategy adopted for MPC6 synthesis may therefore represent a more general way to deliver chemical inhibitors to cells of the innate immune system, especially at sites of inflammation.

THE SITE DEMONSTRATION OF CHEMFIX SOLIDIFICATION/ STABILIZATION PROCESS AT THE PORTABLE EQUIPMENT SALVAGE COMPANY SITE

EPA Science Inventory

A demonstration of the GHEMFIX solidification/stabilization process was conducted under the United States Environmental Protection Agency`s (EPA) Superfund Innovative Technology Evaluation (SITE) program. The demonstration was conducted in March 1989, at the Portable Equipment Sa...

A Comprehensive Study of Neutralizing Antigenic Sites on the Hepatitis E Virus (HEV) Capsid by Constructing, Clustering, and Characterizing a Tool Box*

PubMed Central

Zhao, Min; Li, Xiao-Jing; Tang, Zi-Min; Yang, Fan; Wang, Si-Ling; Cai, Wei; Zhang, Ke; Xia, Ning-Shao; Zheng, Zi-Zheng

2015-01-01

The hepatitis E virus (HEV) ORF2 encodes a single structural capsid protein. The E2s domain (amino acids 459–606) of the capsid protein has been identified as the major immune target. All identified neutralizing epitopes are located on this domain; however, a comprehensive characterization of antigenic sites on the domain is lacking due to its high degree of conformation dependence. Here, we used the statistical software SPSS to analyze cELISA (competitive ELISA) data to classify monoclonal antibodies (mAbs), which recognized conformational epitopes on E2s domain. Using this novel analysis method, we identified various conformational mAbs that recognized the E2s domain. These mAbs were distributed into 6 independent groups, suggesting the presence of at least 6 epitopes. Twelve representative mAbs covering the six groups were selected as a tool box to further map functional antigenic sites on the E2s domain. By combining functional and location information of the 12 representative mAbs, this study provided a complete picture of potential neutralizing epitope regions and immune-dominant determinants on E2s domain. One epitope region is located on top of the E2s domain close to the monomer interface; the other is located on the monomer side of the E2s dimer around the groove zone. Besides, two non-neutralizing epitopes were also identified on E2s domain that did not stimulate neutralizing antibodies. Our results help further the understanding of protective mechanisms induced by the HEV vaccine. Furthermore, the tool box with 12 representative mAbs will be useful for studying the HEV infection process. PMID:26085097

A 1H NMR method for the analysis of antigen-antibody interactions: binding of a peptide fragment of lysozyme to anti-lysozyme monoclonal antibody.

PubMed

Ito, W; Nishimura, M; Sakato, N; Fujio, H; Arata, Y

1987-09-01

A proton nuclear magnetic resonance (NMR) study is reported of the molecular structural basis of antigen-antibody interactions. An immunologically reactive proteolytic fragment corresponding to one of the antigenic regions on hen egg-white lysozyme (HEL) was used in combination with a monoclonal antibody that recognizes this site. Using spin diffusion, we prepared an antibody in which the magnetization of the antigen binding site was saturated by non-specific nuclear Overhauser effect. Under these conditions the effect of the saturation of the antibody was observed to spread over the peptide fragment through the antigen binding site. On the basis of the results obtained for the intermolecular nuclear Overhauser effect, we discuss how the peptide fragment interacts with the antibody. The side chains of aromatic residues, Trp, Tyr, and His, and of ionic residues, especially Arg, Lys, and Glu, are suggested to be important in the antigen-antibody interaction.

An outer membrane protein (porin) as an eliciting antigen for delayed-type hypersensitivity in murine salmonellosis.

PubMed Central

Udhayakumar, V; Muthukkaruppan, V R

1987-01-01

The porin, an outer membrane protein of Salmonella typhimurium, was found to be a suitable antigen for eliciting delayed-type hypersensitivity in mouse salmonellosis. Histological examination of the reaction site revealed that the porin was superior to other antigenic preparations in eliciting a typical delayed-type hypersensitivity reaction consisting of mononuclear cell infiltration without polymorphonuclear cell contamination. This study indicates the importance of using a suitable protein antigen from S. typhi for human application. Images PMID:3028963

Rods and cones contain antigenically distinctive S-antigens.

PubMed

Nork, T M; Mangini, N J; Millecchia, L L

1993-09-01

S-antigen (48 kDa protein or arrestin) is known to be present in rod photoreceptors. Its localization in cones is less clear with several conflicting reports among various species examined. This study employed three different anti-S-antigen antibodies (a48K, a polyclonal antiserum and two monoclonal antibodies, MAb A9-C6 and MAb 5c6.47) and examined their localization in rods and cones of human and cat retinas. To identify the respective cone types, an enzyme histochemical technique for carbonic anhydrase (CA) was employed to distinguish blue cones (CA-negative) from red or green cones (CA-positive). S-antigen localization was then examined by immunocytochemical staining of adjacent sections. In human retinas, a similar labeling pattern was seen with both a48K and MAb A9-C6, i.e., the rods and blue-sensitive cones were strongly positive, whereas the red- or green-sensitive cones showed little immunoreactivity. All human photoreceptors showed reactivity to MAb 5c6.47. In the cat retina, only CA-positive cones could be found. As in the human retina, both rods and cones of the cat were positive for MAb 5c6.47. A difference from the labeling pattern in human retina was noted for the other S-antigen antibodies; a48K labeled rods and all of the cones, whereas MAb A9-C6 reacted strongly with the rods but showed no cone staining. These results suggest that both rods and cones contain S-antigen but that they are antigenically distinctive.

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

PubMed

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.

Direct measurement of IgM-Antigen interaction energy on individual red blood cells.

PubMed

Yeow, Natasha; Tabor, Rico F; Garnier, Gil

2017-07-01

Most blood grouping tests rely on the principle of red blood cells (RBCs) agglutination. Agglutination is triggered by the binding of specific blood grouping antibodies to the corresponding RBC surface antigen on multiple cells. The interaction energies between blood grouping antibodies and antigens have been poorly defined in immunohaematology. Here for the first time, we functionalized atomic force microscope (AFM) cantilevers with the IgM form of blood grouping antibodies to probe populations of individual RBCs of different groups under physiological conditions. The force-mapping mode of AFM allowed us to measure specific antibody - antigen interactions, and simultaneously localize and quantify antigen sites on the scanned cell surface. This study provides a new insight of the interactions between IgM antibodies and its corresponding antigen. The technique and information can be translated to develop better blood typing diagnostics and optimize target-specific drug delivery for medical applications. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

Ligand Binding Site Detection by Local Structure Alignment and Its Performance Complementarity

PubMed Central

Lee, Hui Sun; Im, Wonpil

2013-01-01

Accurate determination of potential ligand binding sites (BS) is a key step for protein function characterization and structure-based drug design. Despite promising results of template-based BS prediction methods using global structure alignment (GSA), there is a room to improve the performance by properly incorporating local structure alignment (LSA) because BS are local structures and often similar for proteins with dissimilar global folds. We present a template-based ligand BS prediction method using G-LoSA, our LSA tool. A large benchmark set validation shows that G-LoSA predicts drug-like ligands’ positions in single-chain protein targets more precisely than TM-align, a GSA-based method, while the overall success rate of TM-align is better. G-LoSA is particularly efficient for accurate detection of local structures conserved across proteins with diverse global topologies. Recognizing the performance complementarity of G-LoSA to TM-align and a non-template geometry-based method, fpocket, a robust consensus scoring method, CMCS-BSP (Complementary Methods and Consensus Scoring for ligand Binding Site Prediction), is developed and shows improvement on prediction accuracy. The G-LoSA source code is freely available at http://im.bioinformatics.ku.edu/GLoSA. PMID:23957286

Monoclonal antibodies against simian virus 40 T antigens: evidence for distinct sublcasses of large T antigen and for similarities among nonviral T antigens.

PubMed Central

Gurney, E G; Harrison, R O; Fenno, J

1980-01-01

We have isolated three clones of hybrid cells which synthesize antibodies specific for determinants on simian virus 40 (SV40) T antigens. Mouse myeloma NS1 cells were fused with spleen cells from mice that had been immunized with SV40-transformed mouse cells. Hybrid cells were selected in HAT medium and cloned in soft agar. We used an enzyme-linked immunosorbent assay for detection and quantification of mouse antibodies against SV40 T antigens. Monoclonal antibodies from 3 of the 24 clones that scored as positive in the enzyme-linked immunosorbent assay were verified by immunoprecipitation to be specific for SV40 T antigens. Two clones (7 and 412) produced antibodies that recognized denaturation-sensitive antigenic determinants unique to large T antigen. Antibodies from clone 7 appeared to have a low affinity for large T antigen. Antibodies from clone 412 had a higher affinity for large T antigen but did not recognize a subclass of large T antigen that was recognized by tumor serum. Antibodies of the third clone, clone 122, recognized a denaturation-stable antigenic determinant of the 53,000-dalton mouse nonviral T antigen in SV40-transformed cells. Antibodies from clone 122 also recognized similar (51,000- to 56,000-dalton) nonviral T antigens in SV40-transormed or lytically infected cells from five mammalian species and in four uninfected mouse lines. From these observations, we have concluded that (i) the 94,000-dalton SV40 large T antigen may exist as immunologically distinguishable subclasses, and (ii) the nonviral T antigens of five mammalian species share at least one antigenic determinant. Images PMID:6155477

Antigen-antibody interaction. The immunodominant region of EDP208 pili.

PubMed

Worobec, E A; Paranchych, W; Parker, J M; Taneja, A K; Hodges, R S

1985-01-25

The EDP208 pilus contains a major antigenic determinant in the N-terminal dodecapeptide, as shown by E. A. Worobec, A. K. Taneja, R. S. Hodges, and W. Paranchych ((1983) J. Bacteriol. 153, 955-961). This peptide was chemically synthesized, coupled to bovine serum albumin with N-hydroxysuccinimidyl p-azido-benzoate, and used in immunoblot and enzyme-linked immunosorbent assays to show it was capable of reacting with anti-EDP208 pilus antibodies. Antibodies raised against the synthetic peptide conjugate were also capable of reacting with whole pili in these assays. To further examine the specific residues responsible for the antigenicity of this site, several peptide analogs were chemically synthesized. The relative affinity of these peptides for anti-EDP208 pilus antibodies was determined by a competitive enzyme-linked immunosorbent assay using the Fab fragment of anti-EDP208 pilus immunoglobulin G. From these results we established that the antigenic region of this peptide was the N-terminal pentapeptide, N-acetyl-Thr-Asp-Leu-Leu-Ala, and the key residues responsible for the antibody-antigen interaction are the N-acetyl-Thr1, Leu3, and Leu4. Hydrophobic interactions involving the methyl of the acetyl group and the leucine side chains make the largest contributions to the antigen-antibody interaction, while a lesser contribution is made by the Thr1 hydroxyl. The side chains of Asp2 and Ala5 contribute only weakly to the stabilization of the antigen-antibody complex.

Neutrophils and monocytes transport tumor cell antigens from the peritoneal cavity to secondary lymphoid tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Terasawa, Masao; Nagata, Kisaburo; Kobayashi, Yoshiro

2008-12-12

Antigen-transporting cells take up pathogens, and then migrate from sites of inflammation to secondary lymphoid tissues to induce an immune response. Among antigen-transporting cells, dendritic cells (DCs) are believed to be the most potent and professional antigen-presenting cells that can stimulate naive T cells. However, the cells that transport antigens, tumor cell antigens in particular, have not been clearly identified. In this study we have analyzed what types of cells transport tumor cell antigens to secondary lymphoid tissues. We show that neutrophils, monocytes and macrophages but not DCs engulf X-irradiated P388 leukemic cells after their injection into the peritoneal cavity,more » and that neutrophils and monocytes but not macrophages migrate to the parathymic lymph nodes (pLN), the blood, and then the spleen. The monocytes in the pLN comprise Gr-1{sup -} and Gr-1{sup +} ones, and some of these cells express CD11c. Overall, this study demonstrates that neutrophils and monocytes transport tumor cell antigens from the peritoneal cavity to secondary lymphoid tissues.« less

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bidart, J.M.; Troalen, F.; Salesse, R.

1987-06-25

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

Antigenic Drift Defines a New D4 Subgenotype of Measles Virus.

PubMed

Muñoz-Alía, Miguel Ángel; Muller, Claude P; Russell, Stephen J

2017-06-01

The measles virus hemagglutinin (MeV-H) protein is the main target of protective neutralizing antibodies. Using a panel of monoclonal antibodies (MAbs) that recognize known major antigenic sites in MeV-H, we identified a D4 genotype variant that escapes neutralization by MAbs targeting the neutralizing epitope (NE) antigenic site. By site-directed mutagenesis, L249P was identified as the critical mutation disrupting the NE in this genotype D4 variant. Forty-two available D4 genotype gene sequences were subsequently analyzed and divided into 2 groups according to the presence or absence of the L249P MeV-H mutation. Further analysis of the MeV-N gene sequences of these 2 groups confirmed that they represent clearly definable, sequence-divergent D4 subgenotypes, which we named subgenotypes D4.1 and D4.2. The subgenotype D4.1 MeVs were isolated predominantly in Kenya and Ethiopia, whereas the MAb-resistant subgenotype D4.2 MeVs were isolated predominantly in France and Great Britain, countries with higher vaccine coverage rates. Interestingly, D4.2 subgenotype viruses showed a trend toward diminished susceptibility to neutralization by human sera pooled from approximately 60 to 80 North American donors. Escape from MAb neutralization may be a powerful epidemiological surveillance tool to monitor the evolution of new MeV subgenotypes. IMPORTANCE Measles virus is a paradigmatic RNA virus, as the antigenic composition of the vaccination has not needed to be updated since its discovery. The vaccine confers protection by inducing neutralizing antibodies that interfere with the function of the hemagglutinin protein. Viral strains are indistinguishable serologically, although characteristic nucleotide sequences differentiate 24 genotypes. In this work, we describe a distant evolutionary branch within genotype D4. Designated subgenotype D4.2, this virus is distinguishable by neutralization with vaccine-induced monoclonal antibodies that target the neutralizing epitope (NE

Antigenic Drift Defines a New D4 Subgenotype of Measles Virus

PubMed Central

Muller, Claude P.

2017-01-01

ABSTRACT The measles virus hemagglutinin (MeV-H) protein is the main target of protective neutralizing antibodies. Using a panel of monoclonal antibodies (MAbs) that recognize known major antigenic sites in MeV-H, we identified a D4 genotype variant that escapes neutralization by MAbs targeting the neutralizing epitope (NE) antigenic site. By site-directed mutagenesis, L249P was identified as the critical mutation disrupting the NE in this genotype D4 variant. Forty-two available D4 genotype gene sequences were subsequently analyzed and divided into 2 groups according to the presence or absence of the L249P MeV-H mutation. Further analysis of the MeV-N gene sequences of these 2 groups confirmed that they represent clearly definable, sequence-divergent D4 subgenotypes, which we named subgenotypes D4.1 and D4.2. The subgenotype D4.1 MeVs were isolated predominantly in Kenya and Ethiopia, whereas the MAb-resistant subgenotype D4.2 MeVs were isolated predominantly in France and Great Britain, countries with higher vaccine coverage rates. Interestingly, D4.2 subgenotype viruses showed a trend toward diminished susceptibility to neutralization by human sera pooled from approximately 60 to 80 North American donors. Escape from MAb neutralization may be a powerful epidemiological surveillance tool to monitor the evolution of new MeV subgenotypes. IMPORTANCE Measles virus is a paradigmatic RNA virus, as the antigenic composition of the vaccination has not needed to be updated since its discovery. The vaccine confers protection by inducing neutralizing antibodies that interfere with the function of the hemagglutinin protein. Viral strains are indistinguishable serologically, although characteristic nucleotide sequences differentiate 24 genotypes. In this work, we describe a distant evolutionary branch within genotype D4. Designated subgenotype D4.2, this virus is distinguishable by neutralization with vaccine-induced monoclonal antibodies that target the neutralizing epitope

Computational studies of H5N1 hemagglutinin binding with SA-{alpha}-2, 3-Gal and SA-{alpha}-2, 6-Gal

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li Minyong; Wang Binghe

2006-09-01

For influenza H5N1 hemagglutinin, a switch from SA-{alpha}-2, 3-Gal to SA-{alpha}-2, 6-Gal receptor specificity is a critical step leading to the conversion from avian-to-human to human-to-human infection. Therefore, the understanding of the binding modes of SA-{alpha}-2, 3-Gal and SA-{alpha}-2, 6-Gal to H5N1 hemagglutinin will be very important for the examination of possible mutations needed for going from an avian to a human flu virus. Based on the available H5N1 hemagglutinin crystal structure, the binding profiles between H5N1 hemagglutinin and two saccharide ligands, SA-{alpha}-2, 3-Gal and SA-{alpha}-2, 6-Gal, were investigated by ab initio quantum mechanics, molecular docking, molecular mechanics, and molecularmore » dynamics simulations. It was found that SA-{alpha}-2, 3-Gal has strong multiple hydrophobic and hydrogen bond interactions in its trans conformation with H5N1 hemagglutinin, whereas the SA-{alpha}-2, 6-Gal only shows weak interactions in a different conformation (cis type)« less

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

PubMed

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

2012-01-01

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

Inference of Genotype–Phenotype Relationships in the Antigenic Evolution of Human Influenza A (H3N2) Viruses

PubMed Central

Steinbrück, Lars; McHardy, Alice Carolyn

2012-01-01

Distinguishing mutations that determine an organism's phenotype from (near-) neutral ‘hitchhikers’ is a fundamental challenge in genome research, and is relevant for numerous medical and biotechnological applications. For human influenza viruses, recognizing changes in the antigenic phenotype and a strains' capability to evade pre-existing host immunity is important for the production of efficient vaccines. We have developed a method for inferring ‘antigenic trees’ for the major viral surface protein hemagglutinin. In the antigenic tree, antigenic weights are assigned to all tree branches, which allows us to resolve the antigenic impact of the associated amino acid changes. Our technique predicted antigenic distances with comparable accuracy to antigenic cartography. Additionally, it identified both known and novel sites, and amino acid changes with antigenic impact in the evolution of influenza A (H3N2) viruses from 1968 to 2003. The technique can also be applied for inference of ‘phenotype trees’ and genotype–phenotype relationships from other types of pairwise phenotype distances. PMID:22532796

Cutaneous basophil anaphylaxis. Immediate vasopermeability increases and anaphylactic degranulation of basophils at delayed hypersensitivity reactions challenged with additional antigen.

PubMed Central

Askenase, P W; Debernardo, R; Tauben, D; Kashgarian, M

1978-01-01

Many delayed-type reactions contain large infiltrates of basophils whose function is unknown. We have studied these cutaneous basophil hypersensitivity (CBH) reactions in guinea-pigs to ascertain whether basophils that are recruited to delayed reaction sites could be triggered for immediate reactivity. We compared 24 h CBH reactions with nearby skin for immediate hypersensitivity by challenging each site with small amounts of antigen. CBH sites had augmented immediate increases in vascular permeability detected by extravasation of Evan's blue dye. The ability to elicit this augmented anaphylactic phenomenon correlated with the local presence of basophils, and light microscopy at CBH reactions 15 min after antigen challenge showed a 50% decline in basophil counts. Electron microscopy showed that progressive anaphylactic-type degranulation of local basophils occurred within minutes following reintroduction of antigen. There was fusion of vacuoles containing granules, exocytosis of granules, and dissolution of granules, without ultrastructural disruption of cellular integrity. These results establish that basophils in CBH reactions can be triggered with soluble antigen to undergo anaphylactic degranulation, with the immediate release of vasoactive mediators. We have termed this phenomenon 'cutaneous basophil anaphylaxis'. Thus, one function of basophils at sites of delayed hypersensitivity may be to provide the potential for augmented, local, immediate anaphylactic reactivity. Images Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 PMID:721140

Cell-permeable nanobodies for targeted immunolabelling and antigen manipulation in living cells

NASA Astrophysics Data System (ADS)

Herce, Henry D.; Schumacher, Dominik; Schneider, Anselm F. L.; Ludwig, Anne K.; Mann, Florian A.; Fillies, Marion; Kasper, Marc-André; Reinke, Stefan; Krause, Eberhard; Leonhardt, Heinrich; Cardoso, M. Cristina; Hackenberger, Christian P. R.

2017-08-01

Functional antibody delivery in living cells would enable the labelling and manipulation of intracellular antigens, which constitutes a long-thought goal in cell biology and medicine. Here we present a modular strategy to create functional cell-permeable nanobodies capable of targeted labelling and manipulation of intracellular antigens in living cells. The cell-permeable nanobodies are formed by the site-specific attachment of intracellularly stable (or cleavable) cyclic arginine-rich cell-penetrating peptides to camelid-derived single-chain VHH antibody fragments. We used this strategy for the non-endocytic delivery of two recombinant nanobodies into living cells, which enabled the relocalization of the polymerase clamp PCNA (proliferating cell nuclear antigen) and tumour suppressor p53 to the nucleolus, and thereby allowed the detection of protein-protein interactions that involve these two proteins in living cells. Furthermore, cell-permeable nanobodies permitted the co-transport of therapeutically relevant proteins, such as Mecp2, into the cells. This technology constitutes a major step in the labelling, delivery and targeted manipulation of intracellular antigens. Ultimately, this approach opens the door towards immunostaining in living cells and the expansion of immunotherapies to intracellular antigen targets.

Performance of the cobas MRSA/SA Test for Simultaneous Detection of Methicillin-Susceptible and Methicillin-Resistant Staphylococcus aureus From Nasal Swabs.

PubMed

Peterson, Lance R; Woods, Christopher W; Davis, Thomas E; Wang, Zi-Xuam; Young, Stephen A; Osiecki, John C; Lewinski, Michael A; Liesenfeld, Oliver

2017-08-01

Health care-associated methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus (SA) infections are continuing problems. Rapidly determining the MRSA colonization status of a patient facilitates practice to reduce spread of MRSA clinical disease. Sensitive detection of all SA prior to surgery, followed by decolonization, can significantly reduce postoperative infection from this pathogen. Our goal was to validate a new automated assay for this testing. We compared performance of the cobas MRSA/SA Test on the cobas 4800 System to direct and enriched chromogenic culture using nasal swabs collected from patients at six United States sites. Compared to direct and enriched culture, the sensitivity for MRSA and SA was 93.1% and 93.9%, and the specificity was 97.5% and 94.2%, respectively. After discrepancy analysis, the sensitivity for MRSA and SA was 97.1% and 98.6%, and the specificity was 98.3% and 95.5%, respectively. Compared to direct culture, sensitivity for detecting any SA was 99.6%. The cobas MRSA/SA Test is an effective tool to simultaneously perform surveillance testing for nasal colonization of both MRSA and MSSA. © American Society for Clinical Pathology, 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

Targeted delivery of antigen to hamster nasal lymphoid tissue with M-cell-directed lectins.

PubMed Central

Giannasca, P J; Boden, J A; Monath, T P

1997-01-01

The nasal cavity of a rodent is lined by an epithelium organized into distinct regional domains responsible for specific physiological functions. Aggregates of nasal lymphoid tissue (NALT) located at the base of the nasal cavity are believed to be sites of induction of mucosal immune responses to airborne antigens. The epithelium overlying NALT contains M cells which are specialized for the transcytosis of immunogens, as demonstrated in other mucosal tissues. We hypothesized that NALT M cells are characterized by distinct glycoconjugate receptors which influence antigen uptake and immune responses to transcytosed antigens. To identify glycoconjugates that may distinguish NALT M cells from other cells of the respiratory epithelium (RE), we performed lectin histochemistry on sections of the hamster nasal cavity with a panel of lectins. Many classes of glycoconjugates were found on epithelial cells in this region. While most lectins bound to sites on both the RE and M cells, probes capable of recognizing alpha-linked galactose were found to label the follicle-associated epithelium (FAE) almost exclusively. By morphological criteria, the FAE contains >90% M cells. To determine if apical glycoconjugates on M cells were accessible from the nasal cavity, an M-cell-selective lectin and a control lectin in parallel were administered intranasally to hamsters. The M-cell-selective lectin was found to specifically target the FAE, while the control lectin did not. Lectin bound to M cells in vivo was efficiently endocytosed, consistent with the role of M cells in antigen transport. Intranasal immunization with lectin-test antigen conjugates without adjuvant stimulated induction of specific serum immunoglobulin G, whereas antigen alone or admixed with lectin did not. The selective recognition of NALT M cells by a lectin in vivo provides a model for microbial adhesin-host cell receptor interactions on M cells and the targeted delivery of immunogens to NALT following intranasal

Identification of antigens by monoclonal antibody PD4 and its expression in Escherichia coli

PubMed Central

Ning, Jin-Ying; Sun, Guo-Xun; Huang, Su; Ma, Hong; An, Ping; Meng, Lin; Song, Shu-Mei; Wu, Jian; Shou, Cheng-Chao

2003-01-01

AIM: To clone and express the antigen of monoclonal antibody (MAb) PD4 for further investigation of its function. METHODS: MGC803 cDNA expression library was constructed and screened with PD4 as probes to clone the antigen. After failed in the library screening, immunoprecipitation and SDS-polyacrylamide gel electrophoresis were applied to purify the antigen for sequence analysis. The antigen coming from Mycoplasma hyorhinis (M. hyorhinis) was further confirmed with Western blot analysis by infecting M. hyorhinis -free HeLa cells and eliminating the M. hyorhinis from MGC803 cells. The full p37 gene was cloned by PCR and expressed successfully in Escherichia coli after site-directed mutations. Immunofluorescence assay was used to demonstrate if p37 protein could directly bind to gastric tumor cell AGS. RESULTS: The cDNA library constructed with MGC803 cells was screened by MAb PD4 as probes. Unfortunately, the positive clones identified with MAb PD4 were also reacted with unrelated antibodies. Then, immunoprecipitation was performed and the purified antigen was identified to be a membrane protein of Mycoplasma hyorhinis (M. hyorhinis) by sequencing of N-terminal amino acid residues. The membrane protein was intensively verified with Western blot by eliminating M. hyorhinis from MGC803 cells and by infecting M. hyorhinis-free HeLa cells. The full p37 gene was cloned and expressed successfully in Escherichia coli after site-directed mutations. Immunofluorescence demonstrated that p37 protein could directly bind to gastric tumor cell AGS. CONCLUSION: The antigen recognized by MAb PD4 is from M. hyorhinis, which suggests the actions involved in MAb PD4 is possibly mediated by p37 protein or M. hyorhinis. As p37 protein can bind directly to tumor cells, the pathogenic role of p37 involved in tumorigenesis justifies further investigation. PMID:14562370

Antigen-B Cell Receptor Complexes Associate with Intracellular major histocompatibility complex (MHC) Class II Molecules*

PubMed Central

Barroso, Margarida; Tucker, Heidi; Drake, Lisa; Nichol, Kathleen; Drake, James R.

2015-01-01

Antigen processing and MHC class II-restricted antigen presentation by antigen-presenting cells such as dendritic cells and B cells allows the activation of naïve CD4+ T cells and cognate interactions between B cells and effector CD4+ T cells, respectively. B cells are unique among class II-restricted antigen-presenting cells in that they have a clonally restricted antigen-specific receptor, the B cell receptor (BCR), which allows the cell to recognize and respond to trace amounts of foreign antigen present in a sea of self-antigens. Moreover, engagement of peptide-class II complexes formed via BCR-mediated processing of cognate antigen has been shown to result in a unique pattern of B cell activation. Using a combined biochemical and imaging/FRET approach, we establish that internalized antigen-BCR complexes associate with intracellular class II molecules. We demonstrate that the M1-paired MHC class II conformer, shown previously to be critical for CD4 T cell activation, is incorporated selectively into these complexes and loaded selectively with peptide derived from BCR-internalized cognate antigen. These results demonstrate that, in B cells, internalized antigen-BCR complexes associate with intracellular MHC class II molecules, potentially defining a site of class II peptide acquisition, and reveal a selective role for the M1-paired class II conformer in the presentation of cognate antigen. These findings provide key insights into the molecular mechanisms used by B cells to control the source of peptides charged onto class II molecules, allowing the immune system to mount an antibody response focused on BCR-reactive cognate antigen. PMID:26400081

1HNMR study of methotrexate serum albumin (MTX SA) binding in rheumatoid arthritis

NASA Astrophysics Data System (ADS)

Sułkowska, A.; Maciążek-Jurczyk, M.; Bojko, B.; Równicka, J.; Sułkowski, W. W.

2008-11-01

Rheumatoid arthritis (RA) is an immunologically depended disease. It is characterized by a chronic, progressive inflammatory process. Methotrexate (4-amino-10-methylfolic acid, MTX) is the modifying drug used to treat RA. The aim of the presented studies is to determine the low affinity binding site of MTX in bovine (BSA) and human (HSA) serum albumin with the use of proton nuclear magnetic resonance ( 1HNMR) spectroscopy. The analysis of 1HNMR spectra of MTX in the presence of serum albumin (SA) allows us to observe the interactions between aromatic rings of the drug and the rings of amino acids located in the hydrophobic subdomains of the protein. On the basis of the chemical shifts σ [ppm] and the relaxation times T1 [s] of drug protons the hydrophobic interaction between MTX-SA and the stoichiometric molar ratio of the complex was evaluated. This work is a part of a spectroscopic study on MTX-SA interactions [A. Sułkowska, M. Maciążek, J. Równicka, B. Bojko, D. Pentak, W.W. Sułkowski, J. Mol. Struct. 834-836 (2007) 162-169].

Highly stretchable HA/SA hydrogels for tissue engineering.

PubMed

Zhu, Chengcheng; Yang, Rui; Hua, Xiaobin; Chen, Hong; Xu, Jumei; Wu, Rile; Cen, Lian

2018-04-01

A highly stretchable hyaluronic acid (HA)/sodium alginate (SA) hydrogel was developed in this study based on an interpenetrating polymer network. HA/SA hydrogels were prepared by mixing two polysaccharides followed by covalent crosslinking via epoxy groups on HA molecules and ionic crosslinking via divalent ions on SA chains sequentially. The effect of HA/SA ratio on the pore size and distribution, swelling ratio, elongation and rheological properties as well as protein loading and release properties of HA/SA hydrogels was explored. Moreover, a surface modification method, layer-by-layer (LBL) assembly technique, was applied to modify the hydrogel to evaluate the hydrogel's tenability in varying biological performance. It was then shown that the hydrogels had the pore sizes ranging from 100 to 50 μm. With the increase in SA content of the resulting hydrogels, the pore size, swelling ratio, and storage modulus (G') and loss modulus (G″) of the hydrogel all decreased, whereas the in vitro bulk weight loss was fastened. Moreover, elongation at break (EB) value increased first, reached a peak value and then decreased, that is HA8/SA1 (HA:SA = 8:1) had the highest EB value of 417%. This hydrogel could retain 33.2% of the pre-loaded protein even after 72 h, which could be further attenuated when LBL was used to shell the hydrogel. The growth of fibroblasts on HA8/SA1 hydrogel gave preliminary assessment on its suitability as a cellular carrier, while the LBL modified HA8/SA1 hydrogel also favored the anchoring of keratinocytes, further enhancing its cell carrier role for tissue regeneration, especially skin engineering.

IS THE IMMUNOCROMATOGRAPHIC FECAL ANTIGEN TEST EFFECTIVE FOR PRIMARY DIAGNOSIS OF HELICOBACTER PYLORI INFECTION IN DYSPEPTIC PATIENTS?

PubMed

Dalla Nora, Magali; Hörner, Rosmari; De Carli, Diego Michelon; Rocha, Marta Pires da; Araujo, Amanda Faria de; Fagundes, Renato Borges

2016-01-01

The diagnosis of H. pylori infection can be performed by non-invasive and invasive methods.The identification through a fecal antigen test is a non-invasive, simple, and relatively inexpensive test. To determine the diagnostic performance of fecal antigen test in the identification of H. pylori infection. H. pylori antigens were identified in the stools of dyspeptic patients undergoing upper gastrointestinal endoscopy. For the identification of H. pylori antigen, we use ImmunoCard STAT! HpSA with immunochromatography technique. Histopathology plus urease test were the gold standard. We studied 163 patients, 51% male, mean age of 56.7± 8.5years. H. pylori infection was present in 49%. Fecal test presented: sensitivity 67.5% (CI95% 60.6-72.9); specificity 85.5% (CI95% 78.9-90.7); positive predictive value 81.8% (CI95% 73.4-88.4) and negative predictive value 73,2% (CI95% 67.5-77.6); Positive likelihood ratio was 4.7 (CI95% 2.9-7.9) and Negative Likelihood Ratio 0.4 (CI95% 0.3-0.5). The prevalence odds ratio for a positive test was 12.3 (CI95% 5.7-26.3).The index kappa between FAT and histology/urease test was 0.53 (CI95% 0.39-0.64). Immunochromatographic FAT is less expensive than the other methods and readily accepted by the patients but its diagnostic performance does not recommend its use in the primary diagnosis, when the patient may have an active infection.

The antigenicity in guinea pigs and monkeys of three mycobacterial polysaccharides purified by affinity chromatography with concanavalin A.

PubMed

Daniel, T M

1975-06-01

The antigenicity of 3 polysaccharides purified from culture filtrates of Mycobacterim tuberculosis by affinity chromatography using a concanavalin A-agarose absorbent was studied. All 3 purified polysaccharides were found to be potent elicitors of delayed skin test reactions in sensitized guinea pigs and in a tuberculos monkey. This antigenicity could not be attributed to contaminating protein. Small dermal reactions were also observed in control guinea pigs. All 3 polysaccharides reacted with precipitating antibody in guinea pig sera, the antigenic specificity observed with the guinea pig sera differing from that demonstrated with reference goat antiserum. The 3 polysaccharides were also demonstrated to contain hemagglutination antigenic sites.

Antigenic Evolution of Vaccine-Derived Polioviruses: Changes in Individual Epitopes and Relative Stability of the Overall Immunological Properties

PubMed Central

Yakovenko, Maria L.; Cherkasova, Elena A.; Rezapkin, Gennady V.; Ivanova, Olga E.; Ivanov, Alexander P.; Eremeeva, Tatyana P.; Baykova, Olga Y.; Chumakov, Konstantin M.; Agol, Vadim I.

2006-01-01

The Sabin oral poliovirus vaccine (OPV) readily undergoes changes in antigenic sites upon replication in humans. Here, a set of antigenically altered descendants of the three OPV serotypes (76 isolates) was characterized to determine the driving forces behind these changes and their biological implications. The amino acid residues of OPV derivatives that lie within or close to the known antigenic sites exhibited a marked tendency to be replaced by residues characteristic of homotypic wild polioviruses, and these changes may occur very early in OPV evolution. The specific amino acid alterations nicely correlated with serotype-specific changes in the reactivity of certain individual antigenic sites, as revealed by the recently devised monoclonal antibody-based enzyme-linked immunosorbent assay. In comparison to the original vaccine, small changes, if any, in the neutralizing capacity of human or rabbit sera were observed in highly diverged vaccine polioviruses of three serotypes, in spite of strong alterations of certain epitopes. We propose that the common antigenic alterations in evolving OPV strains largely reflect attempts to eliminate fitness-decreasing mutations acquired either during the original selection of the vaccine or already present in the parental strains. Variability of individual epitopes does not appear to be primarily caused by, or lead to, a significant immune evasion, enhancing only slightly, if at all, the capacity of OPV derivatives to overcome immunity in human populations. This study reveals some important patterns of poliovirus evolution and has obvious implications for the rational design of live viral vaccines. PMID:16501074

Calcium-dependent antigen binding as a novel modality for antibody recycling by endosomal antigen dissociation

PubMed Central

Hironiwa, N; Ishii, S; Kadono, S; Iwayanagi, Y; Mimoto, F; Habu, K; Igawa, T; Hattori, K

2016-01-01

The pH-dependent antigen binding antibody, termed a recycling antibody, has recently been reported as an attractive type of second-generation engineered therapeutic antibody. A recycling antibody can dissociate antigen in the acidic endosome, and thus bind to its antigen multiple times. As a consequence, a recycling antibody can neutralize large amounts of antigen in plasma. Because this approach relies on histidine residues to achieve pH-dependent antigen binding, which could limit the epitopes that can be targeted and affect the rate of antigen dissociation in the endosome, we explored an alternative approach for generating recycling antibodies. Since calcium ion concentration is known to be lower in endosome than in plasma, we hypothesized that an antibody with antigen-binding properties that are calcium-dependent could be used as recycling antibody. Here, we report a novel anti-interleukin-6 receptor (IL-6R) antibody, identified from a phage library that binds to IL-6R only in the presence of a calcium ion. Thermal dynamics and a crystal structure study revealed that the calcium ion binds to the heavy chain CDR3 region (HCDR3), which changes and possibly stabilizes the structure of HCDR3 to make it bind to antigen calcium dependently (PDB 5AZE). In vitro and in vivo studies confirmed that this calcium-dependent antigen-binding antibody can dissociate its antigen in the endosome and accelerate antigen clearance from plasma, making it a novel approach for generating recycling antibody. PMID:26496237

AmeriFlux BR-Sa1 Santarem-Km67-Primary Forest

DOE Data Explorer

Saleska, Scott [University of Arizona

2016-01-01

This is the AmeriFlux version of the carbon flux data for the site BR-Sa1 Santarem-Km67-Primary Forest. Site Description - The LBA Tapajos KM67 Mature Forest site is located in the Tapajos National Forest, a 450,000 ha closed-canopy upland forest in Amazonian Brazil. Bounded by the Tapajos River in the west and highway BR-163 to the east, the tower is located on a flat plateau (or planalto) that extends up to 150 km to the north, south, and east. Within the confines of the National Forest, anthropogenic disturbances are limited to a few small hunting trails. The surrounding stand is classified as primary or "old-growth"" predominantly by its uneven age distribution, emergent trees, numerous epiphytes and abundant large logs. In 2007 falling trees hit the tower guy wires rendering all instrumentation in-operational. After a complete restoration tower measurements resumed in August of 2008.

Antigen presenting capacity of murine splenic myeloid cells.

PubMed

Hey, Ying-Ying; Quah, Benjamin; O'Neill, Helen C

2017-01-11

The spleen is an important site for hematopoiesis. It supports development of myeloid cells from bone marrow-derived precursors entering from blood. Myeloid subsets in spleen are not well characterised although dendritic cell (DC) subsets are clearly defined in terms of phenotype, development and functional role. Recently a novel dendritic-like cell type in spleen named 'L-DC' was distinguished from other known dendritic and myeloid cells by its distinct phenotype and developmental origin. That study also redefined splenic eosinophils as well as resident and inflammatory monocytes in spleen. L-DC are shown to be distinct from known splenic macrophages and monocyte subsets. Using a new flow cytometric procedure, it has been possible to identify and isolate L-DC in order to assess their functional competence and ability to activate T cells both in vivo and in vitro. L-DC are readily accessible to antigen given intravenously through receptor-mediated endocytosis. They are also capable of CD8 + T cell activation through antigen cross presentation, with subsequent induction of cytotoxic effector T cells. L-DC are MHCII - cells and unable to activate CD4 + T cells, a property which clearly distinguishes them from conventional DC. The myeloid subsets of resident monocytes, inflammatory monocytes, neutrophils and eosinophils, were found to have varying capacities to take up antigen, but were uniformly unable to activate either CD4 + T cells or CD8 + T cells. The results presented here demonstrate that L-DC in spleen are distinct from other myeloid cells in that they can process antigen for CD8 + T cell activation and induction of cytotoxic effector function, while both L-DC and myeloid subsets remain unable to activate CD4 + T cells. The L-DC subset in spleen is therefore distinct as an antigen presenting cell.

Chlorphenesin: an Antigen-Associated Immunosuppressant

PubMed Central

Whang, H. Y.; Neter, E.

1970-01-01

Chlorphenesin (3-p-chlorophenoxy-1,2-propanediol), when injected intravenously together with either of two common bacterial antigens, inhibits the antibody response of the rabbit. The antigens studied are those common to Enterobacteriaceae and to gram-positive bacteria. The immunosuppression is contingent upon incubation of chlorphenesin and antigen in vitro prior to administration, since separate injection of antigen and inhibitor or of mixtures without prior incubation yields undiminished antibody response. Chlorphenesin, as shown by hemagglutination-inhibition tests, does not alter the antigenic determinants, because antibody neutralization occurs in the presence or absence of the drug. The immunosuppressive effect is reversible, since precipitation of chlorphenesin at 4 C substantially restores immunogenicity. Animals immunized with antigen-drug mixtures, which fail to respond with significant antibody production, nonetheless are immunologically primed. It is concluded that chlorphenesin represents another example of antigen-associated immunosuppressants. PMID:16557800

Chlorphenesin: an antigen-associated immunosuppressant.

PubMed

Whang, H Y; Neter, E

1970-07-01

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

Proliferating Cell Nuclear Antigen-dependent Rapid Recruitment of Cdt1 and CRL4Cdt2 at DNA-damaged Sites after UV Irradiation in HeLa Cells*

PubMed Central

Ishii, Takashi; Shiomi, Yasushi; Takami, Toshihiro; Murakami, Yusuke; Ohnishi, Naho; Nishitani, Hideo

2010-01-01

The licensing factor Cdt1 is degraded by CRL4Cdt2 ubiquitin ligase dependent on proliferating cell nuclear antigen (PCNA) during S phase and when DNA damage is induced in G1 phase. Association of both Cdt2 and PCNA with chromatin was observed in S phase and after UV irradiation. Here we used a micropore UV irradiation assay to examine Cdt2 accumulation at cyclobutane pyrimidine dimer-containing DNA-damaged sites in the process of Cdt1 degradation in HeLa cells. Cdt2, present in the nucleus throughout the cell cycle, accumulated rapidly at damaged DNA sites during G1 phase. The recruitment of Cdt2 is dependent on prior PCNA chromatin binding because Cdt2 association was prevented when PCNA was silenced. Cdt1 was also recruited to damaged sites soon after UV irradiation through its PIP-box. As Cdt1 was degraded, the Cdt2 signal at damaged sites was reduced, but PCNA, cyclobutane pyrimidine dimer, and XPA (xeroderma pigmentosum, complementation group A) signals remained at the same levels. These findings suggest that Cdt1 degradation following UV irradiation occurs rapidly at damaged sites due to PCNA chromatin loading and the recruitment of Cdt1 and CRL4Cdt2, before DNA damage repair is completed. PMID:20929861

Population structuring of multi-copy, antigen-encoding genes in Plasmodium falciparum

PubMed Central

Artzy-Randrup, Yael; Rorick, Mary M; Day, Karen; Chen, Donald; Dobson, Andrew P; Pascual, Mercedes

2012-01-01

The coexistence of multiple independently circulating strains in pathogen populations that undergo sexual recombination is a central question of epidemiology with profound implications for control. An agent-based model is developed that extends earlier ‘strain theory’ by addressing the var gene family of Plasmodium falciparum. The model explicitly considers the extensive diversity of multi-copy genes that undergo antigenic variation via sequential, mutually exclusive expression. It tracks the dynamics of all unique var repertoires in a population of hosts, and shows that even under high levels of sexual recombination, strain competition mediated through cross-immunity structures the parasite population into a subset of coexisting dominant repertoires of var genes whose degree of antigenic overlap depends on transmission intensity. Empirical comparison of patterns of genetic variation at antigenic and neutral sites supports this role for immune selection in structuring parasite diversity. DOI: http://dx.doi.org/10.7554/eLife.00093.001 PMID:23251784

Chromomycin SA analogs from a marine-derived Streptomyces sp.

PubMed Central

Hu, Youcai; Espindola, Ana Paula D. M; Stewart, Nathan A.; Wei, Shuguang; Posner, Bruce A.; MacMillan, John B.

2011-01-01

Two chromomycin SA analogs, chromomycin SA3 and chromomycin SA2, along with deacetylchromomycin A3 and five previously reported chromomycin analogs were isolated from a marine-derived Streptomyces sp. The structures of the new compounds were determined by spectroscopic methods including 1D and 2D NMR techniques, HRMS and chemical methods. Chromomycin SA3 and chromomycin SA2 are the first naturally occuring chromomycin analogs with truncated side-chains. Biological evaluation of chromomycin analogs for cytotoxicity against two non-small cell lung cancer (NSCLC) cell-lines, A549 and HCC44, demonstrated a decrease in cytotoxicity for the truncated sides chain chromomycin analogs. PMID:21807523

Comprehensive red blood cell and platelet antigen prediction from whole genome sequencing: proof of principle

PubMed Central

Westhoff, Connie M.; Uy, Jon Michael; Aguad, Maria; Smeland‐Wagman, Robin; Kaufman, Richard M.; Rehm, Heidi L.; Green, Robert C.; Silberstein, Leslie E.

2015-01-01

BACKGROUND There are 346 serologically defined red blood cell (RBC) antigens and 33 serologically defined platelet (PLT) antigens, most of which have known genetic changes in 45 RBC or six PLT genes that correlate with antigen expression. Polymorphic sites associated with antigen expression in the primary literature and reference databases are annotated according to nucleotide positions in cDNA. This makes antigen prediction from next‐generation sequencing data challenging, since it uses genomic coordinates. STUDY DESIGN AND METHODS The conventional cDNA reference sequences for all known RBC and PLT genes that correlate with antigen expression were aligned to the human reference genome. The alignments allowed conversion of conventional cDNA nucleotide positions to the corresponding genomic coordinates. RBC and PLT antigen prediction was then performed using the human reference genome and whole genome sequencing (WGS) data with serologic confirmation. RESULTS Some major differences and alignment issues were found when attempting to convert the conventional cDNA to human reference genome sequences for the following genes: ABO, A4GALT, RHD, RHCE, FUT3, ACKR1 (previously DARC), ACHE, FUT2, CR1, GCNT2, and RHAG. However, it was possible to create usable alignments, which facilitated the prediction of all RBC and PLT antigens with a known molecular basis from WGS data. Traditional serologic typing for 18 RBC antigens were in agreement with the WGS‐based antigen predictions, providing proof of principle for this approach. CONCLUSION Detailed mapping of conventional cDNA annotated RBC and PLT alleles can enable accurate prediction of RBC and PLT antigens from whole genomic sequencing data. PMID:26634332

Rules of co-occurring mutations characterize the antigenic evolution of human influenza A/H3N2, A/H1N1 and B viruses.

PubMed

Chen, Haifen; Zhou, Xinrui; Zheng, Jie; Kwoh, Chee-Keong

2016-12-05

The human influenza viruses undergo rapid evolution (especially in hemagglutinin (HA), a glycoprotein on the surface of the virus), which enables the virus population to constantly evade the human immune system. Therefore, the vaccine has to be updated every year to stay effective. There is a need to characterize the evolution of influenza viruses for better selection of vaccine candidates and the prediction of pandemic strains. Studies have shown that the influenza hemagglutinin evolution is driven by the simultaneous mutations at antigenic sites. Here, we analyze simultaneous or co-occurring mutations in the HA protein of human influenza A/H3N2, A/H1N1 and B viruses to predict potential mutations, characterizing the antigenic evolution. We obtain the rules of mutation co-occurrence using association rule mining after extracting HA1 sequences and detect co-mutation sites under strong selective pressure. Then we predict the potential drifts with specific mutations of the viruses based on the rules and compare the results with the "observed" mutations in different years. The sites under frequent mutations are in antigenic regions (epitopes) or receptor binding sites. Our study demonstrates the co-occurring site mutations obtained by rule mining can capture the evolution of influenza viruses, and confirms that cooperative interactions among sites of HA1 protein drive the influenza antigenic evolution.

Serological purification of polysaccharide antigens from Streptococcus mutans serotypes a and d: characterization of multiple antigenic determinants.

PubMed

Linzer, R; Mukasa, H; Slade, H D

1975-10-01

The polysaccharide antigen preparations from serotype a and serotype d strains of Streptococcus mutans contained both a serotype-specific antigenic determinant and a common a-d antigenic determinant, as demonstrated by agar gel diffusion studies and a quantitative cross-precipitin assay. The chromatographically purified antigens were isolated by a method which depended on their serological specificity to determine if these two antigenic determinants were located on the same molecule. The a and d polysaccharides were recovered from specific antigen-antibody complexes and characterized with respect to their immunological specificity and chemical composition. Agar gel diffusion tests demonstrated that, in both the a and d preparations, the serotype-specific antigenic determinant and the common a-d antigenic determinant were present in one molecule.

Definition of a virulence-related antigen of Neisseria gonorrhoeae with monoclonal antibodies and lectins.

PubMed

Demarco de Hormaeche, R; Bundell, C; Chong, H; Taylor, D W; Wildy, P

1986-03-01

Variants of one strain of Neisseria gonorrhoeae, grown in vivo or in vitro, that have been previously shown to differ in infectivity, serum resistance, and capsule production were compared with use of monoclonal antibodies and lectins. Monoclonal antibodies to virulent gonococci recognized an antigenic site of the lipopolysaccharide (LPS) produced in large amounts by gonococci grown in vivo but present only in a small proportion of in vitro-grown gonococci. This antigen (C-LPS) was found in all 85 different gonococcal isolates studied but not among nonpathogenic neisseriae. It was shared by group B and C meningococci but not by groups A and D. Enzyme-linked immunosorbent assay and Western blot analysis showed that N-acetylglucosamine and N-acetylgalactosamine form part of the epitope. The C-LPS antigen was shown by immunofluorescence to be present on the surface of the gonococci and also free as slime. This antigen appears to confer resistance to killing by normal sera.

MimoSA: a system for minimotif annotation

PubMed Central

2010-01-01

Background Minimotifs are short peptide sequences within one protein, which are recognized by other proteins or molecules. While there are now several minimotif databases, they are incomplete. There are reports of many minimotifs in the primary literature, which have yet to be annotated, while entirely novel minimotifs continue to be published on a weekly basis. Our recently proposed function and sequence syntax for minimotifs enables us to build a general tool that will facilitate structured annotation and management of minimotif data from the biomedical literature. Results We have built the MimoSA application for minimotif annotation. The application supports management of the Minimotif Miner database, literature tracking, and annotation of new minimotifs. MimoSA enables the visualization, organization, selection and editing functions of minimotifs and their attributes in the MnM database. For the literature components, Mimosa provides paper status tracking and scoring of papers for annotation through a freely available machine learning approach, which is based on word correlation. The paper scoring algorithm is also available as a separate program, TextMine. Form-driven annotation of minimotif attributes enables entry of new minimotifs into the MnM database. Several supporting features increase the efficiency of annotation. The layered architecture of MimoSA allows for extensibility by separating the functions of paper scoring, minimotif visualization, and database management. MimoSA is readily adaptable to other annotation efforts that manually curate literature into a MySQL database. Conclusions MimoSA is an extensible application that facilitates minimotif annotation and integrates with the Minimotif Miner database. We have built MimoSA as an application that integrates dynamic abstract scoring with a high performance relational model of minimotif syntax. MimoSA's TextMine, an efficient paper-scoring algorithm, can be used to dynamically rank papers with

Telomere interactions may condition the programming of antigen expression in Trypanosoma brucei.

PubMed Central

Van der Werf, A; Van Assel, S; Aerts, D; Steinert, M; Pays, E

1990-01-01

The AnTat 1.1 antigen type typically occurs late in a chronic infection by the EATRO 1125 stock of Trypanosoma brucei. The AnTat 1.1 gene, which is located 24 kb from a chromosome end, seems exclusively expressed by acting as a donor in gene conversion events targeted to the telomeric expression site. We report that this gene is sufficiently provided with the homology blocks required for recombination with the expression site, and is not interrupted by stop codons up to the 3' block of homology. A possible reason for its low probability of activation is an inverse orientation with respect to the proximal chromosome end, since, if correctly positioned, it is readily expressed at an early stage of infection, following gene conversion. This suggests that interactions between chromosome ends may precede and favour the rearrangements leading to antigenic variation. Images Fig. 1. Fig. 3. Fig. 4. Fig. 5. PMID:2323332

Antigenicity-defined conformations of an extremely neutralization-resistant HIV-1 envelope spike

DOE PAGES

Cai, Yongfei; Karaca-Griffin, Selen; Chen, Jia; ...

2017-04-10

Here, the extraordinary genetic diversity of the HIV-1 envelope spike [Env; trimeric (gp160) 3, cleaved to (gp120/gp41) 3] poses challenges for vaccine development. Envs of different clinical isolates exhibit different sensitivities to antibody-mediated neutralization. Envs of difficult-to-neutralize viruses are thought to be more stable and conformationally homogeneous trimers than those of easy-to-neutralize viruses, thereby providing more effective concealment of conserved, functionally critical sites. In this study we have characterized the antigenic properties of an Env derived from one of the most neutralization-resistant HIV-1 isolates, CH120.6. Sequence variation at neutralizing epitopes does not fully account for its exceptional resistance to antibodies.more » The full-length, membrane-bound CH120.6 Env is indeed stable and conformationally homogeneous. Its antigenicity correlates closely with its neutralization sensitivity, and major changes in antigenicity upon CD4 engagement appear to be restricted to the coreceptor site. The CH120.6 gp140 trimer, the soluble and uncleaved ectodomain of (gp160) 3, retains many antigenic properties of the intact Env, consistent with a conformation close to that of Env spikes on a virion, whereas its monomeric gp120 exposes many nonneutralizing or strain-specific epitopes. Thus, trimer organization and stability are important determinants not only for occluding many epitopes but also for conferring resistance to neutralization by all but a small set of antibodies. Env preparations derived from neutralization-resistant viruses may induce irrelevant antibody responses less frequently than do other Envs and may be excellent templates for developing soluble immunogens.« less

Antigenicity-defined conformations of an extremely neutralization-resistant HIV-1 envelope spike

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cai, Yongfei; Karaca-Griffin, Selen; Chen, Jia

Here, the extraordinary genetic diversity of the HIV-1 envelope spike [Env; trimeric (gp160) 3, cleaved to (gp120/gp41) 3] poses challenges for vaccine development. Envs of different clinical isolates exhibit different sensitivities to antibody-mediated neutralization. Envs of difficult-to-neutralize viruses are thought to be more stable and conformationally homogeneous trimers than those of easy-to-neutralize viruses, thereby providing more effective concealment of conserved, functionally critical sites. In this study we have characterized the antigenic properties of an Env derived from one of the most neutralization-resistant HIV-1 isolates, CH120.6. Sequence variation at neutralizing epitopes does not fully account for its exceptional resistance to antibodies.more » The full-length, membrane-bound CH120.6 Env is indeed stable and conformationally homogeneous. Its antigenicity correlates closely with its neutralization sensitivity, and major changes in antigenicity upon CD4 engagement appear to be restricted to the coreceptor site. The CH120.6 gp140 trimer, the soluble and uncleaved ectodomain of (gp160) 3, retains many antigenic properties of the intact Env, consistent with a conformation close to that of Env spikes on a virion, whereas its monomeric gp120 exposes many nonneutralizing or strain-specific epitopes. Thus, trimer organization and stability are important determinants not only for occluding many epitopes but also for conferring resistance to neutralization by all but a small set of antibodies. Env preparations derived from neutralization-resistant viruses may induce irrelevant antibody responses less frequently than do other Envs and may be excellent templates for developing soluble immunogens.« less

Beyond SaGMRotI: Conversion to SaArb, SaSN, and SaMaxRot

USGS Publications Warehouse

Watson-Lamprey, J. A.; Boore, D.M.

2007-01-01

In the seismic design of structures, estimates of design forces are usually provided to the engineer in the form of elastic response spectra. Predictive equations for elastic response spectra are derived from empirical recordings of ground motion. The geometric mean of the two orthogonal horizontal components of motion is often used as the response value in these predictive equations, although it is not necessarily the most relevant estimate of forces within the structure. For some applications it is desirable to estimate the response value on a randomly chosen single component of ground motion, and in other applications the maximum response in a single direction is required. We give adjustment factors that allow converting the predictions of geometric-mean ground-motion predictions into either of these other two measures of seismic ground-motion intensity. In addition, we investigate the relation of the strike-normal component of ground motion to the maximum response values. We show that the strike-normal component of ground motion seldom corresponds to the maximum horizontal-component response value (in particular, at distances greater than about 3 km from faults), and that focusing on this case in exclusion of others can result in the underestimation of the maximum component. This research provides estimates of the maximum response value of a single component for all cases, not just near-fault strike-normal components. We provide modification factors that can be used to convert predictions of ground motions in terms of the geometric mean to the maximum spectral acceleration (SaMaxRot) and the random component of spectral acceleration (SaArb). Included are modification factors for both the mean and the aleatory standard deviation of the logarithm of the motions.

Ginger (Zingiber officinale) phytochemicals-gingerenone-A and shogaol inhibit SaHPPK: molecular docking, molecular dynamics simulations and in vitro approaches.

PubMed

Rampogu, Shailima; Baek, Ayoung; Gajula, Rajesh Goud; Zeb, Amir; Bavi, Rohit S; Kumar, Raj; Kim, Yongseong; Kwon, Yong Jung; Lee, Keun Woo

2018-04-02

Antibiotic resistance is a defense mechanism, harbored by pathogens to survive under unfavorable conditions. Among several antibiotic resistant microbial consortium, Staphylococcus aureus is one of the most havoc microorganisms. Staphylococcus aureus encodes a unique enzyme 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (SaHPPK), against which, none of existing antibiotics have been reported. Computational approaches have been instrumental in designing and discovering new drugs for several diseases. The present study highlights the impact of ginger phytochemicals on Staphylococcus aureus SaHPPK. Herein, we have retrieved eight ginger phytochemicals from published literature and investigated their inhibitory interactions with SaHPPK. To authenticate our work, the investigation proceeds considering the known antibiotics alongside the phytochemicals. Molecular docking was performed employing GOLD and CDOCKER. The compounds with the highest dock score from both the docking programmes were tested for their inhibitory capability in vitro. The binding conformations that were seated within the binding pocket showing strong interactions with the active sites residues rendered by highest dock score were forwarded towards the molecular dynamic (MD) simulation analysis. Based on molecular dock scores, molecular interaction with catalytic active residues and MD simulations studies, two ginger phytochemicals, gingerenone-A and shogaol have been proposed as candidate inhibitors against Staphylococcus aureus. They have demonstrated higher dock scores than the known antibiotics and have represented interactions with the key residues within the active site. Furthermore, these compounds have rendered considerable inhibitory activity when tested in vitro. Additionally, their superiority was corroborated by stable MD results conducted for 100 ns employing GROMACS package. Finally, we suggest that gingerenone-A and shogaol may either be potential SaHPPK inhibitors or can be used

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

PubMed

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

2004-10-01

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

Identification of the Mycobacterium marinum Apa antigen O-mannosylation sites reveals important glycosylation variability with the M. tuberculosis Apa homologue.

PubMed

Coddeville, Bernadette; Wu, Sz-Wei; Fabre, Emeline; Brassart, Colette; Rombouts, Yoann; Burguière, Adeline; Kremer, Laurent; Khoo, Kay-Hooi; Elass-Rochard, Elisabeth; Guérardel, Yann

2012-10-22

The 45/47 kDa Apa, an immuno-dominant antigen secreted by Mycobacterium tuberculosis is O-mannosylated at multiple sites. Glycosylation of Apa plays a key role in colonization and invasion of the host cells by M. tuberculosis through interactions of Apa with the host immune system C-type lectins. Mycobacterium marinum (M.ma) a fish pathogen, phylogenetically close to M. tuberculosis, induces a granulomatous response with features similar to those described for M. tuberculosis in human. Although M.ma possesses an Apa homologue, its glycosylation status is unknown, and whether this represents a crucial element in the pathophysiology induced by M.ma remains to be addressed. To this aim, we have identified two concanavalin A-reactive 45/47 kDa proteins from M.ma, which have been further purified by a two-step anion exchange chromatography process. Advanced liquid chromatography-nanoESI mass spectrometry-based proteomic analyses of peptides, derived from either tryptic digestion alone or in combination with the Asp-N endoproteinase, established that M.ma Apa possesses up to seven distinct O-mannosylated sites with mainly single mannose substitutions, which can be further extended at the Ser/Thr/Pro rich region near the N-terminus. This opens the way to further studies focussing on the involvement and biological functions of Apa O-mannosylation using the M.ma/zebrafish model. Copyright © 2012 Elsevier B.V. All rights reserved.

In vitro antigen-induced, antigen-specific antibody production in man. Specific and polyclonal components, kinetics, and cellular requirements

PubMed Central

1981-01-01

A highly specific and reproducible antigen-induced, antigen-specific culture and assay system for antibody production by human peripheral blood B lymphocytes has been developed. The system is clearly T cell and monocyte dependent and is independent of exogenous mitogens. The major factors in our ability to trigger specific antibody production with antigen alone have been the use of extremely low concentrations of antigen in vitro (doses several orders of magnitude below those inducing a peak blastogenic response), careful attention to in vitro cell density and culture vessel geometry, and appreciation of the kinetics of the circulating antigen-inducible B cell repertoire. A dichotomy and overlap between antigen-induced, antigen-specific and antigen-induced, polyclonal responses was observed in the study of doubly immunized individuals. Whereas antibody responses highly specific for the antigen in culture were observed under one set of culture conditions (flat-bottomed vessels, 1.5 x 10(6) cells), switching to another culture system (round-bottomed vessels, 5 x 10(5) cells) resulted in polyclonal responses to antigen. Despite these culture condition-related differences in the induction of antibody synthesis, the suppression of specific antibody production that occurred at high concentrations of antigen was specific only for the antigen in culture. The capability to easily and reproducibly look at truly antigen-induced, antigen specific antibody production should be a major tool in furthering the understanding of human B cell activation and immunoregulation. PMID:6169778

Genetic and antigenic relationship of foot-and-mouth disease virus serotype O isolates with the vaccine strain O1/BFS.

PubMed

Xu, Wanhong; Zhang, Zhidong; Nfon, Charles; Yang, Ming

2018-05-15

Foot-and-mouth disease serotype O viruses (FMDV/O) are responsible for the most outbreaks in FMD endemic countries. O1/BFS is one of the recommended FMD/O vaccine strains by World Reference Laboratory for FMD. In the current study, FMDV/O1 BFS vaccine strain and serotype O field isolates (45) were analyzed phylogenetically and antigenically to gain more insight into the genetic and antigenic characteristics of the vaccine strain and field isolates. O1/BFS showed similarity with 89% of the field isolates using a virus neutralization test (VNT). The P1 region encoding the FMDV capsid was sequenced and analysed for 46 strains of FMDV/O. Phylogenetic analysis showed these viruses originated from five continents and covered eight of 11 reported topotypes. Five isolates that demonstrated low antigenic similarities with O1/BFS were analyzed for their antigenic variation at the known neutralizing antigenic sites. Three of the five isolates demonstrated unique amino acid substitutions at various antigenic sites. No unique amino acid substitutions were observed for the other two unmatched isolates. Positively selected residues were identified on the surface of the FMD virus capsid supporting that it is important to continuously monitor field isolates for their antigenic and phenotypic changes. In conclusion, the vaccine strain O1/BFS is likely to confer protection against 89% of the 45 FMDV/O isolates based on VNT. Thus O1/BFS vaccine strain is still suitable for use in global FMD serotype O outbreak control. Combining data from phylogenetic, molecular and antigenic analysis can provide improvements in the process of vaccine selection. Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.

The spacing between adjacent binding sites in the family of repeats affects the functions of Epstein-Barr nuclear antigen 1 in transcription activation and stable plasmid maintenance.

PubMed

Hebner, Christy; Lasanen, Julie; Battle, Scott; Aiyar, Ashok

2003-07-05

Epstein-Barr virus (EBV) and the closely related Herpesvirus papio (HVP) are stably replicated as episomes in proliferating latently infected cells. Maintenance and partitioning of these viral plasmids requires a viral sequence in cis, termed the family of repeats (FR), that is bound by a viral protein, Epstein-Barr nuclear antigen 1 (EBNA1). Upon binding FR, EBNA1 maintains viral genomes in proliferating cells and activates transcription from viral promoters required for immortalization. FR from either virus encodes multiple binding sites for the viral maintenance protein, EBNA1, with the FR from the prototypic B95-8 strain of EBV containing 20 binding sites, and FR from HVP containing 8 binding sites. In addition to differences in the number of EBNA1-binding sites, adjacent binding sites in the EBV FR are typically separated by 14 base pairs (bp), but are separated by 10 bp in HVP. We tested whether the number of binding sites, as well as the distance between adjacent binding sites, affects the function of EBNA1 in transcription activation or plasmid maintenance. Our results indicate that EBNA1 activates transcription more efficiently when adjacent binding sites are separated by 10 bp, the spacing observed in HVP. In contrast, using two separate assays, we demonstrate that plasmid maintenance is greatly augmented when adjacent EBNA1-binding sites are separated by 14 bp, and therefore, presumably lie on the same face of the DNA double helix. These results provide indication that the functions of EBNA1 in transcription activation and plasmid maintenance are separable.

Streptococci and Actinomyces induce antibodies which cross react with epithelial antigens in periodontitis

PubMed Central

Ye, P; Harty, D W S; Chapple, C C; Nadkarni, M A; Carlo, A A D E; Hunter, N

2003-01-01

Perturbation of epithelial structure is a prominent but poorly understood feature of the immunopathological response to bacterial antigens which characterizes the destructive lesion of periodontitis. Western analysis of sera from 22 patients with periodontitis detected multiple antigens in extracts of epithelial cells whereas sera from 12 periodontally healthy subjects displayed only trace reaction with epithelial antigens. To investigate a possible relationship between the bacterial flora adjacent to diseased sites and the presence of antibodies reactive with epithelium, subgingival plaque samples were taken from deep periodontal pockets and cultured anaerobically. Gram positive bacteria containing antigens cross-reactive with epithelial cells were reproducibly isolated by probing membrane colony-lifts with affinity-isolated (epithelium-specific) antibodies and identified by 16S rDNA sequence homology as streptococci (S. mitis, S. constellatus and two S. intermedius strains) and Actinomyces (A. georgiae, and A. sp. oral clone). Conversely, when serum from patients with periodontitis was absorbed with the captured bacterial species the number of epithelial antigens recognized was specifically reduced. It was concluded that development of cross-reactive antibodies related to these organisms may contribute to perturbation of the epithelial attachment to the tooth and the progression of periodontitis. These autoreactive antibodies could also be a contributing factor in other diseases affecting epithelia. PMID:12605700

High-temperature elastic-plastic and creep properties for SA533 Grade B Class I and SA508 materials

DOE Office of Scientific and Technical Information (OSTI.GOV)

Reddy, G.B.; Ayres, D.J.

1982-12-01

High temperature elastic-plastic and creep properties are presented for SA533 Grade B Class I and SA508 Class II materials. These properties are derived from tests conducted at Combustion Engineering Material and Metallurgical Laboratories and cover the temperature range of 70/sup 0/F to 1200/sup 0/F.

ANTIGENIC MODULATION

PubMed Central

Old, Lloyd J.; Stockert, Elisabeth; Boyse, Edward A.; Kim, Jae Ho

1968-01-01

Antigenic modulation (the loss of TL antigens from TL+ cells exposed to TL antibody in the absence of lytic complement) has been demonstrated in vitro. An ascites leukemia, phenotype TL.1,2,3, which modulates rapidly and completely when incubated with TL antiserum in vitro, was selected for further study of the phenomenon. Over a wide range of TL antibody concentrations modulation at 37°C was detectable within 10 min and was complete within approximately 1 hr. The cells were initially sensitized to C' by their contact with antibody, thereafter losing this sensitivity to C' lysis together with their sensitivity to TL antibody and C' in the cytotoxic test. The capacity of the cells to undergo modulation was abolished by actinomycin D and by iodoacetamide, and by reducing the temperature of incubation to 0°C. Thus modulation apparently is an active cellular process. Antigens TL. 1,2, and 3 are all modulated by anti-TL.1,3 serum and by anti-TL.3 serum. This modulation affects all three TL components together, even when antibody to one or two of them is lacking. aAnti-TL.2 serum does not induce modulation and in fact impairs modulation by the other TL antibodies. The influence of the TL phenotype of cells upon the demonstrable content of H-2 (D region) isoantigen, first shown in cells modulated in vivo, has been observed with cells modulated in vitro. Cells undergoing modulation show a progressive increase in H-2 (D region) antigen over a period of 4 hr, with no change in H-2 antigens of the K region. Restoration of the TL+ phenotype of modulated cells after removal of antibody is less rapid than TL+ → TL- modulation and may require several cell divisions. PMID:5636556

Bacterial colonization patterns in daily chlorhexidine care at the exit site in peritoneal dialysis patients-A prospective, randomized controlled trial.

PubMed

Wang, Hsi-Hao; Hung, Shih-Yuan; Chang, Min-Yu; Lee, Yi-Che; Lin, Hsiu-Fang; Lin, Tsun-Mei; Yang, Su-Pen; Lin, Hsi-Hsun; Yang, Su-Ching; Wang, Jiun-Ling

2017-01-01

Bacterial colonization patterns in daily chlorhexidine care at the exit site in peritoneal dialysis (PD) patients were not known. We performed a prospective, randomized controlled trial enrolling 89 PD patients. After stratification by initial Staphylococcus aureus (SA) carrier status, patients were randomly assigned to receive daily 4% chlorhexidine care (intervention group) or normal saline (control group) at the exit site. Monthly, we cultured bacteria from the exit site and nasal swabs for 1 year. The SA colonization rates at exit site at 6 and 12 months were significantly lower in the intervention group than the control group (5.0% vs. 22.9%, p = 0.023 and 8.6% vs. 28.1%, p = 0.037 for 6 and 12 months, respectively). The Methicillin-resistant SA (MRSA) colonization rate at exit site at 6 months was similar (5.7% vs. 2.5%,p = 0.596) in control and intervention group, but significantly lower in the intervention group than the control group at exit site at 12months (0% vs. 12.5%, p = 0.047). The gram-negative bacilli (GNB) colonization rates were similar between the intervention and control groups at 6 and 12 months. Genotyping of all MRSA isolates showed ST (sequence type) 59 was the most predominant clone. In conclusion, chlorhexidine care at the exit site in PD patients may be a good strategy for SA and MRSA decolonization. ClinicalTrials.gov NCT02446158.

Central Tolerance to Tissue-specific Antigens Mediated by Direct and Indirect Antigen Presentation

PubMed Central

Gallegos, Alena M.; Bevan, Michael J.

2004-01-01

Intrathymic expression of tissue-specific antigens (TSAs) by medullary thymic epithelial cells (Mtecs) leads to deletion of autoreactive T cells. However, because Mtecs are known to be poor antigen-presenting cells (APCs) for tolerance to ubiquitous antigens, and very few Mtecs express a given TSA, it was unclear if central tolerance to TSA was induced directly by Mtec antigen presentation or indirectly by thymic bone marrow (BM)-derived cells via cross-presentation. We show that professional BM-derived APCs acquire TSAs from Mtecs and delete autoreactive CD8 and CD4 T cells. Although direct antigen presentation by Mtecs did not delete the CD4 T cell population tested in this study, Mtec presentation efficiently deleted both monoclonal and polyclonal populations of CD8 T cells. For developing CD8 T cells, deletion by BM-derived APC and by Mtec presentation occurred abruptly at the transitional, CD4high CD8low TCRintermediate stage, presumably as the cells transit from the cortex to the medulla. These studies reveal a cooperative relationship between Mtecs and BM-derived cells in thymic elimination of autoreactive T cells. Although Mtecs synthesize TSAs and delete a subset of autoreactive T cells, BM-derived cells extend the range of clonal deletion by cross-presenting antigen captured from Mtecs. PMID:15492126

Cellular Pathway(S) of Antigen Processing and Presentation in Fish APC: Endosomal Involvement and Cell-Free Antigen Presentation

PubMed Central

Vallejo, Abbe N.; Miller, Norman W.; Harvey, Nancy E.; Cuchens, Marvin A.; Warr, Gregory W.

1992-01-01

Studies were conducted to address further the role(s) of antigen processing and presentation in the induction of immune responses in a phylogenetically lower vertebrate, specifically a teleost, the channel catfish. In particular, studies were aimed at determining the subcellular compartments involved in antigen degradation by channel catfish antigen-presenting cells (APC) as well as ascertaining the reexpression of immunogenic peptides on the surfaces of APC. The results showed that exogenous protein antigens were actively endocytosed by APC as detected by flow cytometry. Use of radiolabeled antigen and subcellular fractionation protocols also showed that antigen localized in endosomes/lysosomes. Furthermore, there was an apparent redistribution of antigen between these organelles and the plasma membrane during the course of antigen pulsing. Functional assays for the induction of in vitro antigen-specific proliferation of immune catfish peripheral blood leukocytes (PBL) showed that membrane preparations from antigen-pulsed autologous APC were highly stimulatory. The magnitude of responses elicited with such membrane preparations was very similar to that of PBL cultures stimulated with native antigen-pulsed and fixed intact APC or prefixed intact APC incubated with a peptide fragment of the nominal antigen. Current data further corroborate our previous findings that steps akin to antigen processing and presentation are clearly important in the induction of immune responses in lower vertebrates like fish, in a manner similar to that seen in mammalian systems. Consequently, it would appear that many immune functions among the diverse taxa of vertebrates are remarkably conserved. PMID:1343103

Mapping Antigenic Motifs in the Trypomastigote Small Surface Antigen from Trypanosoma cruzi

PubMed Central

Balouz, Virginia; Cámara, María de los Milagros; Cánepa, Gaspar E.; Carmona, Santiago J.; Volcovich, Romina; Gonzalez, Nicolás; Altcheh, Jaime; Agüero, Fernán

2015-01-01

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. PMID:25589551

Diversifying Selection Analysis Predicts Antigenic Evolution of 2009 Pandemic H1N1 Influenza A Virus in Humans.

PubMed

Lee, Alexandra J; Das, Suman R; Wang, Wei; Fitzgerald, Theresa; Pickett, Brett E; Aevermann, Brian D; Topham, David J; Falsey, Ann R; Scheuermann, Richard H

2015-05-01

Although a large number of immune epitopes have been identified in the influenza A virus (IAV) hemagglutinin (HA) protein using various experimental systems, it is unclear which are involved in protective immunity to natural infection in humans. We developed a data mining approach analyzing natural H1N1 human isolates to identify HA protein regions that may be targeted by the human immune system and can predict the evolution of IAV. We identified 16 amino acid sites experiencing diversifying selection during the evolution of prepandemic seasonal H1N1 strains and found that 11 sites were located in experimentally determined B-cell/antibody (Ab) epitopes, including three distinct neutralizing Caton epitopes: Sa, Sb, and Ca2 [A. J. Caton, G. G. Brownlee, J. W. Yewdell, and W. Gerhard, Cell 31:417-427, 1982, http://dx.doi.org/10.1016/0092-8674(82)90135-0]. We predicted that these diversified epitope regions would be the targets of mutation as the 2009 H1N1 pandemic (pH1N1) lineage evolves in response to the development of population-level protective immunity in humans. Using a chi-squared goodness-of-fit test, we identified 10 amino acid sites that significantly differed between the pH1N1 isolates and isolates from the recent 2012-2013 and 2013-2014 influenza seasons. Three of these sites were located in the same diversified B-cell/Ab epitope regions as identified in the analysis of prepandemic sequences, including Sa and Sb. As predicted, hemagglutination inhibition (HI) assays using human sera from subjects vaccinated with the initial pH1N1 isolate demonstrated reduced reactivity against 2013-2014 isolates. Taken together, these results suggest that diversifying selection analysis can identify key immune epitopes responsible for protective immunity to influenza virus in humans and thereby predict virus evolution. The WHO estimates that approximately 5 to 10% of adults and 20 to 30% of children in the world are infected by influenza virus each year. While an adaptive

Radioimmunoassays of hidden viral antigens

DOE Office of Scientific and Technical Information (OSTI.GOV)

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

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 Bmore » virus DNA were detected. Antibody-bound adenovirus, herpesvirus, and measles virus antigens were discerned by the procedure.« less

Quantitative analysis of antigen for the induction of tolerance in carcinoembryonic antigen transgenic mice.

PubMed Central

Hasegawa, T; Isobe, K; Nakashima, I; Shimokata, K

1992-01-01

In order to analyse the amounts of antigen in the thymus for the induction of tolerance, several carcinoembryonic antigen (CEA) transgenic lines were established which expressed human CEA antigen with different amounts. The chimeric KSN nude mice transplanted with the thymus of the B601 line (in which CEA mRNA and CEA protein could be detected in various tissues) to kidney capsule showed tolerance to human CEA. On the other hand, the chimeric KSN nude mice transplanted with the thymus of the B602 or BC60 line (in which neither CEA mRNA nor CEA protein could be detected by Northern blot analysis and flow cytometry analysis) or normal C57BL/6 (B6) did not develop the tolerance to human CEA. However, the chimeric KSN nude mice transplanted simultaneously with thymus of the B6 and spleen of the B601 line became tolerant to human CEA antigen. In the case of systemic immunization with cells which had CEA antigen, the B601 line was tolerant to human CEA. Surprisingly, the B602 and BC60 lines were also tolerant to CEA molecule. These results indicate that not only the antigen present in the thymus but also the antigen which flows from the peripheral organs to the thymus may be necessary for the induction of CEA tolerance. Images Figure 1 PMID:1493931

T cell receptor–induced phosphoinositide-3-kinase p110δ activity is required for T cell localization to antigenic tissue in mice

PubMed Central

Jarmin, Sarah J.; David, Rachel; Ma, Liang; Chai, Jan-Guo; Dewchand, Hamlata; Takesono, Aya; Ridley, Anne J.; Okkenhaug, Klaus; Marelli-Berg, Federica M.

2008-01-01

The establishment of T cell–mediated inflammation requires the migration of primed T lymphocytes from the blood stream and their retention in antigenic sites. While naive T lymphocyte recirculation in the lymph and blood is constitutively regulated and occurs in the absence of inflammation, the recruitment of primed T cells to nonlymphoid tissue and their retention at the site are enhanced by various inflammatory signals, including TCR engagement by antigen-displaying endothelium and resident antigen-presenting cells. In this study, we investigated whether signals downstream of TCR ligation mediated by the phosphoinositide-3-kinase (PI3K) subunit p110δ contributed to the regulation of these events. T lymphocytes from mice expressing catalytically inactive p110δ displayed normal constitutive trafficking and migratory responses to nonspecific stimuli. However, these cells lost susceptibility to TCR-induced migration and failed to localize efficiently to antigenic tissue. Importantly, we showed that antigen-induced T cell trafficking and subsequent inflammation was abrogated by selective pharmacological inhibition of PI3K p110δ activity. These observations suggest that pharmacological targeting of p110δ activity is a viable strategy for the therapy of T cell–mediated pathology. PMID:18259608

Characterization and storage of malaria antigens: Localization and chemical characterization of Plasmodium knowlesi schizont antigens

PubMed Central

Deans, J. A.; Cohen, S.

1979-01-01

The identification of malarial antigens that induce protective immunity could provide a rational basis for developing an effective antimalarial vaccine as well as specific serodiagnostic tests indicative of clinical immune status. Since protective immunity is probably induced by stage-dependent rather than stage-independent antigens, the antigenic composition of different stages of Plasmodium knowlesi has been compared, and a limited chemical characterization undertaken. This information should provide some insight into the types of preparative procedure appropriate for the purification of functionally important malarial antigens. PMID:120777

Leukemia-associated antigens in man.

PubMed

Brown, G; Capellaro, D; Greaves, M

1975-12-01

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

Structural flexibility of a conserved antigenic region in hepatitis C virus glycoprotein E2 recognized by broadly neutralizing antibodies.

PubMed

Meola, Annalisa; Tarr, Alexander W; England, Patrick; Meredith, Luke W; McClure, C Patrick; Foung, Steven K H; McKeating, Jane A; Ball, Jonathan K; Rey, Felix A; Krey, Thomas

2015-02-01

Neutralizing antibodies (NAbs) targeting glycoprotein E2 are important for the control of hepatitis C virus (HCV) infection. One conserved antigenic site (amino acids 412 to 423) is disordered in the reported E2 structure, but a synthetic peptide mimicking this site forms a β-hairpin in complex with three independent NAbs. Our structure of the same peptide in complex with NAb 3/11 demonstrates a strikingly different extended conformation. We also show that residues 412 to 423 are essential for virus entry but not for E2 folding. Together with the neutralizing capacity of the 3/11 Fab fragment, this indicates an unexpected structural flexibility within this epitope. NAbs 3/11 and AP33 (recognizing the extended and β-hairpin conformations, respectively) display similar neutralizing activities despite converse binding kinetics. Our results suggest that HCV utilizes conformational flexibility as an immune evasion strategy, contributing to the limited immunogenicity of this epitope in patients, similar to the conformational flexibility described for other enveloped and nonenveloped viruses. Approximately 180 million people worldwide are infected with hepatitis C virus (HCV), and neutralizing antibodies play an important role in controlling the replication of this major human pathogen. We show here that one of the most conserved antigenic sites within the major glycoprotein E2 (amino acids 412 to 423), which is disordered in the recently reported crystal structure of an E2 core fragment, can adopt different conformations in the context of the infectious virus particle. Recombinant Fab fragments recognizing different conformations of this antigenic site have similar neutralization activities in spite of converse kinetic binding parameters. Of note, an antibody response targeting this antigenic region is less frequent than those targeting other more immunogenic regions in E2. Our results suggest that the observed conformational flexibility in this conserved antigenic

T-cell antigenic sites involved in myasthenia gravis: correlations with antibody titre and disease severity.

PubMed

Berrih-Aknin, S; Cohen-Kaminsky, S; Lepage, V; Neumann, D; Bach, J F; Fuchs, S

1991-02-01

We have evaluated the ability of eight synthetic peptides corresponding to selected regions of the alpha-subunit from human (H) or Torpedo (T) acetylcholine receptor (AChR) to stimulate proliferative responses of peripheral blood lymphocytes (PBL) and thymic cells from patients with Myasthenia Gravis (MG) in comparison to healthy controls. Using PBL, two of the peptides were most reactive: in the 40 myasthenic patients tested, peptide 169-181 (H) induced significant proliferative responses in 10 patients and peptide 351-368 (H) in five, while there was no response in any of the 34 healthy controls tested. Interestingly, clear associations between proliferation to peptides and clinical data were observed. Indeed, among responding patients, all presented thymic hyperplasia and most showed a high anti-AChR Ab titre and/or a severe form of the disease. In addition, responses to AChR cytoplasmic sequences were observed only in severely affected patients. Correlation with HLA-DR haplotype, sought in a subgroup of patients, indicated that response to 169-181 (H) is associated with HLA-DR5 in the patients presenting a high anti-AChR antibody titre. Using thymic lymphocytes, few responses were obtained with the human peptides, suggesting that the frequency of autoreactive cells is lower than in the blood. Similar to PBL, responses to peptides were observed only with lymphocytes isolated from hyperplastic thymuses. The correlations observed between responses to peptides and clinical parameters underline the pathophysiological relevance of our data and indicate that pathogenic and nonpathogenic T-cell antigenic sites involved in the anti-AChR response could be identified by this approach.

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

PubMed

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.

R4SA for Controlling Robots

NASA Technical Reports Server (NTRS)

Aghazarian, Hrand

2009-01-01

The R4SA GUI mentioned in the immediately preceding article is a userfriendly interface for controlling one or more robot(s). This GUI makes it possible to perform meaningful real-time field experiments and research in robotics at an unmatched level of fidelity, within minutes of setup. It provides such powerful graphing modes as that of a digitizing oscilloscope that displays up to 250 variables at rates between 1 and 200 Hz. This GUI can be configured as multiple intuitive interfaces for acquisition of data, command, and control to enable rapid testing of subsystems or an entire robot system while simultaneously performing analysis of data. The R4SA software establishes an intuitive component-based design environment that can be easily reconfigured for any robotic platform by creating or editing setup configuration files. The R4SA GUI enables event-driven and conditional sequencing similar to those of Mars Exploration Rover (MER) operations. It has been certified as part of the MER ground support equipment and, therefore, is allowed to be utilized in conjunction with MER flight hardware. The R4SA GUI could also be adapted to use in embedded computing systems, other than that of the MER, for commanding and real-time analysis of data.

Antigenic variation of the human influenza A (H3N2) virus during the 2014-2015 winter season.

PubMed

Hua, Sha; Li, XiYan; Liu, Mi; Cheng, YanHui; Peng, YouSong; Huang, WeiJuan; Tan, MinJu; Wei, HeJiang; Guo, JunFeng; Wang, DaYan; Wu, AiPing; Shu, YueLong; Jiang, TaiJiao

2015-09-01

The human influenza A (H3N2) virus dominated the 2014-2015 winter season in many countries and caused massive morbidity and mortality because of its antigenic variation. So far, very little is known about the antigenic patterns of the recent H3N2 virus. By systematically mapping the antigenic relationships of H3N2 strains isolated since 2010, we discovered that two groups with obvious antigenic divergence, named SW13 (A/Switzerland/9715293/2013-like strains) and HK14 (A/Hong Kong/5738/2014-like strains), co-circulated during the 2014-2015 winter season. HK14 group co-circulated with SW13 in Europe and the United States during this season, while there were few strains of HK14 in mainland China, where SW13 has dominated since 2012. Furthermore, we found that substitutions near the receptor-binding site on hemagglutinin played an important role in the antigenic variation of both the groups. These findings provide a comprehensive understanding of the recent antigenic evolution of H3N2 virus and will aid in the selection of vaccine strains.

Oral Vaccination of Fish – Antigen Preparations, Uptake, and Immune Induction

PubMed Central

Mutoloki, Stephen; Munang’andu, Hetron Mweemba; Evensen, Øystein

2015-01-01

The oral route offers the most attractive approach of immunization of fish for a number of reasons: the ease of administration of antigens, it is less stressful than parenteral delivery and in principle, it is applicable to small and large sized fish; it also provides a procedure for oral boosting during grow-out periods in cages or ponds. There are, however, not many commercial vaccines available at the moment due to lack of efficacy and challenges associated with production of large quantities of antigens. These are required to stimulate an effective immune response locally and systemically, and need to be protected against degradation before they reach the sites where immune induction occurs. The hostile stomach environment is believed to be particularly important with regard to degradation of antigens in certain species. There is also a poor understanding about the requirements for proper immune induction following oral administration on one side, and the potential for induction of tolerance on the other. To what extent primary immunization via the oral route will elicit both local and systemic responses is not understood in detail. Furthermore, to what extent parenteral delivery will protect mucosal/gut surfaces and vice-versa is also not fully understood. We review the work that has been done on the subject and discuss it in light of recent advances that include mass production of antigens, including the use of plant systems. Different encapsulation techniques that have been developed in the quest to protect antigens against digestive degradation, as well as to target them for appropriate immune induction are also highlighted. PMID:26539192

Marked differences in the antigenic structure of human respiratory syncytial virus F and G glycoproteins.

PubMed Central

García-Barreno, B; Palomo, C; Peñas, C; Delgado, T; Perez-Breña, P; Melero, J A

1989-01-01

Monoclonal antibodies directed against the glycoproteins of human respiratory syncytial virus were used in competitive enzyme-linked immunosorbent assays for topological mapping of epitopes. Whereas epitopes of the F glycoprotein could be ascribed to five nonoverlapping antigenic sites, anti-G antibodies recognized unique epitopes, many of whose competition profiles overlapped extensively. Variant viruses selected with a neutralizing (47F) anti-F antibody lost the binding for only 47F and 49F antibodies, which mapped in the same antigenic area. In contrast, viruses selected with an anti-G antibody lost the capacity to bind most of the anti-G antibodies, and their G protein was not recognized by an anti-virus antiserum, indicating major changes in the antigenic structure of the G molecule. Finally, we found great antigenic variation of the G protein among viral isolates. This occurred even within viruses of the same subtype with only limited divergence of amino acid sequence between strains. All of these data indicate marked differences in the antigenic organization of the G and F glycoproteins of respiratory syncytial virus; we discuss these differences in terms of the chemical structure of the glycoproteins. Images PMID:2463385

Induced Resistance to Ixodid Tick Infestation: Analysis and Isolation of Antigens

DTIC Science & Technology

1988-01-01

female Ixodes ricinus. This resistance could be in- hibited by daily treatment with mepyramine, a type-1 histamine receptor antagonist. Bagnall (1975...attempted to mimic naturally acquired resistance . This approach caused no reaction at attachment sites. Immunization with midgut antigens resulted in the...particulate and soluble components prepared from midgut induced resistance . This agrees with immunization results using 27,000 x g supernatant

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

PubMed

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

2014-12-01

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

Closed-loop control of SaO2 in the neonate.

PubMed

Morozoff, P E; Evans, R W

1992-01-01

A microprocessor-based device has been designed to control oxygen saturation (SaO2) in neonates by adjusting the inspired air-oxygen mixture (FiO2) delivered by a mechanical blender. The user sets a target SaO2, which the controller attempts to maintain. Alarms are actuated if the neonate's SaO2 is outside predefined limits. SaO2 levels are extracted from a commercial pulse oximeter and analyzed by an eight-bit microprocessing unit (MPU). Delivered percentages of FiO2 are adjusted by a motorized air-oxygen blender. The controller has a menu-driven user interface and can graphically present four-hour trends of the SaO2, FiO2, or blender setting. Sixteen hours of collected data can be stored and later downloaded to a personal computer. A real-time multitasking operating system forms the nucleus of the controller's software. Major tasks that share MPU time are control, filtering, user display, data collection, data archiving, alarm monitoring, and user input. Analog SaO2 levels are read and converted to digital values, which are then filtered to extract noise. A differential control algorithm is used to determine the required FiO2 blender setting. The blender is then adjusted to the new setting, after which the controller waits to repeat the process of sampling SaO2 and adjusting FiO2. System response time and blender increments are adjustable to allow a user to tune the controller to the patient's needs. Alarm conditions of concern within the device are SaO2 and FiO2 sensor disconnection, blender disconnection, and SaO2 limiting errors. In preliminary trials, for a target of 92.0% SaO2, a prototype controller maintained an average of 91.6% with a standard deviation of 5.0% over a one-hour period.(ABSTRACT TRUNCATED AT 250 WORDS)

Do FY antigens act as minor histocompatibility antigens in the graft-versus-host disease paradigm after human leukocyte antigen-identical sibling hematopoietic stem cell transplantation?

PubMed

Sellami, Mohamed Hichem; Chaabane, Manel; Kaabi, Houda; Torjemane, Lamia; Ladeb, Saloua; Ben Othmane, Tarek; Hmida, Slama

2012-03-01

FY antigens are candidate minor histocompatibility antigens relevant to renal allograft rejection, but no data have been reported about their role in graft-versus-host disease (GVHD) incidence after human leukocyte antigen (HLA)-identical siblings hematopoietic stem cell transplantation (HSCT). The aim of this study was to examine the effect of donor/recipient disparity at FY antigens on the incidence of GVHD in Tunisian patients receiving an HLA-identical HSCT. This work enrolled 105 Tunisian pairs of recipients and their HLA-identical sibling donors of HSCs. FY genotyping was performed with the polymerase chain reaction-sequence-specific primer method and donor/recipient disparity for these antigens was analyzed at two levels: incompatibility and nonidentity. The case-control analyses showed no significant correlation between FY disparity and the incidence of either acute or chronic GVHD. Sample size calculation showed that 572 cases and 1716 controls would be necessary to be able to detect a significant association with 80% power and two-sided type I error level of 5% (α=0.05). The lack of association in the studied cohort may be explained by the low immunogenicity of FY antigens in HSCT context, compared with other antigens such as HA-1 and CD31.

The new technological solution for the JT-60SA quench protection circuits

NASA Astrophysics Data System (ADS)

Gaio, E.; Maistrello, A.; Novello, L.; Matsukawa, M.; Perna, M.; Ferro, A.; Yamauchi, K.; Piovan, R.

2018-07-01

An advanced technology has been developed and employed for the main circuit breakers (CB) of the quench protection circuits (QPC) of the superconducting coils of JT-60SA: it consists in a Hybrid mechanical-static CB (HCB) composed of a mechanical Bypass switch (BPS) for conducting the continuous current, in parallel to a static circuit breaker (SCB) based on integrated gate commutated thyristor (IGCT) for current interruption. It was the result of a R&D program carried out since 2006 to identify innovative solutions for the interruption of high dc current, able to improve the maintainability and availability of the CB. The HCB developed for the JT-60SA QPC is the first realization of a dc circuit breaker based on this design approach for interrupting current of some tens of kA with reapplied voltage of some kV. It also represents the first application of hybrid technology with IGCT for protection of superconducting magnets in fusion experiments. The paper aims at giving a comprehensive overview of the main R&D activities devoted to the development of this new technological approach; then, the key aspects of the design, manufacturing and testing of the QPCs for JT-60SA, successfully completed in Naka Site in summer 2015 are presented. Finally, the significance of this research is discussed and the possible future developments, in particular in view of DEMO fusion reactor, are outlined.

Structural insights into substrate and inhibitor binding sites in human indoleamine 2,3-dioxygenase 1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lewis-Ballester, Ariel; Pham, Khoa N.; Batabyal, Dipanwita

Human indoleamine 2,3-dioxygenase 1 (hIDO1) is an attractive cancer immunotherapeutic target owing to its role in promoting tumoral immune escape. However, drug development has been hindered by limited structural information. Here, we report the crystal structures of hIDO1 in complex with its substrate, Trp, an inhibitor, epacadostat, and/or an effector, indole ethanol (IDE). The data reveal structural features of the active site (Sa) critical for substrate activation; in addition, they disclose a new inhibitor-binding mode and a distinct small molecule binding site (Si). Structure-guided mutation of a critical residue, F270, to glycine perturbs the Si site, allowing structural determination ofmore » an inhibitory complex, where both the Sa and Si sites are occupied by Trp. The Si site offers a novel target site for allosteric inhibitors and a molecular explanation for the previously baffling substrate-inhibition behavior of the enzyme. Taken together, the data open exciting new avenues for structure-based drug design.« less

Interconversion of the Specificities of Human Lysosomal Enzymes Associated with Fabry and Schindler Diseases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tomasic, Ivan B.; Metcalf, Matthew C.; Guce, Abigail I.

2010-09-03

The human lysosomal enzymes {alpha}-galactosidase ({alpha}-GAL, EC 3.2.1.22) and {alpha}-N-acetylgalactosaminidase ({alpha}-NAGAL, EC 3.2.1.49) share 46% amino acid sequence identity and have similar folds. The active sites of the two enzymes share 11 of 13 amino acids, differing only where they interact with the 2-position of the substrates. Using a rational protein engineering approach, we interconverted the enzymatic specificity of {alpha}-GAL and {alpha}-NAGAL. The engineered {alpha}-GAL (which we call {alpha}-GALSA) retains the antigenicity of {alpha}-GAL but has acquired the enzymatic specificity of {alpha}-NAGAL. Conversely, the engineered {alpha}-NAGAL (which we call {alpha}-NAGAL{sup EL}) retains the antigenicity of {alpha}-NAGAL but has acquired themore » enzymatic specificity of the {alpha}-GAL enzyme. Comparison of the crystal structures of the designed enzyme {alpha}-GAL{sup SA} to the wild-type enzymes shows that active sites of {alpha}-GAL{sup SA} and {alpha}-NAGAL superimpose well, indicating success of the rational design. The designed enzymes might be useful as non-immunogenic alternatives in enzyme replacement therapy for treatment of lysosomal storage disorders such as Fabry disease.« less

Antigenic characterization of bovine ephemeral fever rhabdovirus G and GNS glycoproteins expressed from recombinant baculoviruses.

PubMed

Johal, Jasjit; Gresty, Karryn; Kongsuwan, Kritaya; Walker, Peter J

2008-01-01

Recombinant baculoviruses expressing the BEFV envelope glycoprotein G and non-structural glycoprotein G(NS) were constructed. The G protein expressed in insect cells was located on the cell surface and induced spontaneous cell fusion at mildly acidic pH. The expressed G protein reacted with MAbs to continuous and conformational neutralization sites (G1, G2, G3b and G4), but not to conformational site G3a. The expressed G(NS) protein was also located on the cell surface but did not exhibit fusogenic activity. The G(NS) protein reacted with polyclonal antiserum produced from vaccinia-virus-expressed recombinant G(NS) but did not react with G protein antibodies. A His(6)-tagged, soluble form of the G protein was expressed and purified by Ni(2+)-NTA chromatography. The purified G protein reacted with BEFV-neutralizing MAbs to all continuous and conformational antigenic sites. The highly protective characteristics of the native BEFV G protein suggest that the secreted, baculovirus-expressed product may be a useful vaccine antigen.

Antigen recognition in the islets changes with progression of autoimmune islet infiltration

PubMed Central

Lindsay, Robin S.; Corbin, Kaitlin; Mahne, Ashley; Levitt, Bonnie E.; Gebert, Matthew J.; Wigton, Eric J.; Bradley, Brenda J.; Haskins, Kathryn; Jacobelli, Jordan; Tang, Qizhi; Krummel, Matthew F.; Friedman, Rachel S.

2014-01-01

In type 1 diabetes, the pancreatic islets are an important site for therapeutic intervention since immune infiltration of the islets is well established at diagnosis. Therefore, understanding the events that underlie the continued progression of the autoimmune response and islet destruction is critical. Islet infiltration and destruction is an asynchronous process, making it important to analyze the disease process on a single islet basis. To understand how T cell stimulation evolves through the process of islet infiltration we analyzed the dynamics of T cell movement and interactions within individual islets of spontaneously autoimmune non-obese diabetic (NOD) mice. Using both intra-vital and explanted 2-photon islet imaging, we defined a correlation between increased islet infiltration and increased T cell motility. Early T cell arrest was antigen dependent and due, at least in part, to antigen recognition through sustained interactions with CD11c+ antigen presenting cells (APCs). As islet infiltration progressed, T cell motility became antigen-independent, with a loss of T cell arrest and sustained interactions with CD11c+ APCs. These studies suggest that the autoimmune T cell response in the islets may be temporarily dampened during the course of islet infiltration and disease progression. PMID:25505281

Sole existence of antithrombin antibody in patients with systemic lupus erythematosus showing tendency of its antigenic determinants directing against exosite II (antithrombin/heparin binding site) of thrombin.

PubMed

Matsuda, Juzo; Matsuyama, Atsushi; Atsumi, Gen; Ohkura, Naoki

2008-01-01

We conducted an investigation to determine the antigenic determinants of antithrombin antibody (aThr), which has recently been recognized as a new antiphospholipid antibody mostly co-existing with antiprothrombin antibody, employing patients with systemic lupus erythematosus and antiphospholipid syndrome. Using an enzyme-linked immunosorbent assay we found aThr in 34 of 83 patients (40.9%), and 27 of these 34 patients (79.4%) with aThr were all negative for other antiphospholipid antibodies. An optical density value of six of 30 patients (20.0%) with aThr showed more than a 40% reduction of reactivity to thrombin with the addition of antithrombin in a dose-dependent manner. The inhibition percentage of aThr to thrombin was prominently increased to 11 of 30 (37%) along with its inhibition rate (100% at the highest) by the co-existence of heparin. Seven out of 30 patients with aThr (23.3%) showed a reduction of the optical density value with the addition of hirudin. Our findings suggest that aThr exists solely in patients with systemic lupus erythematosus without other antiphospholipid antibodies, and the antigenic determinants of aThr are directed to exosite I (hirudin binding site) and exosite II (antithrombin/heparin binding site) on the thrombin surface, with exosite II predominance. Accordingly, aThr could be an isolated and additional new marker of thrombosis/hemostasis. Since our patients who were positive only for aThr do not have a past history of antiphospholipid-associated complications at this stage, however, further long-term follow-up and additional studies in these clinical settings are needed to verify our hypothesis in the future.

The world after SA : benefits to GPS integrity.

DOT National Transportation Integrated Search

2000-03-01

The Presidential Decision Directive (PDD) on the Global Positioning System (GPS) recommends that selective availability (SA) be removed by 2006. The question remains: if SA were to be turned off, how significant are the benefits to the GPS community?...

Isolation and Genome Characterization of the Virulent Staphylococcus aureus Bacteriophage SA97

PubMed Central

Chang, Yoonjee; Shin, Hakdong; Lee, Ju-Hoon; Park, Chul Jong; Paik, Soon-Young; Ryu, Sangryeol

2015-01-01

A novel bacteriophage that infects S. aureus, SA97, was isolated and characterized. The phage SA97 belongs to the Siphoviridae family, and the cell wall teichoic acid (WTA) was found to be a host receptor of the phage SA97. Genome analysis revealed that SA97 contains 40,592 bp of DNA encoding 54 predicted open reading frames (ORFs), and none of these genes were related to virulence or drug resistance. Although a few genes associated with lysogen formation were detected in the phage SA97 genome, the phage SA97 produced neither lysogen nor transductant in S. aureus. These results suggest that the phage SA97 may be a promising candidate for controlling S. aureus. PMID:26437428

A small antigenic determinant of the Chikungunya virus E2 protein is sufficient to induce neutralizing antibodies which are partially protective in mice.

PubMed

Weber, Christopher; Büchner, Sarah M; Schnierle, Barbara S

2015-04-01

The mosquito-borne Chikungunya virus (CHIKV) causes high fever and severe joint pain in humans. It is expected to spread in the future to Europe and has recently reached the USA due to globalization, climate change and vector switch. Despite this, little is known about the virus life cycle and, so far, there is no specific treatment or vaccination against Chikungunya infections. We aimed here to identify small antigenic determinants of the CHIKV E2 protein able to induce neutralizing immune responses. E2 enables attachment of the virus to target cells and a humoral immune response against E2 should protect from CHIKV infections. Seven recombinant proteins derived from E2 and consisting of linear and/or structural antigens were created, and were expressed in and purified from E. coli. BALB/c mice were vaccinated with these recombinant proteins and the mouse sera were screened for neutralizing antibodies. Whereas a linear N-terminally exposed peptide (L) and surface-exposed parts of the E2 domain A (sA) alone did not induce neutralizing antibodies, a construct containing domain B and a part of the β-ribbon (called B+) was sufficient to induce neutralizing antibodies. Furthermore, domain sA fused to B+ (sAB+) induced the highest amount of neutralizing antibodies. Therefore, the construct sAB+ was used to generate a recombinant modified vaccinia virus Ankara (MVA), MVA-CHIKV-sAB+. Mice were vaccinated with MVA-CHIKV-sAB+ and/or the recombinant protein sAB+ and were subsequently challenged with wild-type CHIKV. Whereas four vaccinations with MVA-CHIKV-sAB+ were not sufficient to protect mice from a CHIKV infection, protein vaccination with sAB+ markedly reduced the viral titers of vaccinated mice. The recombinant protein sAB+ contains important structural antigens for a neutralizing antibody response in mice and its formulation with appropriate adjuvants might lead to a future CHIKV vaccine.

VIEW LOOKING EAST, SA WETSIDE (DISTILLATION BUILDING) ON THE RIGHT, ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

VIEW LOOKING EAST, SA WETSIDE (DISTILLATION BUILDING) ON THE RIGHT, STD (SODA TOWER DRYERS?), SA DRYSIDE ON RIGHT. BEHIND STD BUILDING IS SHD BUILDING (SODA HORIZONTAL DRYERS?) THE ENTIRE DRYING COMPLEX WAS KNOWN AS THE DRYSIDE. - Solvay Process Company, SA Wetside Building, Between Willis & Milton Avenue, Solvay, Onondaga County, NY

Integrating influenza antigenic dynamics with molecular evolution

PubMed Central

Bedford, Trevor; Suchard, Marc A; Lemey, Philippe; Dudas, Gytis; Gregory, Victoria; Hay, Alan J; McCauley, John W; Russell, Colin A; Smith, Derek J; Rambaut, Andrew

2014-01-01

Influenza viruses undergo continual antigenic evolution allowing mutant viruses to evade host immunity acquired to previous virus strains. Antigenic phenotype is often assessed through pairwise measurement of cross-reactivity between influenza strains using the hemagglutination inhibition (HI) assay. Here, we extend previous approaches to antigenic cartography, and simultaneously characterize antigenic and genetic evolution by modeling the diffusion of antigenic phenotype over a shared virus phylogeny. Using HI data from influenza lineages A/H3N2, A/H1N1, B/Victoria and B/Yamagata, we determine patterns of antigenic drift across viral lineages, showing that A/H3N2 evolves faster and in a more punctuated fashion than other influenza lineages. We also show that year-to-year antigenic drift appears to drive incidence patterns within each influenza lineage. This work makes possible substantial future advances in investigating the dynamics of influenza and other antigenically-variable pathogens by providing a model that intimately combines molecular and antigenic evolution. DOI: http://dx.doi.org/10.7554/eLife.01914.001 PMID:24497547

Prostate-Specific Membrane Antigen-Targeted Site-Directed Antibody-Conjugated Apoferritin Nanovehicle Favorably Influences In Vivo Side Effects of Doxorubicin.

PubMed

Dostalova, Simona; Polanska, Hana; Svobodova, Marketa; Balvan, Jan; Krystofova, Olga; Haddad, Yazan; Krizkova, Sona; Masarik, Michal; Eckschlager, Tomas; Stiborova, Marie; Heger, Zbynek; Adam, Vojtech

2018-06-11

Herein, we describe the in vivo effects of doxorubicin (DOX) encapsulated in ubiquitous protein apoferritin (APO) and its efficiency and safety in anti-tumor treatment. APODOX is both passively (through Enhanced Permeability and Retention effect) and actively targeted to tumors through prostate-specific membrane antigen (PSMA) via mouse antibodies conjugated to the surface of horse spleen APO. To achieve site-directed conjugation of the antibodies, a HWRGWVC heptapeptide linker was used. The prostate cancer-targeted and non-targeted nanocarriers were tested using subcutaneously implanted LNCaP cells in athymic mice models, and compared to free DOX. Prostate cancer-targeted APODOX retained the high potency of DOX in attenuation of tumors (with 55% decrease in tumor volume after 3 weeks of treatment). DOX and non-targeted APODOX treatment caused damage to liver, kidney and heart tissues. In contrast, no elevation in liver or kidney enzymes and negligible changes were revealed by histological assessment in prostate cancer-targeted APODOX-treated mice. Overall, we show that the APO nanocarrier provides an easy encapsulation protocol, reliable targeting, high therapeutic efficiency and very low off-target toxicity, and is thus a promising delivery system for translation into clinical use.

Chemotherapy Enhances Cross-Presentation of Nuclear Tumor Antigens

PubMed Central

Anyaegbu, Chidozie C.; Lake, Richard A.; Heel, Kathy; Robinson, Bruce W.; Fisher, Scott A.

2014-01-01

Cross-presentation of tumor antigen is essential for efficient priming of naïve CD8+ T lymphocytes and induction of effective anti-tumor immunity. We hypothesized that the subcellular location of a tumor antigen could affect the efficiency of cross-presentation, and hence the outcome of anti-tumor responses to that antigen. We compared cross-presentation of a nominal antigen expressed in the nuclear, secretory, or cytoplasmic compartments of B16 melanoma tumors. All tumors expressed similar levels of the antigen. The antigen was cross-presented from all compartments but when the concentration was low, nuclear antigen was less efficiently cross-presented than antigen from other cellular locations. The efficiency of cross-presentation of the nuclear antigen was improved following chemotherapy-induced tumor cell apoptosis and this correlated with an increase in the proportion of effector CTL. These data demonstrate that chemotherapy improves nuclear tumor antigen cross-presentation and could be important for anti-cancer immunotherapies that target nuclear antigens. PMID:25243472

Immunotoxicity of gallium arsenide on antigen presentation: comparative study of intratracheal and intraperitoneal exposure routes.

PubMed

Hartmann, Constance B; Harrison, M Travis; McCoy, Kathleen L

2005-01-01

Gallium arsenide (GaAs) is a semiconductor utilized in electronics and computer industries. GaAs exposure of animals causes local inflammation and systemic immune suppression. Mice were administered 2 to 200 mg/kg GaAs. On day 5, intratracheal instillation increased lung weights in a dose-dependent manner and induced pulmonary inflammation exemplified by mononuclear cell infiltration and mild epithelial hyperplasia. No fibrosis, pneumocyte hyperplasia, proteinosis, or bronchial epithelial damage was observed in the lungs. Splenic cellularity and composition were unaffected. GaAs' effect on antigen presentation by macrophages was similar after intratracheal and intraperitoneal exposure, although the lowest observable adverse effect levels differed. Macrophages from the exposure site displayed an enhanced ability to activate an antigen-specific CD4(+) helper T-cell hybridoma compared with vehicle controls, whereas splenic macrophages were defective in this function. The chemical's impact on peritoneal macrophages depended on the exposure route. GaAs exposure augmented thiol cathepsins B and L activities in macrophages from the exposure site, but decreased proteolytic activities in splenic macrophages. Alveolar macrophages had increased expression of major histocompatibility complex (MHC) Class II molecules, whereas MHC Class II expression on splenic and peritoneal macrophages was unaffected. Modified thiol cathepsin activities statistically correlated with altered efficiency of antigen presentation, whereas MHC Class II expression did not. Our study is the first one to examine the functional capability of alveolar macrophages after intratracheal GaAs instillation. Therefore, thiol cathepsins may be potential target molecules by which GaAs exposure modulates antigen presentation.

Graphene oxide@gold nanorods-based multiple-assisted electrochemiluminescence signal amplification strategy for sensitive detection of prostate specific antigen.

PubMed

Cao, Jun-Tao; Yang, Jiu-Jun; Zhao, Li-Zhen; Wang, Yu-Ling; Wang, Hui; Liu, Yan-Ming; Ma, Shu-Hui

2018-01-15

A novel and competitive electrochemiluminescence (ECL) aptasensor for prostate specific antigen (PSA) assay was constructed using gold nanorods functionalized graphene oxide (GO@AuNRs) multilabeled with glucose oxidase (GOD) and streptavidin (SA) toward luminol-based ECL system. A strong initial ECL signal was achieved by electrodeposited gold (DpAu) on the electrode because of gold nanoparticles (AuNPs) motivating the luminol ECL signal. The signal probes prepared by loading GOD and SA-biotin-DNA on GO@AuNRs were used for achieving multiple signal amplification. In the absence of PSA, the signal probes can be attached on the electrode by hybridization reaction between PSA aptamer and biotin-DNA. In this state, the GOD loaded on the probe could catalyze glucose to in situ produce H 2 O 2 and then AuNRs catalyze H 2 O 2 to generate abundant reactive oxygen species (ROSs) in luminol ECL reaction. Both the high-content GOD and AuNRs in the signal probe amplified the ECL signal in the ECL system. Moreover, the combination of SA with biotin-DNA further expands ECL intensity. The integration of such amplifying effects in this protocol endows the aptasensor with high sensitivity and good selectivity for PSA detection. This aptasensor exhibits a linear relation in the range of 0.5pgmL -1 to 5.0ngmL -1 with the detection limit of 0.17pgmL -1 (S/N = 3). Besides, the strategy was successfully applied in determination of human serum samples with recovery of 81.4-116.0%. Copyright © 2017 Elsevier B.V. All rights reserved.

Genotypic evolution and antigenicity of H9N2 influenza viruses in Shanghai, China.

PubMed

Ge, Feifei; Li, Xin; Ju, Houbin; Yang, Dequan; Liu, Jian; Qi, Xinyong; Wang, Jian; Yang, Xianchao; Qiu, Yafeng; Liu, Peihong; Zhou, Jinping

2016-06-01

H9N2 influenza viruses have been circulating in China since 1994, but a systematic investigation of H9N2 in Shanghai has not previously been undertaken. Here, using 14 viruses we isolated from poultry and pigs in Shanghai during 2002 and 2006-2014, together with the commercial vaccine A/chicken/Shanghai/F/1998 (Ck/SH/F/98), we analyzed the evolution of H9N2 influenza viruses in Shanghai and showed that all 14 isolates originated from Ck/SH/F/98 antigenically. We evaluated the immune protection efficiency of the vaccine. Our findings demonstrate that H9N2 viruses in Shanghai have undergone extensive reassortment. Various genotypes emerged in 2002, 2006 and 2007, while during 2009-2014 only one genotype was found. Four antigenic groups, A-D, could be identified among the 14 isolates and a variety of antigenically distinct H9N2-virus-derived avian influenza viruses (AIVs) circulated simultaneously in Shanghai during this period. Challenge experiments using vaccinated chickens indicated that the vaccine prevented shedding of antigenic group A and B viruses, but not those of the more recent groups C and D. Genetic analysis showed that compared to the vaccine strain, representative viruses of antigenic groups C and D possess greater numbers of amino acid substitutions in the hemagglutinin (HA) protein than viruses in antigenic groups A and B. Many of these substitutions are located in antigenic sites. Our results indicate that the persistence of H9N2 AIV in China might be due to incomplete vaccine protection and that the avian influenza vaccine should be regularly evaluated and updated to maintain optimal protection.

Concepts and applications for influenza antigenic cartography

PubMed Central

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

2011-01-01

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

The molecular relationship between antigenic domains and epitopes on hCG.

PubMed

Berger, Peter; Lapthorn, Adrian J

2016-08-01

Antigenic domains are defined to contain a limited number of neighboring epitopes recognized by antibodies (Abs) but their molecular relationship remains rather elusive. We thoroughly analyzed the antigenic surface of the important pregnancy and tumor marker human chorionic gonadotropin (hCG), a cystine knot (ck) growth factor, and set antigenic domains and epitopes in molecular relationships to each other. Antigenic domains on hCG, its free hCGα and hCGβ subunits are dependent on appropriate inherent molecular features such as molecular accessibility and protrusion indices that determine bulging structures accessible to Abs. The banana-shaped intact hCG comprises ∼7500Å(2) of antigenic surface with minimally five antigenic domains that encompass a continuum of overlapping non-linear composite epitopes, not taking into account the C-terminal peptide extension of hCGβ (hCGβCTP). Epitopes within an antigenic domain are defined by specific Abs, that bury nearly 1000Å(2) of surface accessible area on the antigen and recognize a few up to 15 amino acid (aa) residues, whereby between 2 and 5 of these provide the essential binding energy. Variability in Ab binding modes to the contact aa residues are responsible for the variation in affinity and intra- and inter-species specificity, e.g. cross-reactions with luteinizing hormone (LH). Each genetically distinct fragment antigen binding (Fab) defines its own epitope. Consequently, recognition of the same epitope by different Abs is only possible in cases of genetically identical sequences of its binding sites. Due to combinatorial V(D)J gene segment variability of heavy and light chains, Abs defining numerous epitopes within an antigenic domain can be generated by different individuals and species. Far more than hundred Abs against the immuno-dominant antigenic domains of either subunit at both ends of the hCG-molecule, the tips of peptide loops one and three (Ł1+3) protruding from the central ck, encompassing h

Detection of specific IgE antibodies to major and minor antigenic determinants in sera of penicillin allergic patients.

PubMed

Zhao, Yongxing; Qiao, Hailing

2003-12-01

To investigate the mechanism(s) of penicillins allergic reaction. The radioallergosorbent test (RAST) was used to detect 9 specific IgE antibodies, including major antigenic determinants: benzylpenicilloyl (BPO), ampicilloyl (APO), amoxicilloyl (AXO), phenoxomethylpenicilloyl (PVO) and flucloxacilloyl (FLUO), and minor antigenic determinants: benzylpenicillanyl (BPA), amoxicillanyl (AXA), 6-aminopenicillanic (APA) and phenoxomethylpenicillany (PVA), in the sera of 32 penicillin allergic patients. The relationship between specific IgE antibodies and penicillins chemical structures was studied by radioallergosorbent inhibition test. Nineteen of 32 patients (59.4%) were RAST positive, among whom, five cases were positive only to one or two antigenic minor determinants, and three cases were positive only to one or three major antigenic determinants. The remaining 11 patients were positive not only to major antigenic determinants but also minor antigenic determinants. In 9 specific IgE antibodies, the positive rate of PVA-IgE was the highest (34.38%), followed by BPO-IgE (31.25%). The positive rate of FLUO-IgE was the lowest (15.63%). Of the total patient group, 53.13% were positive to one or more minor antigenic determinants, while 37.5% (12/32) were positive to one or more major antigenic determinants. The percentage of patients with urticarial reactions who were positive to minor antigenic determinants (63.16%) was significantly higher than observed in the anaphylactic shock group (38.5%, P < 0.05). The minor antigenic determinant was important in allergic reaction. The combining sites of the specific IgE antibodies were likely to be the side-chain of drug or the overwhelming drug molecule.

Localization of sequential antigenic determinants on bovine beta-casein with synthetic peptides and antisera from mouse, rabbit, and goat.

PubMed

Mizumachi, K; Kurisaki, J; Kaminogawa, S

1999-05-01

The antigenic determinants of bovine beta-casein (beta-CN) were localized by using twenty overlapping peptides encompassing the entire sequence of beta-CN and anti-beta-CN antisera from outbred mouse, rabbit and goat. The profile of the reactions was characteristic to the species, the dominant antigenic regions being 80-95, 143-158 and 195-209 in mouse, 1-16 in rabbit and 100-115 in goat. Regions 1-16, 100-115, 121-136 and 143-158 were antigenic in all three species. The number of antigenic regions recognized by goat was much fewer than that by mouse and rabbit, possibly because of the homology between bovine and goat beta-CN. A mixture of the twenty peptides could absorb about 50-60% of beta-CN specific antibodies from each species. Furthermore, the mouse and rabbit anti-beta-CN antibodies were also specific to the phosphorylated regions. We therefore conclude that the major antigenic determinants on beta-CN would be largely sequential and include the phosphorylated sites.

Prediction of site-specific interactions in antibody-antigen complexes: the proABC method and server.

PubMed

Olimpieri, Pier Paolo; Chailyan, Anna; Tramontano, Anna; Marcatili, Paolo

2013-09-15

Antibodies or immunoglobulins are proteins of paramount importance in the immune system. They are extremely relevant as diagnostic, biotechnological and therapeutic tools. Their modular structure makes it easy to re-engineer them for specific purposes. Short of undergoing a trial and error process, these experiments, as well as others, need to rely on an understanding of the specific determinants of the antibody binding mode. In this article, we present a method to identify, on the basis of the antibody sequence alone, which residues of an antibody directly interact with its cognate antigen. The method, based on the random forest automatic learning techniques, reaches a recall and specificity as high as 80% and is implemented as a free and easy-to-use server, named prediction of Antibody Contacts. We believe that it can be of great help in re-design experiments as well as a guide for molecular docking experiments. The results that we obtained also allowed us to dissect which features of the antibody sequence contribute most to the involvement of specific residues in binding to the antigen. http://www.biocomputing.it/proABC. anna.tramontano@uniroma1.it or paolo.marcatili@gmail.com Supplementary data are available at Bioinformatics online.

Heparin-associated thrombocytopenia: antibody binding specificity to platelet antigens.

PubMed

Lynch, D M; Howe, S E

1985-11-01

Sera from four patients with heparin-associated thrombocytopenia (HAT) were evaluated by a quantitative enzyme-linked immunosorbent assay (ELISA) to detect heparin-dependent serum platelet-bindable immunoglobulin (S-PBIg) and by Western blotting and immunoprecipitation to investigate the specificity of the antibody binding. All HAT sera showed mildly increased S-PBIg (mean, 7.8 fg per platelet; normal, less than 6.0 fg per platelet) to intact target platelets in the ELISA, which was markedly increased in the presence of heparin (mean, 20.9 fg per platelet). This increase was 20-fold greater than normal control sera, which showed a mean differential increase of only 0.5 fg per platelet. Immunoglobulin binding specificity to platelet antigens was investigated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis of platelet lysate with transfer of the platelet fractions onto nitrocellulose strips (Western blotting) and subsequent immunoassay using HAT and normal sera. In the presence of heparin, the four HAT patients demonstrated increased binding of immunoglobulin to platelet antigens of apparent molecular weights of 180, 124, and 82 kd. Radiolabeled heparin when incubated with HAT sera, normal sera, or albumin blanks bound to platelet proteins of the same apparent molecular weights. These observations are consistent with current hypotheses suggesting that HAT antibody is directed to heparin-platelet complexes or, alternatively, that heparin induces conformational change of antigenic sites on the platelet membrane.

Enhanced expression of interferon-gamma-induced antigen-processing machinery components in a spontaneously occurring cancer.

PubMed

Cerruti, Fulvia; Martano, Marina; Petterino, Claudio; Bollo, Enrico; Morello, Emanuela; Bruno, Renato; Buracco, Paolo; Cascio, Paolo

2007-11-01

In human tumors, changes in the surface expression and/or function of major histocompatibility complex (MHC) class I antigens are frequently found and may provide malignant cells with a mechanism to escape control of the immune system. This altered human lymphocyte antigen (HLA) class I phenotype can be caused by either structural alterations or dysregulation of genes encoding subunits of HLA class I antigens and/or components of the MHC class I antigen-processing machinery (APM). Herein we analyze the expression of several proteins involved in the generation of MHC class I epitopes in feline injection site sarcoma, a spontaneously occurring tumor in cats that is an informative model for the study of tumor biology in other species, including humans. Eighteen surgically removed primary fibrosarcoma lesions were analyzed, and an enhanced expression of two catalytic subunits of immunoproteasomes, PA28 and leucine aminopeptidase, was found in tumors compared to matched normal tissues. As a functional counterpart of these changes in protein levels, proteasomal activities were increased in tissue extracts from fibrosarcomas. Taken together, these results suggest that alterations in the APM system may account for reduced processing of selected tumor antigens and may potentially provide neoplastic fibroblasts with a mechanism for escape from T-cell recognition and destruction.

Substitutions near the hemagglutinin receptor-binding site determine the antigenic evolution of influenza A H3N2 viruses in U.S. swine.

PubMed

Lewis, Nicola S; Anderson, Tavis K; Kitikoon, Pravina; Skepner, Eugene; Burke, David F; Vincent, Amy L

2014-05-01

Swine influenza A virus is an endemic and economically important pathogen in pigs, with the potential to infect other host species. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major component in swine influenza A vaccines. However, as a result of antigenic drift, vaccine strains must be regularly updated to reflect currently circulating strains. Characterizing the cross-reactivity between strains in pigs and seasonal influenza virus strains in humans is also important in assessing the relative risk of interspecies transmission of viruses from one host population to the other. Hemagglutination inhibition (HI) assay data for swine and human H3N2 viruses were used with antigenic cartography to quantify the antigenic differences among H3N2 viruses isolated from pigs in the United States from 1998 to 2013 and the relative cross-reactivity between these viruses and current human seasonal influenza A virus strains. Two primary antigenic clusters were found circulating in the pig population, but with enough diversity within and between the clusters to suggest updates in vaccine strains are needed. We identified single amino acid substitutions that are likely responsible for antigenic differences between the two primary antigenic clusters and between each antigenic cluster and outliers. The antigenic distance between current seasonal influenza virus H3 strains in humans and those endemic in swine suggests that population immunity may not prevent the introduction of human viruses into pigs, and possibly vice versa, reinforcing the need to monitor and prepare for potential incursions. Influenza A virus (IAV) is an important pathogen in pigs and humans. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major target of vaccines. However, vaccine strains must be updated to reflect current strains. Characterizing the differences between seasonal IAV in humans and swine IAV is important in

Substitutions near the Hemagglutinin Receptor-Binding Site Determine the Antigenic Evolution of Influenza A H3N2 Viruses in U.S. Swine

PubMed Central

Lewis, Nicola S.; Anderson, Tavis K.; Kitikoon, Pravina; Skepner, Eugene; Burke, David F.

2014-01-01

ABSTRACT Swine influenza A virus is an endemic and economically important pathogen in pigs, with the potential to infect other host species. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major component in swine influenza A vaccines. However, as a result of antigenic drift, vaccine strains must be regularly updated to reflect currently circulating strains. Characterizing the cross-reactivity between strains in pigs and seasonal influenza virus strains in humans is also important in assessing the relative risk of interspecies transmission of viruses from one host population to the other. Hemagglutination inhibition (HI) assay data for swine and human H3N2 viruses were used with antigenic cartography to quantify the antigenic differences among H3N2 viruses isolated from pigs in the United States from 1998 to 2013 and the relative cross-reactivity between these viruses and current human seasonal influenza A virus strains. Two primary antigenic clusters were found circulating in the pig population, but with enough diversity within and between the clusters to suggest updates in vaccine strains are needed. We identified single amino acid substitutions that are likely responsible for antigenic differences between the two primary antigenic clusters and between each antigenic cluster and outliers. The antigenic distance between current seasonal influenza virus H3 strains in humans and those endemic in swine suggests that population immunity may not prevent the introduction of human viruses into pigs, and possibly vice versa, reinforcing the need to monitor and prepare for potential incursions. IMPORTANCE Influenza A virus (IAV) is an important pathogen in pigs and humans. The hemagglutinin (HA) protein is the primary target of protective immune responses and the major target of vaccines. However, vaccine strains must be updated to reflect current strains. Characterizing the differences between seasonal IAV in humans and swine

Intellectual Property and Strategic Agreements (IP/SA) | FNLCR Staging

Cancer.gov

IP/SA handles all invention issues including patents and copyrights. All employee inventionreports are filed through the IP/SA office for all activities under the OTS contract.Additionally,request for assignment ofcopyri

Structure-based non-canonical amino acid design to covalently crosslink an antibody–antigen complex

PubMed Central

Xu, Jianqing; Tack, Drew; Hughes, Randall A.; Ellington, Andrew D.; Gray, Jeffrey J.

2014-01-01

Engineering antibodies to utilize non-canonical amino acids (NCAA) should greatly expand the utility of an already important biological reagent. In particular, introducing crosslinking reagents into antibody complementarity determining regions (CDRs) should provide a means to covalently crosslink residues at the antibody–antigen interface. Unfortunately, finding the optimum position for crosslinking two proteins is often a matter of iterative guessing, even when the interface is known in atomic detail. Computer-aided antibody design can potentially greatly restrict the number of variants that must be explored in order to identify successful crosslinking sites. We have therefore used Rosetta to guide the introduction of an oxidizable crosslinking NCAA, l-3,4-dihydroxyphenylalanine (l-DOPA), into the CDRs of the anti-protective antigen scFv antibody M18, and have measured crosslinking to its cognate antigen, domain 4 of the anthrax protective antigen. Computed crosslinking distance, solvent accessibility, and interface energetics were three factors considered that could impact the efficiency of l-DOPA-mediated crosslinking. In the end, 10 variants were synthesized, and crosslinking efficiencies were generally 10% or higher, with the best variant crosslinking to 52% of the available antigen. The results suggest that computational analysis can be used in a pipeline for engineering crosslinking antibodies. The rules learned from l-DOPA crosslinking of antibodies may also be generalizable to the formation of other crosslinked interfaces and complexes. PMID:23680795

Selection of tumor antigens as targets for immune attack using immunohistochemistry: protein antigens.

PubMed

Zhang, S; Zhang, H S; Cordon-Cardo, C; Ragupathi, G; Livingston, P O

1998-11-01

The relative expression of mucin antigens MUC1, MUC2, MUC3, MUC4, MUC5AC, MUC5B, and MUC7 and glycoprotein antigens KSA, carcinoembryonic antigen, prostate-specific membrane antigen (PSMA), HER-2/neu, and human chorionic gonadotropin-beta on different cancers and normal tissues is difficult to determine from available reports. We have compared the distribution of these antigens by immunohistology on a broad range of malignant and normal tissues. MUC1 expression was most intense in cancers of breast, lung, ovarian, and endometrial origin; MUC2 was most intense in cancers of colon and prostate origin; and MUC5AC was most intense in cancers of breast and gastric origin. MUC4 was intensely expressed in 50% of cancers of colon and pancreas origin, and MUC3, MUC5B, and MUC7 were expressed in a variety of epithelial cancers, but not so intensely. KSA was intensely and uniformly expressed on all epithelial cancers; carcinoembryonic antigen was expressed in most cancers of breast, lung, colon, pancreas, and gastric origin; and PSMA was expressed only in cancers of prostate origin. Human chorionic gonadotropin-beta was expressed on the majority of sarcomas and cancers of breast, lung, and pancreas origin, although intense staining was not seen. Staining on normal tissues was restricted to one or many normal epithelial tissues ranging from MUC3, MUC4, and PSMA, which were expressed only on epithelia of pancreas, stomach, and prostate origin, respectively, to MUC1 and KSA, which were expressed on most normal epithelia. Expression was restricted to the secretory borders of these epithelia while stroma and other normal tissues were completely negative. These results plus the results of the two previous papers (S. Zhang et al, Int. J. Cancer, 73: 42-49, 1997; S. Zhang et al., Int. J. Cancer, 73: 50-56, 1997) in this series provide the basis for selection of multiple cell surface antigens as targets for antibody-mediated attack against these cancers.

Antigen Cross-Priming of Cell-Associated Proteins is Enhanced by Macroautophagy within the Antigen Donor Cell

PubMed Central

Joubert, Pierre-Emmanuel; Albert, Matthew L.

2012-01-01

Phagocytosis of dying cells constitutes an important mechanism of antigen capture for the cross-priming of CD8+ T cells. This process has been shown to be critical for achieving tumor and viral immunity. While most studies have focused on the mechanisms inherent in the dendritic cell that account for exogenous antigen accessing MHC I, several recent reports have highlighted the important contribution made by the antigen donor cell. Specifically, the cell stress and cell death pathways that precede antigen transfer are now known to impact cross-presentation and cross-priming. Herein, we review the current literature regarding a role for macroautophagy within the antigen donor cell. Further examination of this point of immune regulation is warranted and may contribute to a better understanding of how to optimize immunotherapy for treatment of cancer and chronic infectious disease. PMID:22566942

76 FR 13063 - Airworthiness Directives; EUROCOPTER FRANCE Model SA330F, SA330G, and SA330J Helicopters

Federal Register 2010, 2011, 2012, 2013, 2014

2011-03-10

... beyond the end limit (``tall pilot'' position). This resulted in the separation of the pedal adjustment... the ``tall pilot'' stop nut was damaged, most likely due to aging of the adhesive. The nut came loose... the pedal unit on a SA 330 helicopter, the copilot set the position beyond the end limit (``tall pilot...

Mutational landscape of antibody variable domains reveals a switch modulating the interdomain conformational dynamics and antigen binding

PubMed Central

Koenig, Patrick; Lee, Chingwei V.; Walters, Benjamin T.; Janakiraman, Vasantharajan; Stinson, Jeremy; Patapoff, Thomas W.; Fuh, Germaine

2017-01-01

Somatic mutations within the antibody variable domains are critical to the immense capacity of the immune repertoire. Here, via a deep mutational scan, we dissect how mutations at all positions of the variable domains of a high-affinity anti-VEGF antibody G6.31 impact its antigen-binding function. The resulting mutational landscape demonstrates that large portions of antibody variable domain positions are open to mutation, and that beneficial mutations can be found throughout the variable domains. We determine the role of one antigen-distal light chain position 83, demonstrating that mutation at this site optimizes both antigen affinity and thermostability by modulating the interdomain conformational dynamics of the antigen-binding fragment. Furthermore, by analyzing a large number of human antibody sequences and structures, we demonstrate that somatic mutations occur frequently at position 83, with corresponding domain conformations observed for G6.31. Therefore, the modulation of interdomain dynamics represents an important mechanism during antibody maturation in vivo. PMID:28057863

Antigenic Distance Measurements for Seasonal Influenza Vaccine Selection

PubMed Central

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

2011-01-01

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. PMID:22063385

Antigen-specific T cell therapies for cancer

PubMed Central

Manzo, Teresa; Heslop, Helen E.; Rooney, Cliona M.

2015-01-01

Adoptively transferred antigen-specific T cells that recognize tumor antigens through their native receptors have many potential benefits as treatment for virus-associated diseases and malignancies, due to their ability to selectively recognize tumor antigens, expand and persist to provide long-term protection. Infusions of T cells targeting Epstein–Barr virus (EBV) antigens have shown encouraging response rates in patients with post-transplant lymphoproliferative disease as well as EBV-positive lymphomas and nasopharyngeal cancer, although a recent study also showed that human papilloma virus-reactive T cells can induce complete regression of metastatic cervical cancer. This strategy is also being evaluated to target non-viral tumor-associated antigens. Targeting these less immunogenic antigens is more challenging, as tumor antigens are generally weak, and high avidity T cells specific for self-antigens are deleted in the thymus, but tumor responses have been reported. Current research focusses on defining factors that promote in vivo persistence of transferred cells and ameliorate the immunosuppressive microenvironment. To this end, investigators are evaluating the effects of combining adoptive transfer of antigen-specific T cells with other immunotherapy moieties such as checkpoint inhibitors. Genetic modification of infused T cells may also be used to overcome tumor evasion mechanisms, and vaccines may be used to promote in vivo proliferation. PMID:26160910

Enumeration of antigen-specific CD8+ T lymphocytes by single-platform, HLA tetramer-based flow cytometry: a European multicenter evaluation.

PubMed

Heijnen, Ingmar A F M; Barnett, David; Arroz, Maria J; Barry, Simon M; Bonneville, Marc; Brando, Bruno; D'hautcourt, Jean-Luc; Kern, Florian; Tötterman, Thomas H; Marijt, Erik W A; Bossy, David; Preijers, Frank W M B; Rothe, Gregor; Gratama, Jan W

2004-11-01

HLA class I peptide tetramers represent powerful diagnostic tools for detection and monitoring of antigen-specific CD8(+) T cells. The impetus for the current multicenter study is the critical need to standardize tetramer flow cytometry if it is to be implemented as a routine diagnostic assay. Hence, the European Working Group on Clinical Cell Analysis set out to develop and evaluate a single-platform tetramer-based method that used cytomegalovirus (CMV) as the antigenic model. Absolute numbers of CMV-specific CD8(+) T cells were obtained by combining the percentage of tetramer-binding cells with the absolute CD8(+) T-cell count. Six send-outs of stabilized blood from healthy individuals or CMV-carrying donors with CMV-specific CD8(+) T-cell counts of 3 to 10 cells/microl were distributed to 7 to 16 clinical sites. These sites were requested to enumerate CD8(+) T cells and, in the case of CMV-positive donors, CMV-specific subsets on three separate occasions using the standard method. Between-site coefficients of variation of less than 10% (absolute CD8(+) T-cell counts) and approximately 30% (percentage and absolute numbers of CMV-specific CD8(+) T cells) were achieved. Within-site coefficients of variation were approximately 5% (absolute CD8(+) T-cell counts), approximately 9% (percentage CMV-specific CD8(+) T cells), and approximately 17% (absolute CMV-specific CD8(+) T-cell counts). The degree of variation tended to correlate inversely with the proportion of CMV-specific CD8(+) T-cell subsets. The single-platform MHC tetramer-based method for antigen-specific CD8(+) T-cell counting has been evaluated by a European group of laboratories and can be considered a reproducible assay for routine enumeration of antigen-specific CD8(+) T cells. (c) 2004 Wiley-Liss, Inc.

Presentation of lipid antigens to T cells.

PubMed

Mori, Lucia; De Libero, Gennaro

2008-04-15

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

ABH antigens as recognition sites for the activation of red blood cell anion exchange by the lectin ulex europaeus agglutinin I.

PubMed

Engelmann, B

1993-11-01

The blood group antigen H (blood group O) and fucose-specific lectin Ulex europaeus agglutinin I (UEA1) (10 micrograms/ml) was found to increase the rate constant of Cl- efflux into 100 mM Na+ oxalate media by about 40% in erythrocytes taken from antigen H donors. In 100 mM K+ oxalate, 150 mM Na+ pyruvate and in 150 mM Na+ acetate media the lectin elevated the rate constant of Cl- efflux by 20-50%. The acceleration of Cl- efflux by UEA1 was completely blocked by 10 microM 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS) indicating that the effect of the lectin is mediated by the anion exchanger of human erythrocytes (band 3 protein). In antigen A1 erythrocytes no significant stimulation of anion exchange by UEA1 was seen. The activation of Cl- efflux was completely prevented by addition of 1 mM fucose to the medium. These results suggest that the effect of UEA1 is mediated through interaction with the fucose residues of H antigens. Increasing extracellular Ca++ from 0.5 to 5 mM in Na+ pyruvate or Na+ acetate media slightly reduced the acceleration of anion exchange by the lectin. On the other hand, replacing part of extracellular chloride by bicarbonate did not considerably alter the (previously reported) stimulatory effect of UEA1 on red blood cell Ca++ uptake. This suggests that the acceleration of anion exchange and of Ca++ uptake by UEA1, respectively, are mediated by different mechanisms. It is concluded that UEA1 activates anion exchange of human erythrocytes most probably by a direct interaction with H antigens present on extracellular domains of the band 3 protein.

Encryption of agonistic motifs for TLR4 into artificial antigens augmented the maturation of antigen-presenting cells.

PubMed

Ito, Masaki; Hayashi, Kazumi; Minamisawa, Tamiko; Homma, Sadamu; Koido, Shigeo; Shiba, Kiyotaka

2017-01-01

Adjuvants are indispensable for achieving a sufficient immune response from vaccinations. From a functional viewpoint, adjuvants are classified into two categories: "physical adjuvants" increase the efficacy of antigen presentation by antigen-presenting cells (APC) and "signal adjuvants" induce the maturation of APC. Our previous study has demonstrated that a physical adjuvant can be encrypted into proteinous antigens by creating artificial proteins from combinatorial assemblages of epitope peptides and those peptide sequences having propensities to form certain protein structures (motif programming). However, the artificial antigens still require a signal adjuvant to maturate the APC; for example, co-administration of the Toll-like receptor 4 (TLR4) agonist monophosphoryl lipid A (MPLA) was required to induce an in vivo immunoreaction. In this study, we further modified the previous artificial antigens by appending the peptide motifs, which have been reported to have agonistic activity for TLR4, to create "adjuvant-free" antigens. The created antigens with triple TLR4 agonistic motifs in their C-terminus have activated NF-κB signaling pathways through TLR4. These proteins also induced the production of the inflammatory cytokine TNF-α, and the expression of the co-stimulatory molecule CD40 in APC, supporting the maturation of APC in vitro. Unexpectedly, these signal adjuvant-encrypted proteins have lost their ability to be physical adjuvants because they did not induce cytotoxic T lymphocytes (CTL) in vivo, while the parental proteins induced CTL. These results confirmed that the manifestation of a motif's function is context-dependent and simple addition does not always work for motif-programing. Further optimization of the molecular context of the TLR4 agonistic motifs in antigens should be required to create adjuvant-free antigens.

Characterization of O-antigen delivered by Generalized Modules for Membrane Antigens (GMMA) vaccine candidates against nontyphoidal Salmonella.

PubMed

De Benedetto, G; Alfini, R; Cescutti, P; Caboni, M; Lanzilao, L; Necchi, F; Saul, A; MacLennan, C A; Rondini, S; Micoli, F

2017-01-11

Invasive nontyphoidal Salmonella disease (iNTS) is a leading cause of death and morbidity in Africa. The most common pathogens are Salmonella enterica serovars Typhimurium and Enteritidis. The O-antigen portion of their lipopolysaccharide is a target of protective immunity and vaccines targeting O-antigen are currently in development. Here we investigate the use of Generalized Modules for Membrane Antigens (GMMA) as delivery system for S. Typhimurium and S. Enteritidis O-antigen. Gram-negative bacteria naturally shed outer membrane in a blebbing process. By deletion of the tolR gene, the level of shedding was greatly enhanced. Further genetic modifications were introduced into the GMMA-producing strains in order to reduce reactogenicity, by detoxifying the lipid A moiety of lipopolysaccharide. We found that genetic mutations can impact on expression of O-antigen chains. All S. Enteritidis GMMA characterized had an O-antigen to protein w/w ratio higher than 0.6, while the ratio was 0.7 for S. Typhimurium ΔtolR GMMA, but decreased to less than 0.1 when further mutations for lipid A detoxification were introduced. Changes were also observed in O-antigen chain length and level and/or position of O-acetylation. When tested in mice, the GMMA induced high levels of anti-O-antigen-specific IgG functional antibodies, despite variation in density and O-antigen structural modifications. In conclusion, simplicity of manufacturing process and low costs of production, coupled with encouraging immunogenicity data, make GMMA an attractive strategy to further investigate for the development of a vaccine against iNTS. Copyright © 2016. Published by Elsevier Ltd.

Structural Basis For Antigenic Peptide Precursor Processing by the Endoplasmic Reticulum Aminopeptidase ERAP1

DOE Office of Scientific and Technical Information (OSTI.GOV)

T Nguyen; S Chang; I Evnouchidou

2011-12-31

ERAP1 trims antigen precursors to fit into MHC class I proteins. To fulfill this function, ERAP1 has unique substrate preferences, trimming long peptides but sparing shorter ones. To identify the structural basis for ERAP1's unusual properties, we determined the X-ray crystal structure of human ERAP1 bound to bestatin. The structure reveals an open conformation with a large interior compartment. An extended groove originating from the enzyme's catalytic center can accommodate long peptides and has features that explain ERAP1's broad specificity for antigenic peptide precursors. Structural and biochemical analyses suggest a mechanism for ERAP1's length-dependent trimming activity, whereby binding of longmore » rather than short substrates induces a conformational change with reorientation of a key catalytic residue toward the active site. ERAP1's unique structural elements suggest how a generic aminopeptidase structure has been adapted for the specialized function of trimming antigenic precursors.« less

Determination of O:4 antigen-antibody affinity level in O:5 antigen positive and negative variants of Salmonella enterica serovar Typhimurium.

PubMed

Nakai, Yuka; Ito, Akihisa; Ogawa, Yohsuke; Aribam, Swarmistha Devi; Elsheimer-Matulova, Marta; Shiraiwa, Kazumasa; Kisaka, Stevens M B; Hikono, Hirokazu; Nishikawa, Sayaka; Akiba, Masato; Kawahara, Kazuyoshi; Shimoji, Yoshihiro; Eguchi, Masahiro

2017-04-01

Salmonella enterica serovar Typhimurium (S. Typhimurium) has two serological variants: one that expresses the O:5 antigen (1,4,5,12:i:1,2) and one that lacks O:5 antigen (1,4,12:i:1,2). For serotyping, S. Typhimurium is agglutinated by diagnostic O:4 antigen serum. This study was carried out to compare the antigen-antibody affinity of O:4 antigen in S. Typhimurium χ3306 O:5-positive and S. Typhimurium χ3306 O:5-negative strains. The affinity of O:4 antigen with O:4 antigen serum was found to be stronger in the O:5-negative strains compared to O:5-positive strains. Next, we investigated the antigen-antibody affinity of O:4 antigen with O:4 antigen serum in field strains of S. Typhimurium, which showed the same tendency in affinity as seen with S. Typhimurium χ3306 O:5-positive and negative strains. This study suggests that the presence or absence of O:5 antigen causes differences in O:4 agglutination reactions with different field strains of S. Typhimurium. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

9 CFR 113.407 - Pullorum antigen.

Code of Federal Regulations, 2011 CFR

2011-01-01

... shall be free from extraneous organisms as determined by Gram staining and microscopic examination. (b... standard for stained antigen K's and 50 ±10 times McFarland No. 1 standard for tube antigen. (c) Preservative requirements. (1) The formalin content of Pullorum Stained Antigen K shall be 1.0 ±0.2 percent as...

9 CFR 113.407 - Pullorum antigen.

Code of Federal Regulations, 2013 CFR

2013-01-01

... shall be free from extraneous organisms as determined by Gram staining and microscopic examination. (b... standard for stained antigen K's and 50 ±10 times McFarland No. 1 standard for tube antigen. (c) Preservative requirements. (1) The formalin content of Pullorum Stained Antigen K shall be 1.0 ±0.2 percent as...

9 CFR 113.407 - Pullorum antigen.

Code of Federal Regulations, 2010 CFR

2010-01-01

... shall be free from extraneous organisms as determined by Gram staining and microscopic examination. (b... standard for stained antigen K's and 50 ±10 times McFarland No. 1 standard for tube antigen. (c) Preservative requirements. (1) The formalin content of Pullorum Stained Antigen K shall be 1.0 ±0.2 percent as...

9 CFR 113.407 - Pullorum antigen.

Code of Federal Regulations, 2014 CFR

2014-01-01

... shall be free from extraneous organisms as determined by Gram staining and microscopic examination. (b... standard for stained antigen K's and 50 ±10 times McFarland No. 1 standard for tube antigen. (c) Preservative requirements. (1) The formalin content of Pullorum Stained Antigen K shall be 1.0 ±0.2 percent as...

9 CFR 113.407 - Pullorum antigen.

Code of Federal Regulations, 2012 CFR

2012-01-01

... shall be free from extraneous organisms as determined by Gram staining and microscopic examination. (b... standard for stained antigen K's and 50 ±10 times McFarland No. 1 standard for tube antigen. (c) Preservative requirements. (1) The formalin content of Pullorum Stained Antigen K shall be 1.0 ±0.2 percent as...

Siting and sizing of distributed generators based on improved simulated annealing particle swarm optimization.

PubMed

Su, Hongsheng

2017-12-18

Distributed power grids generally contain multiple diverse types of distributed generators (DGs). Traditional particle swarm optimization (PSO) and simulated annealing PSO (SA-PSO) algorithms have some deficiencies in site selection and capacity determination of DGs, such as slow convergence speed and easily falling into local trap. In this paper, an improved SA-PSO (ISA-PSO) algorithm is proposed by introducing crossover and mutation operators of genetic algorithm (GA) into SA-PSO, so that the capabilities of the algorithm are well embodied in global searching and local exploration. In addition, diverse types of DGs are made equivalent to four types of nodes in flow calculation by the backward or forward sweep method, and reactive power sharing principles and allocation theory are applied to determine initial reactive power value and execute subsequent correction, thus providing the algorithm a better start to speed up the convergence. Finally, a mathematical model of the minimum economic cost is established for the siting and sizing of DGs under the location and capacity uncertainties of each single DG. Its objective function considers investment and operation cost of DGs, grid loss cost, annual purchase electricity cost, and environmental pollution cost, and the constraints include power flow, bus voltage, conductor current, and DG capacity. Through applications in an IEEE33-node distributed system, it is found that the proposed method can achieve desirable economic efficiency and safer voltage level relative to traditional PSO and SA-PSO algorithms, and is a more effective planning method for the siting and sizing of DGs in distributed power grids.

Retardation of Antigen Release from DNA Hydrogel Using Cholesterol-Modified DNA for Increased Antigen-Specific Immune Response.

PubMed

Umeki, Yuka; Saito, Masaaki; Takahashi, Yuki; Takakura, Yoshinobu; Nishikawa, Makiya

2017-10-01

Our previous study indicates that cationization of an antigen is effective for sustained release of both immunostimulatory DNA containing unmethylated cytosine-phosphate-guanine (CpG) dinucleotides, or CpG DNA, and antigen from a DNA hydrogel. Another approach to sustained antigen release would increase the applicability and versatility of the system. In this study, a hydrophobic interaction-based sustained release system of ovalbumin (OVA), a model antigen, from immunostimulatory CpG DNA hydrogel is developed by the use of cholesterol-modified DNA and urea-denatured OVA (udOVA). Cholesterol-modified DNA forms a hydrogel, Dgel(chol), and induces IL-6 mRNA expression in mouse skin after intradermal injection, as DNA without cholesterol does. Cholesterol-modified DNA associated with OVA and denaturation of OVA using urea increases the interaction. The release of udOVA from Dgel(chol) is significantly slower than that from DNA hydrogel with no cholesterol, Dgel. Moreover, intratumoral injections of udOVA/Dgel(chol) significantly inhibit the growth of EG7-OVA tumors in mice. These results indicate that sustained release of antigen from Dgel can be achieved by the combination of urea denaturation and cholesterol modification, and retardation of antigen release is effective to induce antigen-specific cancer immunity. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Realtime mitigation of GPS SA errors using Loran-C

NASA Technical Reports Server (NTRS)

Braasch, Soo Y.

1994-01-01

The hybrid use of Loran-C with the Global Positioning System (GPS) was shown capable of providing a sole-means of enroute air radionavigation. By allowing pilots to fly direct to their destinations, use of this system is resulting in significant time savings and therefore fuel savings as well. However, a major error source limiting the accuracy of GPS is the intentional degradation of the GPS signal known as Selective Availability (SA). SA-induced position errors are highly correlated and far exceed all other error sources (horizontal position error: 100 meters, 95 percent). Realtime mitigation of SA errors from the position solution is highly desirable. How that can be achieved is discussed. The stability of Loran-C signals is exploited to reduce SA errors. The theory behind this technique is discussed and results using bench and flight data are given.

Antigenic and genetic comparison of foot-and-mouth disease virus serotype O Indian vaccine strain, O/IND/R2/75 against currently circulating viruses.

PubMed

Mahapatra, Mana; Yuvaraj, S; Madhanmohan, M; Subramaniam, S; Pattnaik, B; Paton, D J; Srinivasan, V A; Parida, Satya

2015-01-29

Foot-and-mouth disease (FMD) virus serotype O is the most common cause of FMD outbreaks in India and three of the six lineages that have been described are most frequently detected, namely Ind2001, PanAsia and PanAsia 2. We report the full capsid sequence of 21 serotype O viruses isolated from India between 2002 and 2012. All these viruses belong to the Middle East-South Asia (ME-SA) topotype. The serological cross-reactivity of a bovine post-vaccination serum pool raised against the current Indian vaccine strain, O/IND/R2/75,was tested by virus neutralisation test with the 23 Indian field isolates, revealing a good match between the vaccine and the field isolates. The cross reactivity of the O/IND/R2/75 vaccine with 19 field isolates from other countries (mainly from Asia and Africa) revealed a good match to 79% of the viruses indicating that the vaccine strain is broadly cross-reactive and could be used to control FMD in other countries. Comparison of the capsid sequences of the serologically non-matching isolates with the vaccine strain sequence identified substitutions in neutralising antigenic sites 1 and 2, which could explain the observed serological differences. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Antigenic and genetic comparison of foot-and-mouth disease virus serotype O Indian vaccine strain, O/IND/R2/75 against currently circulating viruses

PubMed Central

Mahapatra, Mana; Yuvaraj, S.; Madhanmohan, M.; Subramaniam, S.; Pattnaik, B.; Paton, D.J.; Srinivasan, V.A.; Parida, Satya

2015-01-01

Foot-and-mouth disease (FMD) virus serotype O is the most common cause of FMD outbreaks in India and three of the six lineages that have been described are most frequently detected, namely Ind2001, PanAsia and PanAsia 2. We report the full capsid sequence of 21 serotype O viruses isolated from India between 2002 and 2012. All these viruses belong to the Middle East–South Asia (ME–SA) topotype. The serological cross-reactivity of a bovine post-vaccination serum pool raised against the current Indian vaccine strain, O/IND/R2/75,was tested by virus neutralisation test with the 23 Indian field isolates, revealing a good match between the vaccine and the field isolates. The cross reactivity of the O/IND/R2/75 vaccine with 19 field isolates from other countries (mainly from Asia and Africa) revealed a good match to 79% of the viruses indicating that the vaccine strain is broadly cross-reactive and could be used to control FMD in other countries. Comparison of the capsid sequences of the serologically non-matching isolates with the vaccine strain sequence identified substitutions in neutralising antigenic sites 1 and 2, which could explain the observed serological differences. PMID:25500306

Encryption of agonistic motifs for TLR4 into artificial antigens augmented the maturation of antigen-presenting cells

PubMed Central

Hayashi, Kazumi; Minamisawa, Tamiko; Homma, Sadamu; Koido, Shigeo; Shiba, Kiyotaka

2017-01-01

Adjuvants are indispensable for achieving a sufficient immune response from vaccinations. From a functional viewpoint, adjuvants are classified into two categories: “physical adjuvants” increase the efficacy of antigen presentation by antigen-presenting cells (APC) and “signal adjuvants” induce the maturation of APC. Our previous study has demonstrated that a physical adjuvant can be encrypted into proteinous antigens by creating artificial proteins from combinatorial assemblages of epitope peptides and those peptide sequences having propensities to form certain protein structures (motif programming). However, the artificial antigens still require a signal adjuvant to maturate the APC; for example, co-administration of the Toll-like receptor 4 (TLR4) agonist monophosphoryl lipid A (MPLA) was required to induce an in vivo immunoreaction. In this study, we further modified the previous artificial antigens by appending the peptide motifs, which have been reported to have agonistic activity for TLR4, to create “adjuvant-free” antigens. The created antigens with triple TLR4 agonistic motifs in their C-terminus have activated NF-κB signaling pathways through TLR4. These proteins also induced the production of the inflammatory cytokine TNF-α, and the expression of the co-stimulatory molecule CD40 in APC, supporting the maturation of APC in vitro. Unexpectedly, these signal adjuvant-encrypted proteins have lost their ability to be physical adjuvants because they did not induce cytotoxic T lymphocytes (CTL) in vivo, while the parental proteins induced CTL. These results confirmed that the manifestation of a motif’s function is context-dependent and simple addition does not always work for motif-programing. Further optimization of the molecular context of the TLR4 agonistic motifs in antigens should be required to create adjuvant-free antigens. PMID:29190754

Design and synthesis of multifunctional gold nanoparticles bearing tumor-associated glycopeptide antigens as potential cancer vaccines.

PubMed

Brinãs, Raymond P; Sundgren, Andreas; Sahoo, Padmini; Morey, Susan; Rittenhouse-Olson, Kate; Wilding, Greg E; Deng, Wei; Barchi, Joseph J

2012-08-15

The development of vaccines against specific types of cancers will offer new modalities for therapeutic intervention. Here, we describe the synthesis of a novel vaccine construction prepared from spherical gold nanoparticles of 3-5 nm core diameters. The particles were coated with both the tumor-associated glycopeptides antigens containing the cell-surface mucin MUC4 with Thomsen Friedenreich (TF) antigen attached at different sites and a 28-residue peptide from the complement derived protein C3d to act as a B-cell activating "molecular adjuvant". The synthesis entailed solid-phase glycopeptide synthesis, design of appropriate linkers, and attachment chemistry of the various molecules to the particles. Attachment to the gold surface was mediated by a novel thiol-containing 33 atom linker which was further modified to be included as a third "spacer" component in the synthesis of several three-component vaccine platforms. Groups of mice were vaccinated either with one of the nanoplatform constructs or with control particles without antigen coating. Evaluation of sera from the immunized animals in enzyme immunoassays (EIA) against each glycopeptide antigen showed a small but statistically significant immune response with production of both IgM and IgG isotypes. Vaccines with one carbohydrate antigen (B, C, and E) gave more robust responses than the one with two contiguous disaccharides (D), and vaccine E with a TF antigen attached to threonine at the 10th position of the peptide was selected for IgG over IgM suggesting isotype switching. The data suggested that this platform may be a viable delivery system for tumor-associated glycopeptide antigens.

Extraction of Cell-Wall Polysaccharide Antigen from Streptococci

PubMed Central

Slade, Hutton D.

1965-01-01

Slade, Hutton D. (Northwestern University Medical School, Chicago, Ill., and Max-Planck Institut für Immunbiologie, Freiburg, Germany). Extraction of cell-wall polysaccharide antigen from streptococci. J. Bacteriol. 90:667–672. 1965.—The carbohydrate grouping antigens in the cell walls of streptococci belonging to groups A, E, G, L, and T were extracted with 5% trichloroacetic acid at 90 C. The antigens were removed also from dry whole cells by extraction with trichloroacetic acid followed by treatment with phenol-water. Details of the methods are presented. The antigens obtained by use of either of these procedures were suitable for studies on immunological specificity and chemical structure. Quantitative enzymatic and chemical analyses of two group E antigens and one group T preparation showed the presence of l-rhamnose (22 to 44%), d-glucose (7 to 22%), d-galactose (T antigen only, 26%), glucosamine (2 to 16%), and galactosamine (T antigen only, 3%). In addition, analyses of A and G antigen preparations are presented. The protein and phosphate content of the A and E antigens were about 1% each. Quantitative precipitin curves of these antigens are presented. PMID:16562065

Inhibitory effects of thymus-independent type 2 antigens on MHC class II-restricted antigen presentation: comparative analysis of carbohydrate structures and the antigen presenting cell.

PubMed

González-Fernández, M; Carrasco-Marín, E; Alvarez-Domínguez, C; Outschoorn, I M; Leyva-Cobián, F

1997-02-25

The role of thymus-independent type 2 (TI-2) antigens (polysaccharides) on the MHC-II-restricted processing of protein antigens was studied in vitro. In general, antigen presentation is inhibited when both peritoneal and splenic macrophages (M phi) as well as Küpffer cells (KC) are preincubated with acidic polysaccharides or branched dextrans. However, the inhibitory effect of neutral polysaccharides was minimal when KC were used as antigen presenting cells (APC). Morphological evaluation of the uptake of fluoresceinated polysaccharides clearly correlates with this selective and differential interference. Polysaccharides do not block MHC-I-restricted antigen presentation. Some chemical characteristics shared by different saccharides seem to be specially related to their potential inhibitory abilities: (i) those where two anomeric carbon atoms of two interlinked sugars and (ii) those containing several sulfate groups per disaccharide repeating unit. No polysaccharide being inhibitory in M phi abrogated antigen processing in other APC: lipopolysaccharide-activated B cells, B lymphoma cells, or dendritic cells (DC). Using radiolabeled polysaccharides it was observed that DC and B cells incorporated less radioactivity as a function of time than M phi. Morphological evaluation of these different APC incubated for extended periods of time with inhibitory concentrations of polysaccharides revealed intense cytoplasmic vacuolization in M phi but not in B cells or DC. The large majority of M phi lysosomes containing polysaccharides fail to fuse with incoming endocytic vesicles and delivery of fluid-phase tracers was reduced, suggesting that indigestible carbohydrates reduced the fusion of these loaded lysosomes with endosomes containing recently internalized tracers. It is suggested that the main causes of this antigen presentation blockade are (i) the chemical characteristics of certain carbohydrates and whether the specific enzymatic machinery for their intracellular

Recognition of Antigen-Specific B Cell Receptors From Chronic Lymphocytic Leukemia Patients By Synthetic Antigen Surrogates

PubMed Central

Sarkar, Mohosin; Liu, Yun; Morimoto, Jumpei; Peng, Haiyong; Aquino, Claudio; Rader, Christoph; Chiorazzi, Nicholas

2014-01-01

In patients with chronic lymphocytic leukemia (CLL), a single neoplastic antigen-specific B cell accumulates and overgrows other B cells, leading to immune deficiency. CLL is often treated with drugs that ablate all B cells, leading to further weakening of humoral immunity, and a more focused therapeutic strategy capable of targeting only the pathogenic B cells would represent a significant advance. One approach to this would be to develop synthetic surrogates of the CLL antigens allowing differentiation of the CLL cells and healthy B cells in a patient. Here, we describe discovery of non-peptidic molecules capable of targeting antigen-specific B cell receptors with good affinity and selectivity using a combinatorial library screen. We demonstrate that our hit compounds act as synthetic antigen surrogates and recognize CLL cells and not healthy B cells. Additionally, we argue that the technology we developed can be used for discovery of other classes of antigen surrogates. PMID:25467125

Recognition of antigen-specific B-cell receptors from chronic lymphocytic leukemia patients by synthetic antigen surrogates.

PubMed

Sarkar, Mohosin; Liu, Yun; Morimoto, Jumpei; Peng, Haiyong; Aquino, Claudio; Rader, Christoph; Chiorazzi, Nicholas; Kodadek, Thomas

2014-12-18

In patients with chronic lymphocytic leukemia (CLL), a single neoplastic antigen-specific B cell accumulates and overgrows other B cells, leading to immune deficiency. CLL is often treated with drugs that ablate all B cells, leading to further weakening of humoral immunity, and a more focused therapeutic strategy capable of targeting only the pathogenic B cells would represent a significant advance. One approach to this would be to develop synthetic surrogates of the CLL antigens allowing differentiation of the CLL cells and healthy B cells in a patient. Here, we describe nonpeptidic molecules capable of targeting antigen-specific B cell receptors with good affinity and selectivity using a combinatorial library screen. We demonstrate that our hit compounds act as synthetic antigen surrogates and recognize CLL cells and not healthy B cells. Additionally, we argue that the technology we developed can be used to identify other classes of antigen surrogates. Copyright © 2014 Elsevier Ltd. All rights reserved.

Antigenic change in feline calicivirus during persistent infection.

PubMed Central

Johnson, R P

1992-01-01

To determine if antigenic variation occurred during persistent infection of cats with feline caliciviruses (FCV), nine persistent (progeny) isolates from nine different carrier cats were compared antigenically to the original infecting parent strain, FCV 255, by two-way cross-neutralization tests with rabbit antisera. Five of the nine progeny viruses isolated 35 to 169 days after initial infection were antigenically different from the parent strain. These five isolates represented four distinct antigenic phenotypes. The emergence of four distinctly different antigenic variants from a single parent strain indicates that FCV, like many other RNA viruses, exhibits considerable antigenic heterogeneity during replication in its natural host, and supports the hypothesis that antigenic variation contributes to chronic FCV infection. PMID:1335833

Covalent binding of C3b to tetanus toxin: influence on uptake/internalization of antigen by antigen-specific and non-specific B cells.

PubMed Central

Villiers, M B; Villiers, C L; Jacquier-Sarlin, M R; Gabert, F M; Journet, A M; Colomb, M G

1996-01-01

Antigen opsonization by the C3b fragment of complement is a significant event in the modulation of cell-mediated immune response, but its mechanism is still largely unknown. The structural characteristics of C3b allow it to act as a bifunctional ligand between antigen and cells via their membrane C3b receptors. It was thus of interest to study the influence of the covalent link between C3b and antigen on the fixation and internalization of this antigen by antigen-presenting cells. Tetanus toxin (TT) was used as antigen, either free or covalently linked to C3b (TT-C3b). The antigen-presenting cells were TT-specific (4.2) or non-specific (BL15) Epstein-Barr virus (EBV)-transformed B cells. C3b was found to play an important role in antigen fixation and internalization by both antigen-specific and antigen non-specific cells. Covalent binding of C3b on TT (1) permitted fixation and internalization of this antigen by non-specific cells via their complement receptors; (2) enhanced antigen fixation and resulted in cross-linking between membrane immunoglobulins and complement receptors on antigen-specific cells. The consequences of covalent C3b binding to TT were analysed using antigen-specific and antigen-nonspecific cells. In both cases, a net increase in antigen fixation was observed. At the intracellular level, covalent C3b binding to TT resulted in a large TT incorporation in endosomes of nonspecific cells, similar to that observed in antigen-specific cells. Thus, C3b covalently linked to antigen enlarges the array of B-cell types capable of presenting antigen, including non-specific cells. Images Figure 2 PMID:8958046

Enhanced Expression of Interferon-γ-Induced Antigen-Processing Machinery Components in a Spontaneously Occurring Cancer1

PubMed Central

Cerruti, Fulvia; Martano, Marina; Petterino, Claudio; Bollo, Enrico; Morello, Emanuela; Bruno, Renato; Buracco, Paolo; Cascio, Paolo

2007-01-01

In human tumors, changes in the surface expression and/or function of major histocompatibility complex (MHC) class I antigens are frequently found and may provide malignant cells with a mechanism to escape control of the immune system. This altered human lymphocyte antigen (HLA) class I phenotype can be caused by either structural alterations or dysregulation of genes encoding subunits of HLA class I antigens and/or components of the MHC class I antigen-processing machinery (APM). Herein we analyze the expression of several proteins involved in the generation of MHC class I epitopes in feline injection site sarcoma, a spontaneously occurring tumor in cats that is an informativemodel for the study of tumor biology in other species, including humans. Eighteen surgically removed primary fibrosarcoma lesions were analyzed, and an enhanced expression of two catalytic subunits of immunoproteasomes, PA28 and leucine aminopeptidase, was found in tumors compared to matched normal tissues. As a functional counterpart of these changes in protein levels, proteasomal activities were increased in tissue extracts from fibrosarcomas. Taken together, these results suggest that alterations in the APM system may account for reduced processing of selected tumor antigens and may potentially provide neoplastic fibroblasts with a mechanism for escape from T-cell recognition and destruction. PMID:18030364

Soluble Antigen Fluorescent-Antibody Technique

PubMed Central

Toussaint, Andre J.; Anderson, Robert I.

1965-01-01

An indirect fluorescent-antibody (FA) procedure employing soluble antigen fixed onto an artificial matrix was developed, and a mechanical means for reading of test results was devised. The method employs two small cellulose acetate paper discs for each test. One disc contains soluble antigen diluted in 1% bovine serum albumin (BSA); the other contains only 1% BSA and serves as a control. After testing by the indirect FA procedure, the results of the tests are read on a fluorometer fitted with a paper chromatogram door. The instrument is set at zero with the control disc as a blank, and the specific fluorescence of the antigen disc is determined. Findings obtained with homologous and heterologous antisera indicated that the method yields excellent results. The soluble antigen fluorescent-antibody technique has definite advantages over the conventional indirect FA procedures. (i) The investigator may objectively select the antigen to be employed. (ii) It is possible to obtain objective mechanical reading of test results rather than the highly subjective readings required by conventional methods. (iii) The system compensates for any nonspecific fluorescence contributed either by the serum (e.g., drugs) or by free fluorescein in the conjugated antiserum. Images Fig. 1 PMID:14339261

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

PubMed

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

2017-08-01

To investigate the predictive value of various molecular forms of prostate-specific antigen in men with baseline prostate-specific antigen <2.0 ng/mL. The case cohort comprised 150 men with a baseline prostate-specific antigen level <2.0 ng/mL, and who developed prostate cancer within 10 years. The control cohort was 300 baseline prostate-specific antigen- and age-adjusted men who did not develop prostate cancer. Serum prostate-specific antigen, free prostate-specific antigen, and [-2] proenzyme prostate-specific antigen were measured at baseline and last screening visit. The predictive impact of baseline prostate-specific antigen- and [-2] proenzyme prostate-specific antigen-related indices on developing prostate cancer was investigated. The predictive impact of those indices at last screening visit and velocities from baseline to final screening on tumor aggressiveness were also investigated. The baseline free to total prostate-specific antigen ratio was a significant predictor of prostate cancer development. The odds ratio was 6.08 in the lowest quintile baseline free to total prostate-specific antigen ratio subgroup. No serum indices at diagnosis were associated with tumor aggressiveness. The Prostate Health Index velocity and [-2] proenzyme prostate-specific antigen/free prostate-specific antigen velocity significantly increased in patients with higher risk D'Amico risk groups and higher Gleason scores. Free to total prostate-specific antigen ratio in men with low baseline prostate-specific antigen levels seems to predict the risk of developing prostate cancer, and it could be useful for a more effective individualized screening system. Longitudinal changes in [-2] proenzyme prostate-specific antigen-related indices seem to correlate with tumor aggressiveness, and they could be used as prognostic tool before treatment and during active surveillance. © 2017 The Japanese Urological Association.

Structural Basis of Preexisting Immunity to the 2009 H1N1 Pandemic Influenza Virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Rui; Ekiert, Damian C.; Krause, Jens C.

The 2009 H1N1 swine flu is the first influenza pandemic in decades. The crystal structure of the hemagglutinin from the A/California/04/2009 H1N1 virus shows that its antigenic structure, particularly within the Sa antigenic site, is extremely similar to those of human H1N1 viruses circulating early in the 20th century. The cocrystal structure of the 1918 hemagglutinin with 2D1, an antibody from a survivor of the 1918 Spanish flu that neutralizes both 1918 and 2009 H1N1 viruses, reveals an epitope that is conserved in both pandemic viruses. Thus, antigenic similarity between the 2009 and 1918-like viruses provides an explanation for themore » age-related immunity to the current influenza pandemic.« less

Specific Fluorine Labeling of the HyHEL10 Antibody Affects Antigen Binding and Dynamics

PubMed Central

Acchione, Mauro; Lee, Yi-Chien; DeSantis, Morgan E.; Lipschultz, Claudia A.; Wlodawer, Alexander; Li, Mi; Shanmuganathan, Aranganathan; Walter, Richard L.; Smith-Gill, Sandra; Barchi, Joseph J.

2012-01-01

To more fully understand the molecular mechanisms responsible for variations in binding affinity with antibody maturation, we explored the use of site specific fluorine labeling and 19F nuclear magnetic resonance (NMR). Several single-chain (scFv) antibodies, derived from an affinity-matured series of anti-hen egg white lysozyme (HEL) mouse IgG1, were constructed with either complete or individual replacement of tryptophan residues with 5-fluorotryptophan (5FW). An array of biophysical techniques was used to gain insight into the impact of fluorine substitution on the overall protein structure and antigen binding. SPR measurements indicated that 5FW incorporation lowered binding affinity for the HEL antigen. The degree of analogue impact was residue-dependent, and the greatest decrease in affinity was observed when 5FW was substituted for residues near the binding interface. In contrast, corresponding crystal structures in complex with HEL were essentially indistinguishable from the unsubstituted antibody. 19F NMR analysis showed severe overlap of signals in the free fluorinated protein that was resolved upon binding to antigen, suggesting very distinct chemical environments for each 5FW in the complex. Preliminary relaxation analysis suggested the presence of chemical exchange in the antibody–antigen complex that could not be observed by X-ray crystallography. These data demonstrate that fluorine NMR can be an extremely useful tool for discerning structural changes in scFv antibody–antigen complexes with altered function that may not be discernible by other biophysical techniques. PMID:22769726

CELL SEPARATION ON ANTIGEN-COATED COLUMNS

PubMed Central

Wigzell, Hans; Andersson, Birger

1969-01-01

Glass and plastic bead columns coated with antigenic protein molecules were used as an immunological filter for cell populations containing immune cells of relevant specificity. A selective elimination of these immune cells from the passing cell suspension was regularly noted and it approached, in some experiments, complete abolition of the specific immune reactivity of the filtered cell population. This specific retention of immune cells by antigenic columns could be selectively blocked by the presence of free antigen molecules in the medium during filtration. The results obtained support the concept of a cell-associated antigen-specific receptor being present on the outer surface of immune cells, displaying the same antigen-binding specificity as the potential product of the cell, the humoral antibody. Using the present bead column system, results were obtained indicating that this receptor was an active product of the immune cells and not any passively adsorbed, cytophilic antibody. Antigenic bead columns may very well constitute a tool for the production in vitro of cell populations being specifically deprived of immune reactivity and allow detailed analysis of the characteristics of the cell-associated antibody of immune cells. PMID:5782770

Antigenic Fingerprinting following Primary RSV Infection in Young Children Identifies Novel Antigenic Sites and Reveals Unlinked Evolution of Human Antibody Repertoires to Fusion and Attachment Glycoproteins

PubMed Central

Fuentes, Sandra; Coyle, Elizabeth M.; Beeler, Judy; Golding, Hana; Khurana, Surender

2016-01-01

Respiratory Syncytial Virus (RSV) is the major cause of pneumonia among infants. Here we elucidated the antibody repertoire following primary RSV infection and traced its evolution through adolescence and adulthood. Whole genome-fragment phage display libraries (GFPDL) expressing linear and conformational epitopes in the RSV fusion protein (F) and attachment protein (G) were used for unbiased epitope profiling of infant sera prior to and following RSV infection. F-GFPDL analyses demonstrated modest changes in the anti-F epitope repertoires post-RSV infection, while G-GFPDL analyses revealed 100-fold increase in number of bound phages. The G-reactive epitopes spanned the N- and C-terminus of the G ectodomain, along with increased reactivity to the central conserved domain (CCD). Panels of F and G antigenic sites were synthesized to evaluate sera from young children (<2 yr), adolescents (14–18 yr) and adults (30–45 yr) in SPR real-time kinetics assays. A steady increase in RSV-F epitope repertoires from young children to adults was observed using peptides and F proteins. Importantly, several novel epitopes were identified in pre-fusion F and an immunodominant epitope in the F-p27. In all age groups, antibody binding to pre-fusion F was 2–3 folds higher than to post-fusion form. For RSV-G, antibody responses were high following early RSV infection in children, but declined significantly in adults, using either G proteins or peptides. This study identified unlinked evolution of anti-F and anti G responses and supportive evidence for immune pressure driven evolution of RSV-G. These findings could help development of effective countermeasures including vaccines. PMID:27100289

Protamine-based nanoparticles as new antigen delivery systems.

PubMed

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

2015-11-01

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

Fusion of antigen to a dendritic cell targeting chemokine combined with adjuvant yields a malaria DNA vaccine with enhanced protective capabilities.

PubMed

Luo, Kun; Zhang, Hong; Zavala, Fidel; Biragyn, Arya; Espinosa, Diego A; Markham, Richard B

2014-01-01

Although sterilizing immunity to malaria can be elicited by irradiated sporozoite vaccination, no clinically practical subunit vaccine has been shown to be capable of preventing the approximately 600,000 annual deaths attributed to this infection. DNA vaccines offer several potential advantages for a disease that primarily affects the developing world, but new approaches are needed to improve the immunogenicity of these vaccines. By using a novel, lipid-based adjuvant, Vaxfectin, to attract immune cells to the immunization site, in combination with an antigen-chemokine DNA construct designed to target antigen to immature dendritic cells, we elicited a humoral immune response that provided sterilizing immunity to malaria challenge in a mouse model system. The chemokine, MIP3αCCL20, did not significantly enhance the cellular infiltrate or levels of cytokine or chemokine expression at the immunization site but acted with Vaxfectin to reduce liver stage malaria infection by orders of magnitude compared to vaccine constructs lacking the chemokine component. The levels of protection achieved were equivalent to those observed with irradiated sporozoites, a candidate vaccine undergoing development for further large scale clinical trial. Only vaccination with the combined regimen of adjuvant and chemokine provided 80-100% protection against the development of bloodstream infection. Treating the immunization process as requiring the independent steps of 1) attracting antigen-presenting cells to the site of immunization and 2) specifically directing vaccine antigen to the immature dendritic cells that initiate the adaptive immune response may provide a rational strategy for the development of a clinically applicable malaria DNA vaccine.

Human heat shock protein 70 enhances tumor antigen presentation through complex formation and intracellular antigen delivery without innate immune signaling.

PubMed

Bendz, Henriette; Ruhland, Sibylle C; Pandya, Maya J; Hainzl, Otmar; Riegelsberger, Stefan; Braüchle, Christoph; Mayer, Matthias P; Buchner, Johannes; Issels, Rolf D; Noessner, Elfriede

2007-10-26

Heat shock proteins (HSPs) have shown promise for the optimization of protein-based vaccines because they can transfer exogenous antigens to dendritic cells and at the same time induce their maturation. Great care must be exercised in interpretating HSP-driven studies, as by-products linked to the recombinant generation of these proteins have been shown to mediate immunological effects. We generated highly purified human recombinant Hsp70 and demonstrated that it strongly enhances the cross-presentation of exogenous antigens resulting in better antigen-specific T cell stimulation. Augmentation of T cell stimulation was a direct function of the degree of complex formation between Hsp70 and peptides and correlated with improved antigen delivery to endosomal compartments. The Hsp70 activity was independent of TAP proteins and was not inhibited by exotoxin A or endosomal acidification. Consequently, Hsp70 enhanced cross-presentation of various antigenic sequences, even when they required different post-uptake processing and trafficking, as exemplified by the tumor antigens tyrosinase and Melan-A/MART-1. Furthermore, Hsp70 enhanced cross-presentation by different antigen-presenting cells (APCs), including dendritic cells and B cells. Importantly, enhanced cross-presentation and antigen-specific T cell activation were observed in the absence of innate signals transmitted by Hsp70. As Hsp70 supports the cross-presentation of different antigens and APCs and is inert to APC function, it may show efficacy in various settings of immune modulation, including induction of antigen-specific immunity or tolerance.

Non-stoichiometric O-acetylation of Shigella flexneri 2a O-specific polysaccharide: synthesis and antigenicity.

PubMed

Gauthier, Charles; Chassagne, Pierre; Theillet, François-Xavier; Guerreiro, Catherine; Thouron, Françoise; Nato, Farida; Delepierre, Muriel; Sansonetti, Philippe J; Phalipon, Armelle; Mulard, Laurence A

2014-06-28

Synthetic functional mimics of the O-antigen from Shigella flexneri 2a are seen as promising vaccine components against endemic shigellosis. Herein, the influence of the polysaccharide non-stoichiometric di-O-acetylation on antigenicity is addressed for the first time. Three decasaccharides, representing relevant internal mono- and di-O-acetylation profiles of the O-antigen, were synthesized from a pivotal protected decasaccharide designed to tailor late stage site-selective O-acetylation. The latter was obtained via a convergent route involving the imidate glycosylation chemistry. Binding studies to five protective mIgGs showed that none of the acetates adds significantly to broad antibody recognition. Yet, one of the five antibodies had a unique pattern of binding. With IC50 in the micromolar to submicromolar range mIgG F22-4 exemplifies a remarkable tight binding antibody against diversely O-acetylated and non-O-acetylated fragments of a neutral polysaccharide of medical importance.

Peptide-dependent Conformational Fluctuation Determines the Stability of the Human Leukocyte Antigen Class I Complex*

PubMed Central

Yanaka, Saeko; Ueno, Takamasa; Shi, Yi; Qi, Jianxun; Gao, George F.; Tsumoto, Kouhei; Sugase, Kenji

2014-01-01

In immune-mediated control of pathogens, human leukocyte antigen (HLA) class I presents various antigenic peptides to CD8+ T-cells. Long-lived peptide presentation is important for efficient antigen-specific T-cell activation. Presentation time depends on the peptide sequence and the stability of the peptide-HLA complex (pHLA). However, the determinant of peptide-dependent pHLA stability remains elusive. Here, to reveal the pHLA stabilization mechanism, we examined the crystal structures of an HLA class I allomorph in complex with HIV-derived peptides and evaluated site-specific conformational fluctuations using NMR. Although the crystal structures of various pHLAs were almost identical independent of the peptides, fluctuation analyses identified a peptide-dependent minor state that would be more tightly packed toward the peptide. The minor population correlated well with the thermostability and cell surface presentation of pHLA, indicating that this newly identified minor state is important for stabilizing the pHLA and facilitating T-cell recognition. PMID:25028510

Structure and genetic variability of envelope glycoproteins of two antigenic variants of caprine arthritis-encephalitis lentivirus.

PubMed

Knowles, D P; Cheevers, W P; McGuire, T C; Brassfield, A L; Harwood, W G; Stem, T A

1991-11-01

To define the structure of the caprine arthritis-encephalitis virus (CAEV) env gene and characterize genetic changes which occur during antigenic variation, we sequenced the env genes of CAEV-63 and CAEV-Co, two antigenic variants of CAEV defined by serum neutralization. The deduced primary translation product of the CAEV env gene consists of a 60- to 80-amino-acid signal peptide followed by an amino-terminal surface protein (SU) and a carboxy-terminal transmembrane protein (TM) separated by an Arg-Lys-Lys-Arg cleavage site. The signal peptide cleavage site was verified by amino-terminal amino acid sequencing of native CAEV-63 SU. In addition, immunoprecipitation of [35S]methionine-labeled CAEV-63 proteins by sera from goats immunized with recombinant vaccinia virus expressing the CAEV-63 env gene confirmed that antibodies induced by env-encoded recombinant proteins react specifically with native virion SU and TM. The env genes of CAEV-63 and CAEV-Co encode 28 conserved cysteines and 25 conserved potential N-linked glycosylation sites. Nucleotide sequence variability results in 62 amino acid changes and one deletion within the SU and 34 amino acid changes within the TM.

Structure and genetic variability of envelope glycoproteins of two antigenic variants of caprine arthritis-encephalitis lentivirus.

PubMed Central

Knowles, D P; Cheevers, W P; McGuire, T C; Brassfield, A L; Harwood, W G; Stem, T A

1991-01-01

To define the structure of the caprine arthritis-encephalitis virus (CAEV) env gene and characterize genetic changes which occur during antigenic variation, we sequenced the env genes of CAEV-63 and CAEV-Co, two antigenic variants of CAEV defined by serum neutralization. The deduced primary translation product of the CAEV env gene consists of a 60- to 80-amino-acid signal peptide followed by an amino-terminal surface protein (SU) and a carboxy-terminal transmembrane protein (TM) separated by an Arg-Lys-Lys-Arg cleavage site. The signal peptide cleavage site was verified by amino-terminal amino acid sequencing of native CAEV-63 SU. In addition, immunoprecipitation of [35S]methionine-labeled CAEV-63 proteins by sera from goats immunized with recombinant vaccinia virus expressing the CAEV-63 env gene confirmed that antibodies induced by env-encoded recombinant proteins react specifically with native virion SU and TM. The env genes of CAEV-63 and CAEV-Co encode 28 conserved cysteines and 25 conserved potential N-linked glycosylation sites. Nucleotide sequence variability results in 62 amino acid changes and one deletion within the SU and 34 amino acid changes within the TM. Images PMID:1656067

Natural Mutations in Streptococcus agalactiae Resulting in Abrogation of β Antigen Production

PubMed Central

Vasilyeva, Anastasia; Santos Sanches, Ilda; Florindo, Carlos; Dmitriev, Alexander

2015-01-01

Streptococcus agalactiae genome encodes 21 two-component systems (TCS) and a variety of regulatory proteins in order to control gene expression. One of the TCS, BgrRS, comprising the BgrR DNA-binding regulatory protein and BgrS sensor histidine kinase, was discovered within a putative virulence island. BgrRS influences cell metabolism and positively control the expression of bac gene, coding for β antigen at transcriptional level. Inactivation of bgrR abrogated bac gene expression and increased virulence properties of S. agalactiae. In this study, a total of 140 strains were screened for the presence of bac gene, and the TCS bgrR and bgrS genes. A total of 53 strains carried the bac, bgrR and bgrS genes. Most of them (48 strains) expressed β antigen, while five strains did not express β antigen. Three strains, in which bac gene sequence was intact, while bgrR and/or bgrS genes had mutations, and expression of β antigen was absent, were complemented with a constructed plasmid pBgrRS(P) encoding functionally active bgrR and bgrS gene alleles. This procedure restored expression of β antigen indicating the crucial regulatory role of TCS BgrRS. The complemented strain A49V/BgrRS demonstrated attenuated virulence in intraperitoneal mice model of S. agalactiae infection compared to parental strain A49V. In conclusion we showed that disruption of β antigen expression is associated with: i) insertion of ISSa4 upstream the bac gene just after the ribosomal binding site; ii) point mutation G342A resulting a stop codon TGA within the bac gene and a truncated form of β antigen; iii) single deletion (G) in position 439 of the bgrR gene resulting in a frameshift and the loss of DNA-binding domain of the BgrR protein, and iv) single base substitutions in bgrR and bgrS genes causing single amino acid substitutions in BgrR (Arg187Lys) and BgrS (Arg252Gln). The fact that BgrRS negatively controls virulent properties of S. agalactiae gives a novel clue for understanding of S

Natural Mutations in Streptococcus agalactiae Resulting in Abrogation of β Antigen Production.

PubMed

Vasilyeva, Anastasia; Santos Sanches, Ilda; Florindo, Carlos; Dmitriev, Alexander

2015-01-01

Streptococcus agalactiae genome encodes 21 two-component systems (TCS) and a variety of regulatory proteins in order to control gene expression. One of the TCS, BgrRS, comprising the BgrR DNA-binding regulatory protein and BgrS sensor histidine kinase, was discovered within a putative virulence island. BgrRS influences cell metabolism and positively control the expression of bac gene, coding for β antigen at transcriptional level. Inactivation of bgrR abrogated bac gene expression and increased virulence properties of S. agalactiae. In this study, a total of 140 strains were screened for the presence of bac gene, and the TCS bgrR and bgrS genes. A total of 53 strains carried the bac, bgrR and bgrS genes. Most of them (48 strains) expressed β antigen, while five strains did not express β antigen. Three strains, in which bac gene sequence was intact, while bgrR and/or bgrS genes had mutations, and expression of β antigen was absent, were complemented with a constructed plasmid pBgrRS(P) encoding functionally active bgrR and bgrS gene alleles. This procedure restored expression of β antigen indicating the crucial regulatory role of TCS BgrRS. The complemented strain A49V/BgrRS demonstrated attenuated virulence in intraperitoneal mice model of S. agalactiae infection compared to parental strain A49V. In conclusion we showed that disruption of β antigen expression is associated with: i) insertion of ISSa4 upstream the bac gene just after the ribosomal binding site; ii) point mutation G342A resulting a stop codon TGA within the bac gene and a truncated form of β antigen; iii) single deletion (G) in position 439 of the bgrR gene resulting in a frameshift and the loss of DNA-binding domain of the BgrR protein, and iv) single base substitutions in bgrR and bgrS genes causing single amino acid substitutions in BgrR (Arg187Lys) and BgrS (Arg252Gln). The fact that BgrRS negatively controls virulent properties of S. agalactiae gives a novel clue for understanding of S

Level of Need for Cognition and Metacognitive Thinking among Undergraduate Kindergarten Female Students at King Sa'ud University in Sa'udi Arabia

ERIC Educational Resources Information Center

Daghistani, Bulquees

2015-01-01

This study aims at examining the level of need for cognition and metacognitive thinking among undergraduate kindergarten female students in Education Faculty at King Sa'ud University in Sa'udi Arabia from their own perceptions. Results showed that the need for the cognition level was moderate, but metacognitive thinking level was high. In…

Chemoselective ligation and antigen vectorization.

PubMed

Gras-Masse, H

2001-01-01

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

Antibody-Antigen-Adjuvant Conjugates Enable Co-Delivery of Antigen and Adjuvant to Dendritic Cells in Cis but Only Have Partial Targeting Specificity

PubMed Central

Abuknesha, Ram; Uematsu, Satoshi; Akira, Shizuo; Nestle, Frank O.; Diebold, Sandra S.

2012-01-01

Antibody-antigen conjugates, which promote antigen-presentation by dendritic cells (DC) by means of targeted delivery of antigen to particular DC subsets, represent a powerful vaccination approach. To ensure immunity rather than tolerance induction the co-administration of a suitable adjuvant is paramount. However, co-administration of unlinked adjuvant cannot ensure that all cells targeted by the antibody conjugates are appropriately activated. Furthermore, antigen-presenting cells (APC) that do not present the desired antigen are equally strongly activated and could prime undesired responses against self-antigens. We, therefore, were interested in exploring targeted co-delivery of antigen and adjuvant in cis in form of antibody-antigen-adjuvant conjugates for the induction of anti-tumour immunity. In this study, we report on the assembly and characterization of conjugates consisting of DEC205-specific antibody, the model antigen ovalbumin (OVA) and CpG oligodeoxynucleotides (ODN). We show that such conjugates are more potent at inducing cytotoxic T lymphocyte (CTL) responses than control conjugates mixed with soluble CpG. However, our study also reveals that the nucleic acid moiety of such antibody-antigen-adjuvant conjugates alters their binding and uptake and allows delivery of the antigen and the adjuvant to cells partially independently of DEC205. Nevertheless, antibody-antigen-adjuvant conjugates are superior to antibody-free antigen-adjuvant conjugates in priming CTL responses and efficiently induce anti-tumour immunity in the murine B16 pseudo-metastasis model. A better understanding of the role of the antibody moiety is required to inform future conjugate vaccination strategies for efficient induction of anti-tumour responses. PMID:22808118

Mathematics teachers' beliefs about scientific approach (SA) and implementation in mathematics learning

NASA Astrophysics Data System (ADS)

Mutholib, Ahmad Abdul; Sujadi, Imam; Subanti, Sri

2017-08-01

SA is the approach used for the exploration of research and answer questions. Teachers' beliefs have a greater influence than the teacher's knowledge of designing lesson plans in the classroom. The objectives of this study are to explore the teachers' beliefs in SA, to reveal how the beliefs are reflected in classroom practices; and to figure out the factors affecting their beliefs and practices of SA to the teaching of mathematics. This qualitative research applied case study. The data was gained from classroom observation, face-to-face interview, and documentation. Interactive models from Miles and Huberman were used to examine the data. Results of the study: 1) The teachers believe about the conception of SA. They also believe that the SA is important and gives impact to students' progress. They believe that by applying SA, the target of mathematics learning is acquired. As to learning procedure, they believe that SA steps are conducted in sequence by combining some steps for each. 2) Teachers formulate their beliefs of applying the five scientific step of integrating all steps by keeping the sequence. Teachers argue that target of mathematics learning can be attained by some ways, namely presence of theoretical and practical support, teachers' guidance, providing variety of media and motivation to students. 3) There are five factors which influence teachers' beliefs and practices of SA, namely learning and teaching experience, teachers' motivation, sharing with colleagues and facility. This study concludes that teachers believe in the importance of SA, therefore they implement it in the classroom.

Cosmetic outcome of skin adhesives versus transcutaneous sutures in laparoscopic port-site wounds: a prospective randomized controlled trial.

PubMed

Buchweitz, Olaf; Frye, Christian; Moeller, Claus Peter; Nugent, Wolfgang; Krueger, Eckart; Nugent, Andreas; Biel, Peter; Juergens, Sven

2016-06-01

In an elective laparoscopic surgery, the cosmetic outcome becomes increasingly important. We conducted a study to evaluate the cosmetic outcome 3 months after a laparoscopic procedure and compared skin adhesive (SA) versus transcutaneous suture (TS). A randomized, controlled, prospective study was conducted at a single study centre in Hamburg, Germany. Seventy-seven patients undergoing laparoscopic surgery with two lower abdominal port sites met the study requirements. It was decided randomly which port site would be closed with SA. The opposite site was closed with TS. Wounds were assessed after 7-12 days and after 3 months. Cosmetic outcome was measured by a visual analogue scale (VAS) completed by the patient, by the Hollander wound evaluation scale (HWES) and by the judgement of blinded investigators. Seventy-seven subjects were randomized. Complete data from the 3-month follow-up visit were available from 56 patients (72.7 %). The VAS scale ranged from 0 to 100 mm with "0" representing the best possible cosmetic outcome. Median satisfaction was 2 mm in the TS group and 3 mm in the SA group. The mean was high in both groups 4.6 (s = 13.1) versus 3.8 mm (s = 4.6). The outcome was neither clinically nor statistically significant. Cosmetic outcome was assessed by an investigator, and the HWES showed no difference. In regard to complications, no difference was found between SA and TS, either. In conclusion this study demonstrated that closure of laparoscopic port-site wounds leads to equivalent outcomes whether SAs or TSs are used. Complications are rare in both methods. Thus, SAs seem to be a valid alternative to sutures in laparoscopic surgery. Registration site: www.clinicaltrials.gov . NCT02179723.

Podocytes Are Nonhematopoietic Professional Antigen-Presenting Cells

PubMed Central

Burkard, Miriam; Ölke, Martha; Daniel, Christoph; Amann, Kerstin; Hugo, Christian; Kurts, Christian; Steinkasserer, Alexander; Gessner, André

2013-01-01

Podocytes are essential to the structure and function of the glomerular filtration barrier; however, they also exhibit increased expression of MHC class II molecules under inflammatory conditions, and they remove Ig and immune complexes from the glomerular basement membrane (GBM). This finding suggests that podocytes may act as antigen-presenting cells, taking up and processing antigens to initiate specific T cell responses, similar to professional hematopoietic cells such as dendritic cells or macrophages. Here, MHC–antigen complexes expressed exclusively on podocytes of transgenic mice were sufficient to activate CD8+ T cells in vivo. In addition, deleting MHC class II exclusively on podocytes prevented the induction of experimental anti-GBM nephritis. Podocytes ingested soluble and particulate antigens, activated CD4+ T cells, and crosspresented exogenous antigen on MHC class I molecules to CD8+ T cells. In conclusion, podocytes participate in the antigen-specific activation of adaptive immune responses, providing a potential target for immunotherapies of inflammatory kidney diseases and transplant rejection. PMID:23539760

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

PubMed

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

2015-12-01

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

Infrequent and low expression of cancer-testis antigens located on the X chromosome in colorectal cancer: implications for immunotherapy in South African populations.

PubMed

Dakshinamurthy, Amirtha Ganesh; Ramesar, Rajkumar; Goldberg, Paul; Blackburn, Jonathan M

2008-11-01

Cancer-testis (CT) antigens are a group of tumor antigens that are expressed in the testis and aberrantly in cancerous tissue but not in somatic tissues. The testis is an immune-privileged site because of the presence of a blood-testis barrier; as a result, CT antigens are considered to be essentially tumor specific and are attractive targets for immunotherapy. CT antigens are classified as the CT-X and the non-X CT antigens depending on the chromosomal location to which the genes are mapped. CT-X antigens are typically highly immunogenic and hence the first step towards tailored immunotherapy is to elucidate the expression profile of CT-X antigens in the respective tumors. In this study we investigated the expression profile of 16 CT-X antigen genes in 34 colorectal cancer (CRC) patients using reverse transcription-polymerase chain reaction. We observed that 12 of the 16 CT-X antigen genes studied did not show expression in any of the CRC samples analyzed. The other 4 CT-X antigen genes showed low frequency of expression and exhibited a highly variable expression profile when compared to other populations. Thus, our study forms the first report on the expression profile of CT-X antigen genes among CRC patients in the genetically diverse South African population. The results of our study suggest that genetic and ethnic variations in population might have a role in the expression of the CT-X antigen genes. Thus our results have significant implications for anti-CT antigen-based immunotherapy trials in this population.

[HL-A antigen distribution in duodenal ulcer patients].

PubMed

Ilieva, P; Minev, M; Etŭrska, M

1980-01-01

The incidence of HLA-antigens was studied in 405 patients with clinically, roentgenologically and gastroscopically confirmed ulcer of duodenum and in 1085 controls, healthy subjects. Increased incidence of both antigens from locus B was established among the patients: HLA-B17 and HLA-BW21. A reduction of HLA-A3 incidence was found from the antigens of locus A. In the determination of incidence of HLA antigenes, depending on blood grouping ABO both of patients and healthy subjects, it was established that antigen HLA-A3 is less frequent in the patients with blood group B, whereas antigen HLA-B12 is found more often among the patients with blood group A.

Development of the JT-60SA Neutral Beam Injectors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hanada, M.; Kojima, A.; Inoue, T.

2011-09-26

This paper describes the development of the neutral beam (NB) systems on JT-60SA, where 30-34 MW D{sup 0} beams are required to be injected for 100 s. A 30 s operation of the NB injectors suggests that existing beamline components and positive ion sources on JT-60U can be reused without the modifications on JT-60 SA. The JT-60 negative ion source was modified to improve the voltage holding capability, which leads to a successful acceleration of 2.8 A H{sup -} ion beam up to 500 keV of the rated acceleration energy for JT-60SA.

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Federal Register 2010, 2011, 2012, 2013, 2014

2012-05-01

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Antigen-mediated regulation in monoclonal gammopathies and myeloma.

PubMed

Nair, Shiny; Sng, Joel; Boddupalli, Chandra Sekhar; Seckinger, Anja; Chesi, Marta; Fulciniti, Mariateresa; Zhang, Lin; Rauniyar, Navin; Lopez, Michael; Neparidze, Natalia; Parker, Terri; Munshi, Nikhil C; Sexton, Rachael; Barlogie, Bart; Orlowski, Robert; Bergsagel, Leif; Hose, Dirk; Flavell, Richard A; Mistry, Pramod K; Meffre, Eric; Dhodapkar, Madhav V

2018-04-19

A role for antigen-driven stimulation has been proposed in the pathogenesis of monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) based largely on the binding properties of monoclonal Ig. However, insights into antigen binding to clonal B cell receptors and in vivo responsiveness of the malignant clone to antigen-mediated stimulation are needed to understand the role of antigenic stimulation in tumor growth. Lysolipid-reactive clonal Ig were detected in Gaucher disease (GD) and some sporadic gammopathies. Here, we show that recombinant Ig (rIg) cloned from sort-purified single tumor cells from lipid-reactive sporadic and GD-associated gammopathy specifically bound lysolipids. Liposome sedimentation and binding assays confirmed specific interaction of lipid-reactive monoclonal Ig with lysolipids. The clonal nature of lysolipid-binding Ig was validated by protein sequencing. Gene expression profiling and cytogenetic analyses from 2 patient cohorts showed enrichment of nonhyperdiploid tumors in lipid-reactive patients. In vivo antigen-mediated stimulation led to an increase in clonal Ig and plasma cells (PCs) in GD gammopathy and also reactivated previously suppressed antigenically related nonclonal PCs. These data support a model wherein antigenic stimulation mediates an initial polyclonal phase, followed by evolution of monoclonal tumors enriched in nonhyperdiploid genomes, responsive to underlying antigen. Targeting underlying antigens may therefore prevent clinical MM.

Structural mimicry of O-antigen by a peptide revealed in a complex with an antibody raised against Shigella flexneri serotype 2a.

PubMed

Theillet, François-Xavier; Saul, Frederick A; Vulliez-Le Normand, Brigitte; Hoos, Sylviane; Felici, Franco; Weintraub, Andrej; Mulard, Laurence A; Phalipon, Armelle; Delepierre, Muriel; Bentley, Graham A

2009-05-15

The use of carbohydrate-mimicking peptides to induce immune responses against surface polysaccharides of pathogenic bacteria offers a novel approach to vaccine development. Factors governing antigenic and immunogenic mimicry, however, are complex and poorly understood. We have addressed this question using the anti-lipopolysaccharide monoclonal antibody F22-4, which was raised against Shigella flexneri serotype 2a and shown to protect against homologous infection in a mouse model. In a previous crystallographic study, we described F22-4 in complex with two synthetic fragments of the O-antigen, the serotype-specific saccharide moiety of lipopolysaccharide. Here, we present a crystallographic and NMR study of the interaction of F22-4 with a dodecapeptide selected by phage display using the monoclonal antibody. Like the synthetic decasaccharide, the peptide binds to F22-4 with micromolar affinity. Although the peptide and decasaccharide use very similar regions of the antigen-binding site, indicating good antigenic mimicry, immunogenic mimicry by the peptide was not observed. The F22-4-antigen interaction is significantly more hydrophobic with the peptide than with oligosaccharides; nonetheless, all hydrogen bonds formed between the peptide and F22-4 have equivalents in the oligosaccharide complex. Two bridging water molecules are also in common, adding to partial structural mimicry. Whereas the bound peptide is entirely helical, its structure in solution, as shown by NMR, is helical in the central region only. Moreover, docking the NMR structure into the antigen-binding site shows that steric hindrance would occur, revealing poor complementarity between the major solution conformation and the antibody that could contribute to the absence of immunogenic mimicry.

Immunity to Intracellular Salmonella Depends on Surface-associated Antigens

PubMed Central

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

Preliminary treatment of bovine mastitis caused by Staphylococcus aureus, with trx-SA1, recombinant endolysin of S. aureus bacteriophage IME-SA1.

PubMed

Fan, Jindai; Zeng, Zhiliang; Mai, Kaijie; Yang, Yu; Feng, Jiaqi; Bai, Yang; Sun, Baoli; Xie, Qingmei; Tong, Yigang; Ma, Jingyun

2016-08-15

Methicillin-resistant Staphylococcus aureus (MRSA) has become a great threat to human and animal health and there is an urgent need to develop novel antibacterial agents to control this pathogen. The objective of this study was to obtain an active recombinant endolysin from the novel bacteriophage (IME-SA1), and conduct an efficacy trial of its effectiveness against bovine mastitis. We isolated a phage that was virulent and specific for S. aureus with an optimal multiplicity of infection of 0.01. Electron microscopy revealed that IME-SA1 was a member of the family Myoviridae, with an isometric head (98nm) and a long contractile tail (200nm). Experimental lysis experiments indicated the phage had an incubation period of 20min with a burst size of 80. When host bacteria were in early exponential growth stages, a multiplicity of infection of 0.01 resulted in a complete bacterial lysis after 9h. The endolysin gene (804bp) was cloned into the pET-32a bacterial expression vector and recombinant endolysin Trx-SA1 was successfully obtained with molecular size of about 47kDa. Preliminary results of therapeutic trials in cow udders showed that Trx-SA1 could effectively control mild clinical mastitis caused by S. aureus. The endolysin Trx-SA1 might be an alternative treatment strategy for infections caused by S. aureus, including MRSA. Copyright © 2016 Elsevier B.V. All rights reserved.

Discussion of CoSA: Clustering of Sparse Approximations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Armstrong, Derek Elswick

2017-03-07

The purpose of this talk is to discuss the possible applications of CoSA (Clustering of Sparse Approximations) to the exploitation of HSI (HyperSpectral Imagery) data. CoSA is presented by Moody et al. in the Journal of Applied Remote Sensing (“Land cover classification in multispectral imagery using clustering of sparse approximations over learned feature dictionaries”, Vol. 8, 2014) and is based on machine learning techniques.

Antigenic variation: Molecular and genetic mechanisms of relapsing disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cruse, J.M.; Lewis, R.E.

1987-01-01

This book contains 10 chapters. They are: Contemporary Concepts of Antigenic Variation; Antigenic Variation in the Influenza Viruses; Mechanisms of Escape of Visna Lentiviruses from Immunological Control; A Review of Antigenic Variation by the Equine Infectious Anemia Virus; Biologic and Molecular Variations in AIDS Retrovirus Isolates; Rabies Virus Infection: Genetic Mutations and the Impact on Viral Pathogenicity and Immunity; Immunobiology of Relapsing Fever; Antigenic Variation in African Trypanosomes; Antigenic Variation and Antigenic Diversity in Malaria; and Mechanisms of Immune Evasion in Schistosomiasis.

Characterization of the Aspergillus nidulans aspnd1 gene demonstrates that the ASPND1 antigen, which it encodes, and several Aspergillus fumigatus immunodominant antigens belong to the same family.

PubMed Central

Calera, J A; Ovejero, M C; López-Medrano, R; Segurado, M; Puente, P; Leal, F

1997-01-01

For the first time, an immunodominant Aspergillus nidulans antigen (ASPND1) consistently reactive with serum samples from aspergilloma patients has been purified and characterized, and its coding gene (aspnd1) has been cloned and sequenced. ASPND1 is a glycoprotein with four N-glycosidically-bound sugar chains (around 2.1 kDa each) which are not necessary for reactivity with immune human sera. The polypeptide part is synthesized as a 277-amino-acid precursor of 30.6 kDa that after cleavage of a putative signal peptide of 16 amino acids, affords a mature protein of 261 amino acids with a molecular mass of 29 kDa and a pI of 4.24 (as deduced from the sequence). The ASPND1 protein is 53.1% identical to the AspfII allergen from Aspergillus fumigatus and 48% identical to an unpublished Candida albicans antigen. All of the cysteine residues and most of the glycosylation sites are perfectly conserved in the three proteins, suggesting a similar but yet unknown function. Analysis of the primary structure of the ASPND1 coding gene (aspnd1) has allowed the establishment of a clear relationship between several previously reported A. fumigatus and A. nidulans immunodominant antigens. PMID:9119471

Intellectual Property and Strategic Agreements (IP/SA) | Frederick National Laboratory for Cancer Research

Cancer.gov

What does IP/SA handle? IP/SA handles all invention issues including patents and copyrights. All employee inventionreports are filed through the IP/SA office for all activities under the OTS contract.Additionally,&nbs

Differential Recognition and Hydrolysis of Host Carbohydrate Antigens by Streptococcus pneumoniae Family 98 Glycoside Hydrolases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Higgins, M.; Whitworth, G; El Warry, N

2009-01-01

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

Sampling and sensitivity analyses tools (SaSAT) for computational modelling

PubMed Central

Hoare, Alexander; Regan, David G; Wilson, David P

2008-01-01

SaSAT (Sampling and Sensitivity Analysis Tools) is a user-friendly software package for applying uncertainty and sensitivity analyses to mathematical and computational models of arbitrary complexity and context. The toolbox is built in Matlab®, a numerical mathematical software package, and utilises algorithms contained in the Matlab® Statistics Toolbox. However, Matlab® is not required to use SaSAT as the software package is provided as an executable file with all the necessary supplementary files. The SaSAT package is also designed to work seamlessly with Microsoft Excel but no functionality is forfeited if that software is not available. A comprehensive suite of tools is provided to enable the following tasks to be easily performed: efficient and equitable sampling of parameter space by various methodologies; calculation of correlation coefficients; regression analysis; factor prioritisation; and graphical output of results, including response surfaces, tornado plots, and scatterplots. Use of SaSAT is exemplified by application to a simple epidemic model. To our knowledge, a number of the methods available in SaSAT for performing sensitivity analyses have not previously been used in epidemiological modelling and their usefulness in this context is demonstrated. PMID:18304361

Specific Fluorine Labeling of the HyHEL10 Antibody Affects Antigen Binding and Dynamics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Acchione, Mauro; Lee, Yi-Chien; DeSantis, Morgan E.

To more fully understand the molecular mechanisms responsible for variations in binding affinity with antibody maturation, we explored the use of site specific fluorine labeling and {sup 19}F nuclear magnetic resonance (NMR). Several single-chain (scFv) antibodies, derived from an affinity-matured series of anti-hen egg white lysozyme (HEL) mouse IgG1, were constructed with either complete or individual replacement of tryptophan residues with 5-fluorotryptophan ({sup 5F}W). An array of biophysical techniques was used to gain insight into the impact of fluorine substitution on the overall protein structure and antigen binding. SPR measurements indicated that {sup 5F}W incorporation lowered binding affinity for themore » HEL antigen. The degree of analogue impact was residue-dependent, and the greatest decrease in affinity was observed when {sup 5F}W was substituted for residues near the binding interface. In contrast, corresponding crystal structures in complex with HEL were essentially indistinguishable from the unsubstituted antibody. {sup 19}F NMR analysis showed severe overlap of signals in the free fluorinated protein that was resolved upon binding to antigen, suggesting very distinct chemical environments for each {sup 5F}W in the complex. Preliminary relaxation analysis suggested the presence of chemical exchange in the antibody-antigen complex that could not be observed by X-ray crystallography. These data demonstrate that fluorine NMR can be an extremely useful tool for discerning structural changes in scFv antibody-antigen complexes with altered function that may not be discernible by other biophysical techniques.« less

Identification of the antigenic determinants of factors 8, 9, and 34 of genus Candida.

PubMed

Kobayashi, H; Oyamada, H; Suzuki, A; Shibata, N; Suzuki, S; Okawa, Y

1996-10-21

We investigated the antigenic determinants of factors 8, 9, and 34 of the genus Candida among pathogenic yeasts by enzyme-linked immunosorbent assay (ELISA) using mannans of Saccharomyces cerevisiae wild type and mutant types, mnn 1-mnn 4 and mnn 2. Results of ELISA including antisera against the antigenic factors of genus Candida (Candida Check, latron; FAbs) indicated that these three types of mannan distinctly react with FAbs 34, 8 and 9, respectively. To identify the recognition sites of these FAbs, we compared the ability of various oligosaccharides to inhibit the binding of the mannans to FAbs. The results indicated that FAb 34 preferentially recognizes linear side chains containing a non-reducing terminal alpha-1,3-linked mannose residue, Man(alpha)1 --> 3Man(alpha)1 --> (2Man(alpha)1 --> )n(2Man) (n > or = 0), and that one of the recognition sites of FAb 9 is linear alpha-1,6-linked oligomannosyl series, Man(alpha)1 --> (6Man(alpha)1 --> )n(6Man) (n > or = 2). On the other hand, the recognition site of FAb 8 apparently consisted of two alpha-1,2-linked oligomannosyl side chains and an alpha-1,6-linked mannose residue that originated from the mannan backbone, Man(alpha)1 --> 2Man(alpha)1 --> 2(Man(alpha)1 -->2Man(alpha)1 --> 6)Man.

The Association Between Broad Antigen HLA Mismatches, Eplet HLA Mismatches and Acute Rejection After Kidney Transplantation.

PubMed

Do Nguyen, Hung Thanh; Wong, Germaine; Chapman, Jeremy R; McDonald, Stephen P; Coates, Patrick T; Watson, Narelle; Russ, Graeme R; D'Orsogna, Lloyd; Lim, Wai Hon

2016-12-01

Epitope matching, which evaluates mismatched amino acids within antigen-antibody interaction sites (eplets), may better predict acute rejection than broad antigen matching alone. We aimed to determine the association between eplet mismatches and acute rejection in kidney transplant recipients. The association between eplet mismatches, broad antigen mismatches and acute rejection was assessed using adjusted Cox proportional hazard regression. Model discrimination for acute rejection was evaluated using the area under receiver operating characteristic curves. Of the 3,499 kidney transplant recipients from 2006 to 2011, the average (SD) number of broad antigen and eplet mismatches were 3.4 (1.7) and 22.8 (12.2), respectively. Compared with 0 to 2 eplet mismatches, the adjusted hazard ratio (HR) for acute rejection among those with 20 or greater eplet mismatches was 2.16 (95% confidence interval [CI], 1.33-3.52; P = 0.001). The adjusted area under the curve for broad antigen mismatches was 0.58 (95% CI, 0.56-0.61), similar to that for eplet mismatches (HR, 0.59; 95% CI, 0.56-0.61; P = 0.365). In recipients who were considered as low immunological risk (0-2 broad antigen HLA-ABDR mismatch), those with 20 or greater eplet mismatches experienced an increased risk of rejection compared to those with less than 20 mismatches (adjusted HR, 1.85; 95% CI, 1.11-3.08; P = 0.019). Increasing number of eplet mismatches is associated with acute rejection in kidney transplant recipients. Consideration of eplet HLA mismatches may improve risk stratification for acute rejection in a selected group of kidney transplant candidates.

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

PubMed Central

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

1993-01-01

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

Dustborne Alternaria alternata antigens in U.S. homes: Results from the National Survey of Lead and Allergens in Housing

PubMed Central

Salo, Päivi M.; Yin, Ming; Arbes, Samuel J.; Cohn, Richard D.; Sever, Michelle; Muilenberg, Michael; Burge, Harriet A.; London, Stephanie J.; Zeldin, Darryl C.

2005-01-01

Background: Alternaria alternata is one of the most common fungi associated with allergic disease. However, Alternaria exposure in indoor environments is not well characterized. Objective: The primary goals of this study were to examine the prevalence of Alternaria exposure and identify independent predictors of Alternaria antigen concentrations in U.S. homes. Methods: Data for this cross-sectional study were obtained from the National Survey of Lead and Allergens in Housing. A nationally representative sample of 831 housing units in 75 different locations throughout the U.S. completed the survey. Information on housing and household characteristics was obtained by questionnaire and environmental assessments. Concentrations of Alternaria antigens in dust collected from various indoor sites were assessed with a polyclonal anti-Alternaria antibody assay. Results: Alternaria antigens were detected in most (95-99%) of the dust samples. The geometric mean concentration, reflecting the average Alternaria concentration in homes, was 4.88 μg/g (SE=0.13 μg/g). In the multivariable linear regression analysis, the age of the housing unit, geographic region, urbanization, poverty, family race, observed mold and moisture problems, use of dehumidifier, and presence of cats and dogs were independent predictors of Alternaria antigen concentrations. Less frequent cleaning and smoking indoors also contributed to higher Alternaria antigen levels in homes. Conclusion: Exposure to Alternaria alternata antigens in U.S. homes is common. Antigen levels in homes are not only influenced by regional factors but also by residential characteristics. Preventing mold and moisture problems, avoiding smoking indoors, and regular household cleaning may help reduce exposure to Alternaria antigens indoors. PMID:16159634

Antigen-mediated regulation in monoclonal gammopathies and myeloma

PubMed Central

Nair, Shiny; Sng, Joel; Boddupalli, Chandra Sekhar; Seckinger, Anja; Fulciniti, Mariateresa; Zhang, Lin; Rauniyar, Navin; Lopez, Michael; Neparidze, Natalia; Parker, Terri; Munshi, Nikhil C.; Sexton, Rachael; Barlogie, Bart; Orlowski, Robert; Bergsagel, Leif; Hose, Dirk; Mistry, Pramod K.; Meffre, Eric; Dhodapkar, Madhav V.

2018-01-01

A role for antigen-driven stimulation has been proposed in the pathogenesis of monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) based largely on the binding properties of monoclonal Ig. However, insights into antigen binding to clonal B cell receptors and in vivo responsiveness of the malignant clone to antigen-mediated stimulation are needed to understand the role of antigenic stimulation in tumor growth. Lysolipid-reactive clonal Ig were detected in Gaucher disease (GD) and some sporadic gammopathies. Here, we show that recombinant Ig (rIg) cloned from sort-purified single tumor cells from lipid-reactive sporadic and GD-associated gammopathy specifically bound lysolipids. Liposome sedimentation and binding assays confirmed specific interaction of lipid-reactive monoclonal Ig with lysolipids. The clonal nature of lysolipid-binding Ig was validated by protein sequencing. Gene expression profiling and cytogenetic analyses from 2 patient cohorts showed enrichment of nonhyperdiploid tumors in lipid-reactive patients. In vivo antigen-mediated stimulation led to an increase in clonal Ig and plasma cells (PCs) in GD gammopathy and also reactivated previously suppressed antigenically related nonclonal PCs. These data support a model wherein antigenic stimulation mediates an initial polyclonal phase, followed by evolution of monoclonal tumors enriched in nonhyperdiploid genomes, responsive to underlying antigen. Targeting underlying antigens may therefore prevent clinical MM. PMID:29669929

Skin-Resident T Cells Drive Dermal Dendritic Cell Migration in Response to Tissue Self-Antigen.

PubMed

Ali, Niwa; Zirak, Bahar; Truong, Hong-An; Maurano, Megan M; Gratz, Iris K; Abbas, Abul K; Rosenblum, Michael D

2018-05-01

Migratory dendritic cell (DC) subsets deliver tissue Ags to draining lymph nodes (DLNs) to either initiate or inhibit T cell-mediated immune responses. The signals mediating DC migration in response to tissue self-antigen are largely unknown. Using a mouse model of inducible skin-specific self-antigen expression, we demonstrate that CD103 + dermal DCs (DDCs) rapidly migrate from skin to skin DLN (SDLNs) within the first 48 h after Ag expression. This window of time was characterized by the preferential activation of tissue-resident Ag-specific effector T cells (Teffs), with no concurrent activation of Ag-specific Teffs in SDLNs. Using genetic deletion and adoptive transfer approaches, we show that activation of skin-resident Teffs is required to drive CD103 + DDC migration in response to tissue self-antigen and this Batf3-dependent DC population is necessary to mount a fulminant autoimmune response in skin. Conversely, activation of Ag-specific Teffs in SDLNs played no role in DDC migration. Our studies reveal a crucial role for skin-resident T cell-derived signals, originating at the site of self-antigen expression, to drive DDC migration during the elicitation phase of an autoimmune response. Copyright © 2018 by The American Association of Immunologists, Inc.

Antigen Masking During Fixation and Embedding, Dissected

PubMed Central

Scalia, Carla Rossana; Boi, Giovanna; Bolognesi, Maddalena Maria; Riva, Lorella; Manzoni, Marco; DeSmedt, Linde; Bosisio, Francesca Maria; Ronchi, Susanna; Leone, Biagio Eugenio; Cattoretti, Giorgio

2016-01-01

Antigen masking in routinely processed tissue is a poorly understood process caused by multiple factors. We sought to dissect the effect on antigenicity of each step of processing by using frozen sections as proxies of the whole tissue. An equivalent extent of antigen masking occurs across variable fixation times at room temperature. Most antigens benefit from longer fixation times (>24 hr) for optimal detection after antigen retrieval (AR; for example, Ki-67, bcl-2, ER). The transfer to a graded alcohol series results in an enhanced staining effect, reproduced by treating the sections with detergents, possibly because of a better access of the polymeric immunohistochemical detection system to tissue structures. A second round of masking occurs upon entering the clearing agent, mostly at the paraffin embedding step. This may depend on the non-freezable water removal. AR fully reverses the masking due both to the fixation time and the paraffin embedding. AR itself destroys some epitopes which do not survive routine processing. Processed frozen sections are a tool to investigate fixation and processing requirements for antigens in routine specimens. PMID:27798289

Peanut oral immunotherapy results in increased antigen-induced regulatory T-cell function and hypomethylation of forkhead box protein 3 (FOXP3).

PubMed

Syed, Aleena; Garcia, Marco A; Lyu, Shu-Chen; Bucayu, Robert; Kohli, Arunima; Ishida, Satoru; Berglund, Jelena P; Tsai, Mindy; Maecker, Holden; O'Riordan, Gerri; Galli, Stephen J; Nadeau, Kari C

2014-02-01

The mechanisms contributing to clinical immune tolerance remain incompletely understood. This study provides evidence for specific immune mechanisms that are associated with a model of operationally defined clinical tolerance. Our overall objective was to study laboratory changes associated with clinical immune tolerance in antigen-induced T cells, basophils, and antibodies in subjects undergoing oral immunotherapy (OIT) for peanut allergy. In a phase 1 single-site study, we studied participants (n = 23) undergoing peanut OIT and compared them with age-matched allergic control subjects (n = 20) undergoing standard of care (abstaining from peanut) for 24 months. Participants were operationally defined as clinically immune tolerant (IT) if they had no detectable allergic reactions to a peanut oral food challenge after 3 months of therapy withdrawal (IT, n = 7), whereas those who had an allergic reaction were categorized as nontolerant (NT; n = 13). Antibody and basophil activation measurements did not statistically differentiate between NT versus IT participants. However, T-cell function and demethylation of forkhead box protein 3 (FOXP3) CpG sites in antigen-induced regulatory T cells were significantly different between IT versus NT participants. When IT participants were withdrawn from peanut therapy for an additional 3 months (total of 6 months), only 3 participants remained classified as IT participants, and 4 participants regained sensitivity along with increased methylation of FOXP3 CpG sites in antigen-induced regulatory T cells. In summary, modifications at the DNA level of antigen-induced T-cell subsets might be predictive of a state of operationally defined clinical immune tolerance during peanut OIT. Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

Antigenic Competition Between and Endotoxic Adjuvant and a Protein Antigen

PubMed Central

Leong, Daniel L. Y.; Rudbach, Jon A.

1971-01-01

Antigenic competition between bovine gamma globulin (BGG) and endotoxin from a smooth strain (S-ET) and a rough (R-ET) heptoseless mutant strain of Salmonella minnesota was studied in mice. Both endotoxins acted as adjuvants for enhancing the antibody response to BGG. However, other work showed that the R-ET had minimal antigenicity, and it was used as a control for the competition studies. Antigenic competition between BGG and endotoxin as expressed by a suppression of the antibody response to BGG could not be demonstrated when varying adjuvant doses of S-ET or R-ET were injected simultaneously with a small constant dose of BGG into normal mice. However, mice presensitized with S-ET several weeks before immunization with the S-ET and BGG combination produced anti-BGG levels which were four to eightfold lower than in normal mice. Nearly complete suppression of the anti-BGG response could be obtained in presensitized mice by reducing the BGG dose 10-fold or by increasing the adjuvant dose of endotoxin. Mice pretreated with R-ET and challenged with BGG plus S-ET or R-ET showed no depression of the anti-BGG response. These and other experiments confirmed the immunological basis of the competitive effect. PMID:16557970

JAL (RH48) blood group antigen: serologic observations

PubMed Central

Lomas-Francis, Christine; Alcantara, Denden; Westhoff, Connie; Uehlinger, Joan; Valvasori, Marilia; Castilho, Lillian; Reid, Marion E.

2009-01-01

BACKGROUND JAL (RH48) is a low-prevalence antigen in the Rh blood group system and anti-JAL has caused hemolytic disease of the newborn. JAL is associated with either a haplotype carrying depressed C and e antigens or one carrying depressed c and e antigens. Blood samples from JAL+ people were tested, published serologic findings were confirmed, serologic studies were extended to include expression of other Rh antigens, and the antibody specificities produced by three sensitized JAL+ probands are reported. STUDY DESIGN AND METHODS Red blood cell (RBC) samples from 17 (12 probands) JAL+ persons were tested by hemagglutination using standard methods. RESULTS RBCs from both the Caucasian JAL+ probands had the (C)(e) haplotype and weakened C, e, hrB, and hrS antigens. JAL+ samples from black persons had the (c)(e) haplotype and expressed weakened c, e, f, V, VS, hrB, and hrS antigens. Plasma from three sensitized c+e+ JAL+ probands contained alloanti-c, alloanti-e, or alloantibody of apparent anti-Rh17 specificity. This study shows that this alloanti-Rh17–like antibody recognizes the high-prevalence antigen antithetical to JAL that has been named CEST. CONCLUSIONS The presence of the JAL antigen has a quantitative (weakening) effect on the expression of C, e, hrB, and hrS antigens in Caucasian persons and of c, e, f, V, VS, hrB, and hrS antigens in people of black African ancestry. A qualitative effect also was demonstrated by the presence of alloanti-c or alloanti-e in the plasma of two transfused c+e+ patients and by an antibody (anti-CEST) that recognizes the high-prevalence antigen antithetical to JAL. PMID:19192256

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

PubMed

Verma, Anita; Ngundi, Miriam M; Price, Gregory A; Takeda, Kazuyo; Yu, James; Burns, Drusilla L

2018-02-27

Toxin neutralizing antibodies represent the major mode of protective immunity against a number of toxin-mediated bacterial diseases, including anthrax; however, the cellular mechanisms that lead to optimal neutralizing antibody responses remain ill defined. Here we show that the cellular binding pathway of anthrax protective antigen (PA), the binding component of anthrax toxin, determines the toxin neutralizing antibody response to this antigen. PA, which binds cellular receptors and efficiently enters antigen-presenting cells by receptor-mediated endocytosis, was found to elicit robust anti-PA IgG and toxin neutralizing antibody responses. In contrast, a receptor binding-deficient mutant of PA, which does not bind receptors and only inefficiently enters antigen-presenting cells by macropinocytosis, elicited very poor antibody responses. A chimeric protein consisting of the receptor binding-deficient PA mutant tethered to the binding subunit of cholera toxin, which efficiently enters cells using the cholera toxin receptor rather than the PA receptor, elicited an anti-PA IgG antibody response similar to that elicited by wild-type PA; however, the chimeric protein elicited a poor toxin neutralizing antibody response. Taken together, our results demonstrate that the antigen capture pathway can dictate the magnitudes of the total IgG and toxin neutralizing antibody responses to PA as well as the ratio of the two responses. IMPORTANCE Neutralizing antibodies provide protection against a number of toxin-mediated bacterial diseases by inhibiting toxin action. Therefore, many bacterial vaccines are designed to induce a toxin neutralizing antibody response. We have used protective antigen (PA), the binding component of anthrax toxin, as a model antigen to investigate immune mechanisms important for the induction of robust toxin neutralizing antibody responses. We found that the pathway used by antigen-presenting cells to capture PA dictates the robustness of the

Plasmid and surface antigen markers of endemic and epidemic Legionella pneumophila strains.

PubMed Central

Brown, A; Vickers, R M; Elder, E M; Lema, M; Garrity, G M

1982-01-01

Environmental and clinical isolates of Legionella pneumophila obtained from the Pittsburgh Veterans Administration Medical Center were studied for the presence of plasmids and for unique surface antigens. The majority of environmental isolates contained a single 80-megadalton plasmid. After an epidemic of nosocomial Legionnaires disease subsided in the Spring of 1981, plasmid-bearing environmental isolates persisted in the environment. Whereas L. pneumophila could not be reisolated from most sites with plasmidless isolates. During this epidemic the attack rate was highest on wards with plasmidless isolates. All clinical isolates were plasmidless. Strains were serotyped by the indirect immunofluorescence method with serum from a single immunized rat which was used both without absorption and after absorption with various plasmid-bearing and plasmidless isolates. These studies suggested that a plasmid-associated surface antigen was present and that the most common plasmidless environmental serotype was similar to the epidemic clinical serotype. Images PMID:7119096

Mutations in yeast proliferating cell nuclear antigen define distinct sites for interaction with DNA polymerase delta and DNA polymerase epsilon.

PubMed Central

Eissenberg, J C; Ayyagari, R; Gomes, X V; Burgers, P M

1997-01-01

The importance of the interdomain connector loop and of the carboxy-terminal domain of Saccharomyces cerevisiae proliferating cell nuclear antigen (PCNA) for functional interaction with DNA polymerases delta (Poldelta) and epsilon (Pol epsilon) was investigated by site-directed mutagenesis. Two alleles, pol30-79 (IL126,128AA) in the interdomain connector loop and pol30-90 (PK252,253AA) near the carboxy terminus, caused growth defects and elevated sensitivity to DNA-damaging agents. These two mutants also had elevated rates of spontaneous mutations. The mutator phenotype of pol30-90 was due to partially defective mismatch repair in the mutant. In vitro, the mutant PCNAs showed defects in DNA synthesis. Interestingly, the pol30-79 mutant PCNA (pcna-79) was most defective in replication with Poldelta, whereas pcna-90 was defective in replication with Pol epsilon. Protein-protein interaction studies showed that pcna-79 and pcna-90 failed to interact with Pol delta and Pol epsilon, respectively. In addition, pcna-90 was defective in interaction with the FEN-1 endo-exonuclease (RTH1 product). A loss of interaction between pcna-79 and the smallest subunit of Poldelta, the POL32 gene product, implicates this interaction in the observed defect with the polymerase. Neither PCNA mutant showed a defect in the interaction with replication factor C or in loading by this complex. Processivity of DNA synthesis by the mutant holoenzyme containing pcna-79 was unaffected on poly(dA) x oligo(dT) but was dramatically reduced on a natural template with secondary structure. A stem-loop structure with a 20-bp stem formed a virtually complete block for the holoenzyme containing pcna-79 but posed only a minor pause site for wild-type holoenzyme, indicating a function of the POL32 gene product in allowing replication past structural blocks. PMID:9343398

Zitkala-Sa and the Problem of Regionalism: Nations, Narratives, and Critical Traditions

ERIC Educational Resources Information Center

Totten, Gary

2005-01-01

Although Yankton Sioux writer Zitkala-Sa (Gertrude Bonnin, 1876-1938) was, as P. Jane Hafen notes, "virtually unknown for many decades," much critical work has appeared since Dexter Fisher's 1979 article,"Zitkala-Sa: Evolution of a Writer." Some critics desiring to bring Zitkala-Sa into the conversation about turn-of-the-century American women…

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

PubMed

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

2012-04-01

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

Prognostic significance of 5T4 oncofetal antigen expression in colorectal carcinoma.

PubMed Central

Starzynska, T.; Marsh, P. J.; Schofield, P. F.; Roberts, S. A.; Myers, K. A.; Stern, P. L.

1994-01-01

The 5T4 oncofetal antigen is a 72 kDa glycoprotein defined by a monoclonal antibody raised against human placental trophoblast and is expressed in many different carcinomas but detected only at low levels in some normal epithelia. Immunohistochemical analysis of the patterns of expression in colorectal carcinomas has indicated a significant association between the presence of the antigen in tumour cells and metastatic spread. The 5T4 antigen phenotype of 72 colorectal cancers has been compared with the clinical outcome of the patients in order to assess its relationship with prognosis. Forty per cent of tumours were 5T4 positive; the remainder were either unlabelled or exhibited stroma-associated labelling only. There was a significant correlation between 5T4 expression in the malignant cells and unfavourable course of disease (P < 0.001). The 5 year survival with 5T4-positive tumours was 22% compared with 75% for patients with 5T4-negative tumours; median survival was 24 versus > 90 months respectively. Stratified analysis showed that 5T4 antigen tumour positivity was acting independently of each of stage, site of tumour, age or sex. There were significant differences in survival for patients with Dukes' B and C stage carcinomas (P = 0.001 and P = 0.034). The results suggest that in colorectal cancer immunohistochemical assessment of 5T4 expression may be useful in identifying patients at high risk for tumour recurrence and for whom additional treatment strategies might be most appropriate. Images Figure 1 PMID:8180020

Antigenic value of lyophilized phenolized antirabies vaccine.

PubMed

VEERARAGHAVAN, N; SUBRAHMANYAN, T P

1961-01-01

The authors present the results of experiments, carried out at the Pasteur Institute of Southern India, Coonoor, in which various preparations of lyophilized and liquid phenolized antirabies vaccines were assessed for antigenicity in relation to the NIH (United States National Institutes of Health) Reference Vaccine 164 (the proposed International Reference Preparation of Rabies Vaccine). The claim that phenolized antirabies vaccines can be lyophilized without loss of antigenicity was fully substantiated: the lyophilized vaccines were found to possess high antigenic values and to retain their antigenicity better than the liquid vaccines during storage under the same conditions.

Antigenic value of lyophilized phenolized antirabies vaccine

PubMed Central

Veeraraghavan, N.; Subrahmanyan, T. P.

1961-01-01

The authors present the results of experiments, carried out at the Pasteur Institute of Southern India, Coonoor, in which various preparations of lyophilized and liquid phenolized antirabies vaccines were assessed for antigenicity in relation to the NIH (United States National Institutes of Health) Reference Vaccine 164 (the proposed International Reference Preparation of Rabies Vaccine). The claim that phenolized antirabies vaccines can be lyophilized without loss of antigenicity was fully substantiated: the lyophilized vaccines were found to possess high antigenic values and to retain their antigenicity better than the liquid vaccines during storage under the same conditions. PMID:13925168

78 FR 11978 - Airworthiness Directives; Reims Aviation S.A. Airplanes

Federal Register 2010, 2011, 2012, 2013, 2014

2013-02-21

... DEPARTMENT OF TRANSPORTATION Federal Aviation Administration 14 CFR Part 39 [Docket No. FAA-2012... Airworthiness Directives; Reims Aviation S.A. Airplanes AGENCY: Federal Aviation Administration (FAA... directive (AD) for Reims Aviation S.A. Model F406 airplanes. This AD results from mandatory continuing...

A Novel Method Linking Antigen Presentation by Human Monocyte-Derived Macrophages to CD8+ T Cell Polyfunctionality

PubMed Central

Short, Kirsty R.; Grant, Emma J.; Vissers, Marloes; Reading, Patrick C.; Diavatopoulos, Dimitri A.; Kedzierska, Katherine

2013-01-01

To understand the interactions between innate and adaptive immunity, and specifically how virally infected macrophages impact T cell function, novel assays examining the ability of macrophages to present antigen to CD8+ T cells are needed. In the present study, we have developed a robust in vitro assay to measure how antigen presentation by human monocyte-derived macrophages (MDMs) affects the functional capacity of autologous CD8+ T cells. The assay is based on the polyfunctional characteristics of antigen-specific CD8+ T cells, and is thus called a Mac-CD8 Polyfunctionality Assay. Following purification of monocytes and their maturation to MDMs, MDMs were pulsed with an antigenic peptide to be presented to CD8+ T cells. Peptide-pulsed MDMs were then incubated with antigen-specific CD8+ T cells in order to assess the efficacy of antigen presentation to T cells. CD8+ T cell polyfunctionality was assessed by staining with mAbs to IFN-γ, TNF-α, and CD107a in a multi-color intracellular cytokine staining assay. To highlight the utility of the Mac-CD8 Polyfunctionality Assay, we assessed the effects of influenza infection on the ability of human macrophages to present antigen to CD8+ T cells. We found that influenza infection of human MDMs can alter the effector efficacy of MDMs to activate more CD8+ T cells with cytotoxic capacity. This has important implications for understanding how the virus-infected macrophages affect adaptive immunity at the site of infection. PMID:24312096

Genetic variants related to disease susceptibility and immunotolerance in the Duffy antigen receptor for chemokines (DARC, Fy) gene in the black lion tamarin (Leontopithecus chrysopygus, primates).

PubMed

Ansel, Ashley; Lewis, James D; Melnick, Don J; Martins, Cristiana; Valladares-Padua, Claudio; Perez-Sweeney, Beatriz

2017-10-01

The DARC (Duffy antigen receptor for chemokines) gene encodes the DARC protein, which serves multiple roles in the immune system, as a binding site for the malarial parasites Plasmodium vivax and Plasmodium knowlesi, a promiscuous chemokine receptor and a blood group antigen. Variation in DARC may play particularly significant roles in innate immunity, immunotolerance and pathogen entry in callitrichines, such as the black lion tamarin (Leontopithecus chrysopygus). We compared amino acid sequences of DARC in the black lion tamarin (BLT) to non-human Haplorhine primates and Homo sapiens. Consistent with prior studies in other Haplorhines, we observed that the chemokine receptor experiences two opposing selection forces: (1) positive selection on the Plasmodium binding site and (2) purifying selection. We observed also that D21N, F22L, and V25L differentiated BLT from humans at a critical site for P. vivax and P. knowlesi binding. One amino acid residue, F22L, was subject to both positive selection and fixation in New World monkeys, suggesting a beneficial role as an adaptive barrier to Plasmodium entry. Unlike in humans, we observed no variation in DARC among BLTs, suggesting that the protein does not play a role in immunotolerance. In addition, lion tamarins differed from humans at the blood compatibility Fy a /Fy b antigen-binding site 44, as well as at the putative destabilizing residues A61, T68, A187, and L215, further supporting a difference in the functional role of DARC in these primates compared with humans. Further research is needed to determine whether changes in the Plasmodium and Fy a /Fy b antigen-binding sites disrupt DARC function in callitrichines. © 2017 Wiley Periodicals, Inc.

The increased flexibility of CDR loops generated in antibodies by Congo red complexation favors antigen binding.

PubMed

Krol, Marcin; Roterman, Irena; Drozd, Anna; Konieczny, Leszek; Piekarska, Barbara; Rybarska, Janina; Spolnik, Paweł; Stopa, Barbara

2006-02-01

The dye Congo red and related self-assembling compounds were found to stabilize immune complexes by binding to antibodies currently engaged in complexation to antigen. In our simulations, it was shown that the site that becomes accessible for binding the supramolecular dye ligand is located in the V domain, and is normally occupied by the N-terminal polypeptide chain fragment. The binding of the ligand disrupts the beta-structure in the domain, increasing the plasticity of the antigen-binding site. The higher fluctuation of CDR-bearing loops enhances antigen binding, and allows even low-affinity antibodies to be engaged in immune complexes. Experimental observations of the enhancement effect were supported by theoretical studies using L lambda chain (4BJL-PDB identification) and the L chain from the complex of IgM-rheumatoid factor bound to the CH3 domain of the Fc fragment (1ADQ-PDB identification) as the initial structures for theoretical studies of dye-induced changes. Commercial IgM-type rheumatoid factor (human) and sheep red blood cells with coupled IgG (human) were used for experimental tests aimed to reveal the dye-enhancement effect in this system. The specificity of antigen-antibody interaction enhanced by dye binding was studied using rabbit anti-sheep red cell antibodies to agglutinate red cells of different species. Red blood cells of hoofed mammals (horse, goat) showed weak enhancement of agglutination in the presence of Congo red. Neither agglutination nor enhancement were observed in the case of human red cells. The dye-enhancement capability in the SRBC-antiSRBC system was lost after pepsin-digestion of antibodies producing (Fab)2 fragments still agglutinating red cells. Monoclonal (myeloma) IgG, L lambda chain and ovoalbumin failed to agglutinate red cells, as expected, and showed no enhancement effect. This indicates that the enhancement effect is specific.

Antigen Loss Variants: Catching Hold of Escaping Foes.

PubMed

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

2017-01-01

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

B-cell acquisition of antigen: Sensing the surface.

PubMed

Knight, Andrew M

2015-06-01

B-cell antigen receptor (BCR) recognition and acquisition of antigen by B cells is the essential first step in the generation of effective antibody responses. As B-cell-mediated antigen presentation is also believed to play a significant role in the activation of CD4(+) Th-cell responses, considerable effort has focused on clarifying the nature of antigen/BCR interactions. Following earlier descriptions of interactions of soluble antigens with the BCR, it is now clear that B cells also recognize, physically extract and present antigens that are tethered to, or integral components of, the surfaces or extracellular matrix of other cells. In this issue of the European Journal of Immunology, Zeng et al. [Eur. J. Immunol. 2015. 45: XXXX-XXXX] examine how the physical property or "stiffness" of the surface displaying antigens to B cells influences the B-cell response. This commentary reports that antigen tethered on "less stiff" surfaces induces increased B-cell activation and antibody responses. I then infer how "sensing the surface" by B cells may represent a new component of the immune system's ability to detect "damage," and how this understanding may influence approaches to clinical therapies where immune activity is either unwanted or desired. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The expression of hematopoietic progenitor cell antigen CD34 is regulated by DNA methylation in a site-dependent manner in gastrointestinal stromal tumours.

PubMed

Bure, Irina; Braun, Alexander; Kayser, Claudia; Geddert, Helene; Schaefer, Inga-Marie; Cameron, Silke; Ghadimi, Michael B; Ströbel, Philipp; Werner, Martin; Hartmann, Arndt; Wiemann, Stefan; Agaimy, Abbas; Haller, Florian; Moskalev, Evgeny A

2017-12-01

The anatomic site-dependent expression of hematopoietic progenitor cell antigen CD34 is a feature of gastrointestinal stromal tumours (GISTs). The basis for the differential CD34 expression is only incompletely understood. This study aimed at understanding the regulation of CD34 in GISTs and clarification of its site-dependent expression. Two sample sets of primary GISTs were interrogated including 52 fresh-frozen and 134 paraffin-embedded and formalin-fixed specimens. DNA methylation analysis was performed by HumanMethylation450 BeadChip array in three cell lines derived from gastric and intestinal GISTs, and differentially methylated CpG sites were established upstream of CD34. The methylation degree was further quantified by pyrosequencing, and inverse correlation with CD34 mRNA and protein abundance was revealed. The gene's expression could be activated upon induction of DNA hypomethylation with 5-aza-2'-deoxycytidine in GIST-T1 cells. In patient samples, a strong inverse correlation of DNA methylation degree with immunohistochemically evaluated CD34 expression was documented. Both CD34 expression and DNA methylation levels were specific to the tumours' anatomic location and mutation status. A constant decrease in methylation levels was observed ranging from almost 100% hypermethylation in intestinal GISTs from duodenum to hypomethylation in rectum. CD34 was heavily methylated in gastric PDGFRA-mutant GISTs in comparison to hypomethylated KIT-mutant counterparts. Next to CD34 hypermethylation, miR-665 was predicted and experimentally confirmed to target CD34 mRNA in GIST-T1 cells. Our results suggest that CD34 expression in GISTs may undergo a complex control by DNA methylation and miR-665. Differential methylation and expression of CD34 in GISTs along the gastrointestinal tract axis and in tumours that harbour different gain-of-function mutations suggest the origin from different cell populations in the gastrointestinal tract. © 2017 UICC.

Nonclassical T Cells and Their Antigens in Tuberculosis

PubMed Central

De Libero, Gennaro; Singhal, Amit; Lepore, Marco; Mori, Lucia

2014-01-01

T cells that recognize nonpeptidic antigens, and thereby are identified as nonclassical, represent important yet poorly characterized effectors of the immune response. They are present in large numbers in circulating blood and tissues and are as abundant as T cells recognizing peptide antigens. Nonclassical T cells exert multiple functions including immunoregulation, tumor control, and protection against infections. They recognize complexes of nonpeptidic antigens such as lipid and glycolipid molecules, vitamin B2 precursors, and phosphorylated metabolites of the mevalonate pathway. Each of these antigens is presented by antigen-presenting molecules other than major histocompatibility complex (MHC), including CD1, MHC class I–related molecule 1 (MR1), and butyrophilin 3A1 (BTN3A1) molecules. Here, we discuss how nonclassical T cells participate in the recognition of mycobacterial antigens and in the mycobacterial-specific immune response. PMID:25059739

Conservation of myeloid surface antigens on primate granulocytes.

PubMed

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

1983-02-01

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

Antigen Cross-Presentation of Immune Complexes

PubMed Central

Platzer, Barbara; Stout, Madeleine; Fiebiger, Edda

2014-01-01

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

Polymer blend particles with defined compositions for targeting antigen to both class I and II antigen presentation pathways

PubMed Central

Tran, Kenny K.; Zhan, Xi; Shen, Hong

2013-01-01

Defense against many persistent and difficult-to-treat diseases requires a combination of humoral, CD4+ and CD8+ T cell responses, which necessitates targeting antigens to both class I and II antigen presentation pathways. In this study, we developed polymer blend particles by mixing two functionally unique polymers, poly(lactide-co-glycolide) (PLGA) and a pH-responsive polymer, poly(dimethylaminoethyl methacrylate-co-propylacrylic acid-co-butyl methacrylate) (DMAEMA-co-PAA-co-BMA). We showed polymer blend particles enabled the delivery of antigens into both class I and II antigen presentation pathways in vitro. Increasing the ratio of the pH-responsive polymer in blend particles increased the degree of class I antigen presentation, while maintaining high levels of class II antigen presentation. In a mouse model, we demonstrated that a significantly higher and sustained level of CD4+ and CD8+ T cell responses, and comparable antibody responses, were elicited with polymer blend particles than PLGA particles and a conventional vaccine, Alum. The polymer blend particles offer a potential vaccine delivery platform to generate a combination of humoral and cell-mediated immune responses that insure robust and long-lasting immunity against many infectious diseases and cancers. PMID:24124123

Antigenic Structure of Rabbit γ Globulin

PubMed Central

Dubiski, S.; Dubiska, Anna; Skalba, Danuta; Kelus, A.

1961-01-01

By iso-immunization, antisera to five rabbit γ globulin antigens were obtained. They are called A (former Da), B, C, D and E. Individual sera of 670 rabbits belonging to six separate populations were tested by precipitation methods. The distribution of the iso-antigens and their combinations into serum groups were studied. Each particular γ globulin iso-antigen was found to be of hereditary character; they seem to form three genetic systems: A, C and BDE, statistically independent. Various antisera from England, Poland and U.S.A were compared. PMID:13724581

Phase Variable O Antigen Biosynthetic Genes Control Expression of the Major Protective Antigen and Bacteriophage Receptor in Vibrio cholerae O1

PubMed Central

Seed, Kimberley D.; Faruque, Shah M.; Mekalanos, John J.; Calderwood, Stephen B.; Qadri, Firdausi; Camilli, Andrew

2012-01-01

The Vibrio cholerae lipopolysaccharide O1 antigen is a major target of bacteriophages and the human immune system and is of critical importance for vaccine design. We used an O1-specific lytic bacteriophage as a tool to probe the capacity of V. cholerae to alter its O1 antigen and identified a novel mechanism by which this organism can modulate O antigen expression and exhibit intra-strain heterogeneity. We identified two phase variable genes required for O1 antigen biosynthesis, manA and wbeL. manA resides outside of the previously recognized O1 antigen biosynthetic locus, and encodes for a phosphomannose isomerase critical for the initial step in O1 antigen biosynthesis. We determined that manA and wbeL phase variants are attenuated for virulence, providing functional evidence to further support the critical role of the O1 antigen for infectivity. We provide the first report of phase variation modulating O1 antigen expression in V. cholerae, and show that the maintenance of these phase variable loci is an important means by which this facultative pathogen can generate the diverse subpopulations of cells needed for infecting the host intestinal tract and for escaping predation by an O1-specific phage. PMID:23028317

Expression of a cloned lipopolysaccharide antigen from Neisseria gonorrhoeae on the surface of Escherichia coli K-12.

PubMed Central

Palermo, D A; Evans, T M; Clark, V L

1987-01-01

A gonococcal gene bank maintained in Escherichia coli K-12 was screened by colony immunoblotting, and a transformant expressing a surface antigen reactive to anti-gonococcal outer membrane antiserum was isolated. The isolate carried a recombinant plasmid, pTME6, consisting of approximately 9 kilobases of Neisseria gonorrhoeae DNA inserted into the BamHI site of pBR322. Surface labeling of E. coli HB101(pTME6) confirmed that the antigen was expressed on the E. coli cell surface. The antigenic material was resistant to proteinase K digestion and sensitive to periodate oxidation, indicating that the material was carbohydrate. Purified lipopolysaccharide (LPS) from HB101(pTME6) produced a unique band on silver-stained polyacrylamide gels that contained immunoreactive material as seen on Western blots of LPS samples. Only two of three E. coli LPS mutant strains carrying pTME6 reacted with the antigonococcal antiserum, suggesting that a certain E. coli core structure is necessary for antigen expression. We conclude that pTME6 contains one or more gonococcal genes encoding an LPS core biosynthetic enzyme(s) which can modify E. coli core LPS to produce a gonococcuslike epitope(s). Images PMID:3117695

Understanding original antigenic sin in influenza with a dynamical system.

PubMed

Pan, Keyao

2011-01-01

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

Immunogenicity of 60 novel latency-related antigens of Mycobacterium tuberculosis

PubMed Central

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

2014-01-01

The aim of our work here was to evaluate the immunogenicity of 60 mycobacterial antigens, some of which have not been previously assessed, notably a novel series of in vivo-expressed Mycobacterium tuberculosis (IVE-TB) antigens. We enrolled 505 subjects and separated them in individuals with and without latent tuberculosis infection (LTBI) vs. patients with active tuberculosis (TB). Following an overnight and 7 days stimulation of whole blood with purified recombinant M. tuberculosis antigens, interferon-γ (IFN-γ) levels were determined by ELISA. Several antigens could statistically significantly differentiate the groups of individuals. We obtained promising antigens from all studied antigen groups [dormancy survival regulon (DosR regulon) encoded antigens; resuscitation-promoting factors (Rpf) antigens; IVE-TB antigens; reactivation associated antigens]. Rv1733, which is a probable conserved transmembrane protein encoded in DosR regulon, turned out to be very immunogenic and able to discriminate between the three defined TB status, thus considered a candidate biomarker. Rv2389 and Rv2435n, belonging to Rpf family and IVE-TB group of antigens, respectively, also stood out as LTBI biomarkers. Although more studies are needed to support our findings, the combined use of these antigens would be an interesting approach to TB immunodiagnosis candidates. PMID:25339944

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

PubMed

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

2014-01-01

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

Broadly neutralizing antibodies from human survivors target a conserved site in the Ebola virus glycoprotein HR2-MPER region.

PubMed

Flyak, Andrew I; Kuzmina, Natalia; Murin, Charles D; Bryan, Christopher; Davidson, Edgar; Gilchuk, Pavlo; Gulka, Christopher P; Ilinykh, Philipp A; Shen, Xiaoli; Huang, Kai; Ramanathan, Palaniappan; Turner, Hannah; Fusco, Marnie L; Lampley, Rebecca; Kose, Nurgun; King, Hannah; Sapparapu, Gopal; Doranz, Benjamin J; Ksiazek, Thomas G; Wright, David W; Saphire, Erica Ollmann; Ward, Andrew B; Bukreyev, Alexander; Crowe, James E

2018-05-07

Ebola virus (EBOV) in humans causes a severe illness with high mortality rates. Several strategies have been developed in the past to treat EBOV infection, including the antibody cocktail ZMapp, which has been shown to be effective in nonhuman primate models of infection 1 and has been used under compassionate-treatment protocols in humans 2 . ZMapp is a mixture of three chimerized murine monoclonal antibodies (mAbs) 3-6 that target EBOV-specific epitopes on the surface glycoprotein 7,8 . However, ZMapp mAbs do not neutralize other species from the genus Ebolavirus, such as Bundibugyo virus (BDBV), Reston virus (RESTV) or Sudan virus (SUDV). Here, we describe three naturally occurring human cross-neutralizing mAbs, from BDBV survivors, that target an antigenic site in the canonical heptad repeat 2 (HR2) region near the membrane-proximal external region (MPER) of the glycoprotein. The identification of a conserved neutralizing antigenic site in the glycoprotein suggests that these mAbs could be used to design universal antibody therapeutics against diverse ebolavirus species. Furthermore, we found that immunization with a peptide comprising the HR2-MPER antigenic site elicits neutralizing antibodies in rabbits. Structural features determined by conserved residues in the antigenic site described here could inform an epitope-based vaccine design against infection caused by diverse ebolavirus species.

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

PubMed

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

2016-08-01

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

MyD88/CD40 Genetic Adjuvant Function in Cutaneous Atypical Antigen-Presenting Cells Contributes to DNA Vaccine Immunogenicity

PubMed Central

Slawin, Kevin M.; Levitt, Jonathan M.; Spencer, David M.

2016-01-01

Therapeutic DNA-based vaccines aim to prime an adaptive host immune response against tumor-associated antigens, eliminating cancer cells primarily through CD8+ cytotoxic T cell-mediated destruction. To be optimally effective, immunological adjuvants are required for the activation of tumor-specific CD8+ T cells responses by DNA vaccination. Here, we describe enhanced anti-tumor efficacy of an in vivo electroporation-delivered DNA vaccine by inclusion of a genetically encoded chimeric MyD88/CD40 (MC) adjuvant, which integrates both innate and adaptive immune signaling pathways. When incorporated into a DNA vaccine, signaling by the MC adjuvant increased antigen-specific CD8+ T cells and promoted elimination of pre-established tumors. Interestingly, MC-enhanced vaccine efficacy did not require direct-expression of either antigen or adjuvant by local antigen-presenting cells, but rather our data supports a key role for MC function in “atypical” antigen-presenting cells of skin. In particular, MC adjuvant-modified keratinocytes increased inflammatory cytokine secretion, upregulated surface MHC class I, and were able to increase in vitro and in vivo priming of antigen-specific CD8+ T cells. Furthermore, in the absence of critical CD8α+/CD103+ cross-priming dendritic cells, MC was still able to promote immune priming in vivo, albeit at a reduced level. Altogether, our data support a mechanism by which MC signaling activates an inflammatory phenotype in atypical antigen-presenting cells within the cutaneous vaccination site, leading to an enhanced CD8+ T cell response against DNA vaccine-encoded antigens, through both CD8α+/CD103+ dendritic cell-dependent and independent pathways. PMID:27741278

Two separable functional domains of simian virus 40 large T antigen: carboxyl-terminal region of simian virus 40 large T antigen is required for efficient capsid protein synthesis.

PubMed Central

Tornow, J; Polvino-Bodnar, M; Santangelo, G; Cole, C N

1985-01-01

The carboxyl-terminal portion of simian virus 40 large T antigen is essential for productive infection of CV-1 and CV-1p green monkey kidney cells. Mutant dlA2459, lacking 14 base pairs at 0.193 map units, was positive for viral DNA replication, but unable to form plaques in CV-1p cells (J. Tornow and C.N. Cole, J. Virol. 47:487-494, 1983). In this report, the defect of dlA2459 is further defined. Simian virus 40 late mRNAs were transcribed, polyadenylated, spliced, and transported in dlA2459-infected cells, but the level of capsid proteins produced in infected CV-1 green monkey kidney cells was extremely low. dlA2459 large T antigen lacks those residues known to be required for adenovirus helper function, and the block to productive infection by dlA2459 occurs at the same stage of infection as the block to productive adenovirus infection of CV-1 cells. These results suggest that the adenovirus helper function is required for productive infection by simian virus 40. Mutant dlA2459 was able to grow on the Vero and BSC-1 lines of African green monkey kidney cells. Additional mutants affecting the carboxyl-terminal portion of large T were prepared. Mutant inv2408 contains an inversion of the DNA between the BamHI and BclI sites (0.144 to 0.189 map units). This inversion causes transposition of the carboxyl-terminal 26 amino acids of large T antigen and the carboxyl-terminal 18 amino acids of VP1. This mutant was viable, even though the essential information absent from dlA2459 large T antigen has been transferred to the carboxyl terminus of VP1 of inv2408. The VP1 polypeptide carrying this carboxyl-terminal portion of large T could overcome the defect of dlA2459. This indicates that the carboxyl terminus of large T antigen is a separate and separable functional domain. Images PMID:2982029

The subcutaneous inoculation of pH 6 antigen mutants of Yersinia pestis does not affect virulence and immune response in mice.

PubMed

Anisimov, Andrey P; Bakhteeva, Irina V; Panfertsev, Evgeniy A; Svetoch, Tat'yana E; Kravchenko, Tat'yana B; Platonov, Mikhail E; Titareva, Galina M; Kombarova, Tat'yana I; Ivanov, Sergey A; Rakin, Alexander V; Amoako, Kingsley K; Dentovskaya, Svetlana V

2009-01-01

Two isogenic sets of Yersinia pestis strains were generated, composed of wild-type strains 231 and I-1996, their non-polar pH 6(-) mutants with deletions in the psaA gene that codes for its structural subunit or the whole operon, as well as strains with restored ability for temperature- and pH-dependent synthesis of adhesion pili or constitutive production of pH 6 antigen. The mutants were generated by site-directed mutagenesis of the psa operon and subsequent complementation in trans. It was shown that the loss of synthesis or constitutive production of pH 6 antigen did not influence Y. pestis virulence or the average survival time of subcutaneously inoculated BALB/c naïve mice or animals immunized with this antigen.

Protective antigens from El Tor vibrios

PubMed Central

Watanabe, Yoshikazu; Verwey, W. F.

1965-01-01

A biochemically and immunologically homogeneous antigenic fraction having the properties of a lipopolysaccharide has been isolated from the culture supernatant of an El Tor vibrio (Ogawa subtype). This antigen was very specifically protective for mice challenged with Ogawa strains of either El Tor vibrios or Vibrio cholerae. Rabbit antisera prepared against the antigen were passively protective for mice and highly vibriocidal but had little agglutinating activity. However, the antigen was able specifically to absorb agglutinins, as well as mouse-protective and vibriocidal antibody from serum prepared against whole bacterial cells. The specific protective activity of this lipopolysaccharide was much greater than that of vaccines made from whole bacterial cells, and its toxicity in animals was about equivalent to that of whole cells. The relationship of activity to toxicity therefore represented an improvement over the vaccines that were studied. ImagesFIG. 1FIG. 3FIG. 4FIG. 5 PMID:5294306

Analysis of density and epitopes of D antigen on the surface of erythrocytes from DEL phenotypic individuals carrying the RHD1227A allele.

PubMed

Gu, Juan; Sun, An-Yuan; Wang, Xue-Dong; Shao, Chao-Peng; Li, Zheng; Huang, Li-Hua; Pan, Zhao-Lin; Wang, Qing-Ping; Sun, Guang-Ming

2014-04-01

The characteristics of the D antigen are important as they influence the immunogenicity of D variant cells. Several studies on antigenic sites have been reported in normal D positive, weak D and partial D cases, including a comprehensive analysis of DEL types in Caucasians. The aim of this study was to assess D antigen density and epitopes on the erythrocyte surface of Asian type DEL phenotypic individuals carrying the RHD1227A allele in the Chinese population. A total of 154 DEL phenotypic individuals carrying the RHD1227A allele were identified through adsorption and elution tests and polymerase chain reaction analysis with sequence-specific primers in the Chinese population. D antigen density on the erythrocyte surface of these individuals was detected using a flow cytometric method. An erythrocyte sample with known D antigen density was used as a standard. Blood samples from D-negative and D-positive individuals were used as controls. In addition, D antigen epitopes on the erythrocyte surface of DEL individuals carrying the RHD1227A allele were investigated with 18 monoclonal anti-D antibodies specific for different D antigen epitopes. The means of the median fluorescence intensity of D antigen on the erythrocyte membrane surface of D-negative, D-positive and DEL individuals were 2.14±0.25, 193.61±11.43 and 2.45±0.82, respectively. The DEL samples were estimated to have approximately 22 D antigens per cell. The samples from all 154 DEL individuals reacted positively with 18 monoclonal anti-D antibodies specific for different D antigen epitopes. In this study, D antigen density on the erythrocyte surface of DEL individuals carrying the RHD1227A allele was extremely low, there being only very few antigenic molecules per cell, but the D antigen epitopes were grossly complete.

Screening Immunomodulators To Skew the Antigen-Specific Autoimmune Response.

PubMed

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

2017-01-03

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

Modeling the impact of hydraulic redistribution on the carbon flux and storages using CLM4.5 at four AmeriFlux Sites

NASA Astrophysics Data System (ADS)

Fu, C.; Wang, G.; Cardon, Z. G.

2015-12-01

Effects of hydraulic redistribution (HR) on the hydrological cycle and ecosystem dynamics have been demonstrated in the field, but few modeling studies have compared HR's influences on the carbon cycle in different ecosystems and climate regions. The soil moisture changes associated with HR could influence plant carbon gain via two mechanisms: (1) improved plant water status supporting stomatal opening, and/or (2) enhanced nutrient availability to plants caused by enhanced soil microbial activity. In this study, using a modified version of the Community Land Model with Century-based soil carbon pool kinetics that includes the "Ryel et al. 2002" scheme for hydraulic redistribution (HR), the influence of HR on the carbon flux and storage is investigated at four Ameriflux sites where HR was detected from soil moisture measurements. The study sites include a Douglas-fir site (US-Wrc) in Washington State with a mediterranean climate, a savanna site (US-SRM) in Arizona with a semi-arid climate, an oak/pine forest site (US-SCf) in Southern California with a mediterranean climate, and an evergreen broadleaf forest site (BR-Sa1) with tropical monsoon climate. Simulations revealed that HR tended to enhance plant growth at all four sites, and incorporating HR into CLM4.5 reduces the temporal fluctuation of soil carbon storage at all four sites. Simulations with HR can capture the net carbon exchange between ecosystem and the atmosphere (NEE) at the US-Wrc, US-SRM, and BR-Sa1 sites over the annual cycle. Incorporation of HR into CLM4.5 clearly improved the weekly and sub-daily NEE simulation during dry periods at US-SCf and BR-Sa1 site. HR-induced increase in Net Primary Productivity (NPP) at the US-Wrc and US-SRM sites was driven approximately equally by the two distinct mechanisms we investigated: increased stomatal conductance and increased nutrient availability to plants.

Avidity of anti-malarial antibodies inversely related to transmission intensity at three sites in Uganda.

PubMed

Ssewanyana, Isaac; Arinaitwe, Emmanuel; Nankabirwa, Joaniter I; Yeka, Adoke; Sullivan, Richard; Kamya, Moses R; Rosenthal, Philip J; Dorsey, Grant; Mayanja-Kizza, Harriet; Drakeley, Chris; Greenhouse, Bryan; Tetteh, Kevin K A

2017-02-10

People living in malaria endemic areas acquire protection from severe malaria quickly, but protection from clinical disease and control of parasitaemia is acquired only after many years of repeated infections. Antibodies play a central role in protection from clinical disease; however, protective antibodies are slow to develop. This study sought to investigate the influence of Plasmodium falciparum exposure on the acquisition of high-avidity antibodies to P. falciparum antigens, which may be associated with protection. Cross-sectional surveys were performed in children and adults at three sites in Uganda with varied P. falciparum transmission intensity (entomological inoculation rates; 3.8, 26.6, and 125 infectious bites per person per year). Sandwich ELISA was used to measure antibody responses to two P. falciparum merozoite surface antigens: merozoite surface protein 1-19 (MSP1-19) and apical membrane antigen 1 (AMA1). In individuals with detectable antibody levels, guanidine hydrochloride (GuHCl) was added to measure the relative avidity of antibody responses by ELISA. Within a site, there were no significant differences in median antibody levels between the three age groups. Between sites, median antibody levels were generally higher in the higher transmission sites, with differences more apparent for AMA-1 and in ≥5 year group. Similarly, median avidity index (proportion of high avidity antibodies) showed no significant increase with increasing age but was significantly lower at sites of higher transmission amongst participants ≥5 years of age. Using 5 M GuHCl, the median avidity indices in the ≥5 year group at the highest and lowest transmission sites were 19.9 and 26.8, respectively (p = 0.0002) for MSP1-19 and 12.2 and 17.2 (p = 0.0007) for AMA1. Avidity to two different P. falciparum antigens was lower in areas of high transmission intensity compared to areas with lower transmission. Appreciation of the mechanisms behind these findings as

γδ T cells recognize a microbial encoded B cell antigen to initiate a rapid antigen specific Interleukin 17 response

PubMed Central

Zeng, Xun; Wei, Yu-ling; Huang, Jun; Newell, Evan W.; Yu, Hongxiang; Kidd, Brian A.; Kuhns, Michael S.; Waters, Ray W.; Davis, Mark M.; Weaver, Casey T.; Chien, Yueh-hsiu

2012-01-01

Summary γδ T cells contribute uniquely to host immune defense. However, how they function remains an enigma. Although it is unclear what most γδ T cells recognize, common dogma asserts that they recognize self-antigens. While they are the major initial Interleukin-17 (IL-17) producers in infections, it is unclear what is required to trigger these cells to act. Here, we report that a noted B cell antigen, the algae protein-phycoerythrin (PE) is an antigen for murine and human γδ T cells. PE also stained specific bovine γδ T cells. Employing this specificity, we demonstrated that antigen recognition, but not extensive clonal expansion, was required to activate naïve γδ T cells to make IL-17. In this activated state, γδ T cells gained the ability to respond to cytokine signals that perpetuated the IL-17 production. These results underscore the adaptability of lymphocyte antigen receptors and suggest a previously unrecognized antigen-driven rapid response in protective immunity prior to the maturation of classical adaptive immunity. PMID:22960222

Definition of neutralizing sites on African horse sickness virus serotype 4 VP2 at the level of peptides.

PubMed

Martínez-Torrecuadrada, J L; Langeveld, J P; Meloen, R H; Casal, J I

2001-10-01

The antigenic structure of African horse sickness virus (AHSV) serotype 4 capsid protein VP2 has been determined at the peptide level by PEPSCAN analysis in combination with a large collection of polyclonal antisera and monoclonal antibodies. VP2, the determinant for the virus serotype and an important target in virus neutralization, was found to contain 15 antigenic sites. A major antigenic region containing 12 of the 15 sites was identified in the region between residues 223 and 400. A second domain between residues 568 and 681 contained the three remaining sites. These sites were used for the synthesis of peptides, which were later tested in rabbits. Of the 15 synthetic peptides, three were able to induce neutralizing antibodies for AHSV-4, defining two neutralizing epitopes, 'a' and 'b', between residues 321 and 339, and 377 and 400, respectively. A combination of peptides representing both sites induced a more effective neutralizing response. Still, the relatively low neutralization titres make the possibility of producing a synthetic vaccine for AHSV unlikely. The complex protein-protein interaction of the outer shell of the viral capsid would probably require the presence of either synthetic peptides in the correct conformation or peptide segments from the different proteins VP2, VP5 and VP7.

Engineering Chimeric Antigen Receptors

PubMed Central

Kulemzin, S. V.; Kuznetsova, V. V.; Mamonkin, M.; Taranin, A. V.; Gorchakov, A. A.

2017-01-01

Chimeric antigen receptors (CARs) are recombinant protein molecules that redirect cytotoxic lymphocytes toward malignant and other target cells. The high feasibility of manufacturing CAR-modified lymphocytes for the therapy of cancer has spurred the development and optimization of new CAR T cells directed against a broad range of target antigens. In this review, we describe the main structural and functional elements constituting a CAR, discuss the roles of these elements in modulating the anti-tumor activity of CAR T cells, and highlight alternative approaches to CAR engineering. PMID:28461969

A novel Arg H52/Tyr H33 conservative motif in antibodies: A correlation between sequence of antibodies and antigen binding.

PubMed

Petrov, Artem; Arzhanik, Vladimir; Makarov, Gennady; Koliasnikov, Oleg

2016-08-01

Antibodies are the family of proteins, which are responsible for antigen recognition. The computational modeling of interaction between an antigen and an antibody is very important when crystallographic structure is unavailable. In this research, we have discovered the correlation between the amino acid sequence of antibody and its specific binding characteristics on the example of the novel conservative binding motif, which consists of four residues: Arg H52, Tyr H33, Thr H59, and Glu H61. These residues are specifically oriented in the binding site and interact with each other in a specific manner. The residues of the binding motif are involved in interaction strictly with negatively charged groups of antigens, and form a binding complex. Mechanism of interaction and characteristics of the complex were also discovered. The results of this research can be used to increase the accuracy of computational antibody-antigen interaction modeling and for post-modeling quality control of the modeled structures.

77 FR 6666 - Airworthiness Directives; CFM International, S.A. Turbofan Engines

Federal Register 2010, 2011, 2012, 2013, 2014

2012-02-09

... International, S.A. model CFM56-5B series turbofan engines. This AD was prompted by a normal quality sampling at... quality sampling at CFM International. S.A. that isolated a production batch of fan blades with...

Paraprofessional Education in SA DETAFE.

ERIC Educational Resources Information Center

Mills, Paul; Cesnich, Janine

A study examined the current outcomes and future of paraprofessional education provided under the auspices of South Australia's Department of Employment and Technical and Further Education (SA DETAFE). The primary objectives of the study were as follows: (1) determine whether an autonomous middle-level employment base exists in selected…

The Molecular Determinants of Antibody Recognition and Antigenic Drift in the H3 Hemagglutinin of Swine Influenza A Virus

PubMed Central

Abente, Eugenio J.; Santos, Jefferson; Lewis, Nicola S.; Gauger, Phillip C.; Stratton, Jered; Skepner, Eugene; Rajao, Daniela S.

2016-01-01

ABSTRACT Influenza A virus (IAV) of the H3 subtype is an important respiratory pathogen that affects both humans and swine. Vaccination to induce neutralizing antibodies against the surface glycoprotein hemagglutinin (HA) is the primary method used to control disease. However, due to antigenic drift, vaccine strains must be periodically updated. Six of the 7 positions previously identified in human seasonal H3 (positions 145, 155, 156, 158, 159, 189, and 193) were also indicated in swine H3 antigenic evolution. To experimentally test the effect on virus antigenicity of these 7 positions, substitutions were introduced into the HA of an isogenic swine lineage virus. We tested the antigenic effect of these introduced substitutions by using hemagglutination inhibition (HI) data with monovalent swine antisera and antigenic cartography to evaluate the antigenic phenotype of the mutant viruses. Combinations of substitutions within the antigenic motif caused significant changes in antigenicity. One virus mutant that varied at only two positions relative to the wild type had a >4-fold reduction in HI titers compared to homologous antisera. Potential changes in pathogenesis and transmission of the double mutant were evaluated in pigs. Although the double mutant had virus shedding titers and transmissibility comparable to those of the wild type, it caused a significantly lower percentage of lung lesions. Elucidating the antigenic effects of specific amino acid substitutions at these sites in swine H3 IAV has important implications for understanding IAV evolution within pigs as well as for improved vaccine development and control strategies in swine. IMPORTANCE A key component of influenza virus evolution is antigenic drift mediated by the accumulation of amino acid substitutions in the hemagglutinin (HA) protein, resulting in escape from prior immunity generated by natural infection or vaccination. Understanding which amino acid positions of the HA contribute to the ability

Structural and functional consequences of antigenic modulation of red blood cells with methoxypoly(ethylene glycol).

PubMed

Murad, K L; Mahany, K L; Brugnara, C; Kuypers, F A; Eaton, J W; Scott, M D

1999-03-15

We previously showed that the covalent modification of the red blood cell (RBC) surface with methoxypoly(ethylene glycol) [mPEG; MW approximately 5 kD] could significantly attenuate the immunologic recognition of surface antigens. However, to make these antigenically silent RBC a clinically viable option, the mPEG-modified RBC must maintain normal cellular structure and functions. To this end, mPEG-derivatization was found to have no significant detrimental effects on RBC structure or function at concentrations that effectively blocked antigenic recognition of a variety of RBC antigens. Importantly, RBC lysis, morphology, and hemoglobin oxidation state were unaffected by mPEG-modification. Furthermore, as shown by functional studies of Band 3, a major site of modification, PEG-binding does not affect protein function, as evidenced by normal SO4- flux. Similarly, Na+ and K+ homeostasis were unaffected. The functional aspects of the mPEG-modified RBC were also maintained, as evidenced by normal oxygen binding and cellular deformability. Perhaps most importantly, mPEG-derivatized mouse RBC showed normal in vivo survival ( approximately 50 days) with no sensitization after repeated transfusions. These data further support the hypothesis that the covalent attachment of nonimmunogenic materials (eg, mPEG) to intact RBC may have significant application in transfusion medicine, especially for the chronically transfused and/or allosensitized patient.

Revealing the glycation sites in synthetic neoglycoconjugates formed by conjugation of the antigenic monosaccharide hapten of Vibrio cholerae O1, serotype Ogawa with the BSA protein carrier using LC-ESI-QqTOF-MS/MS

PubMed Central

Jahouh, Farid; Saksena, Rina; Kováč, Pavol; Banoub, Joseph

2012-01-01

In this manuscript, we present the determination of glycation sites in synthetic neoglycoconjugates formed by conjugation of the antigenic monosaccharide hapten of Vibrio cholerae O1 serotype Ogawa to BSA using nano- liquid chromatography electrospray ionization quadrupole time-of-flight tandem mass spectroscopy (LC-ESI-QqTOF-MS/MS). The matrix-assisted laser desorption/ionization-TOF/TOF-MS/MS analyses of the tryptic digests of the glycoconjugates having a hapten:BSA ratio of 4.3:1, 6.6:1 and 13.2:1 revealed only three glycation sites, on the following lysine residues: Lys 235, Lys 437 and Lys 455. Digestion of the neoglycoconjugates with the proteases trypsin and GluC V8 gave complementary structural information and was shown to maximize the number of recognized glycation sites. Here, we report identification of 20, 27 and 33 glycation sites using LC-ESI-QqTOF-MS/MS analysis of a series of synthetic neoglycoconjugates with a hapten:BSA ratio of, respectively, 4.3:1, 6.6:1 and 13.2:1. We also tentatively propose that all the glycated lysine residues are located mainly near the outer surface of the protein. PMID:22791257

The pharmacokinetics of, and humoral responses to, antigen delivered by microencapsulated liposomes.

PubMed

Cohen, S; Bernstein, H; Hewes, C; Chow, M; Langer, R

1991-12-01

The feasibility of creating a s.c. depot for sustained protein delivery with the goal of enhancing antigen immunogenicity was investigated. The depot was designed as antigen-laden liposomes of hydrogenated egg phosphatidylcholine and cholesterol (1:1 molar ratio) encapsulated in alginate-poly(L-lysine) microcapsules and evaluated using iodinated bovine serum albumin (BSA) as a model antigen. The in vivo release behavior of the liposomes and microencapsulated liposomes (MELs) was evaluated from the BSA serum concentration profiles after s.c. injection into rats and the pharmacokinetic parameters of 125I-labeled BSA appearance after s.c. or i.v. injections of BSA in saline. Maximal BSA concentrations were detected 11 h after s.c. injection in all rats. The BSA serum concentrations decreased rapidly in rats injected with BSA in saline or Freund's adjuvant and less rapidly in rats injected with BSA in liposomes or MELs. Four to 5 weeks after injection, BSA-associated radioactivity was detected only in sera of rats injected with BSA in liposomes or MELs. Fifty days after injection, 50% of the originally injected BSA was recovered form the s.c. sites of rats injected with BSA in MELs; no radioactivity was recovered from the other three groups of rats. The antigen-reactive antibody levels induced in rats immunized with BSA in MELs were 2- to 3-fold higher than those obtained in rats immunized with BSA in liposomes, saline, or Freund's adjuvant. More significantly, high antibody levels were maintained for more than 150 days after a single injection of BSA in MELs, suggesting that MELs can serve as a long-term single-dose immunization vehicle.

[SciELO Public Health: the performance of Cadernos de Saúde Pública and Revista de Saúde Pública].

PubMed

Barata, Rita Barradas

2007-12-01

The aim of this paper was to analyze two Brazilian scientific journals included in the SciELO Library of Public Health, using a group of bibliometric indicators and scrutinizing the articles most viewed. Cadernos de Saúde Pública was accessed 3,743.59 times per month, with an average of 30.31 citations per article. The 50 articles most viewed (6.72 to 524.5 views) were mostly published in Portuguese (92%). 42% were theoretical essays, 20% surveys, and 16% descriptive studies. 42% used argumentative techniques, 34% quantitative techniques, 18% qualitative techniques, and 6% mathematical modeling. The most common themes were: health and work (50%), epidemiology (22%), and environmental health (8%). Revista de Saúde Pública was accessed 1,590.97 times per month, with an average of 26.27 citations per article. The 50 articles most viewed (7.33 and 56.50 views) were all published in Portuguese: 46% were surveys, 14% databases analysis, and 12% systematic reviews. Quantitative techniques were adopted in 66% of such articles, while mathematical modeling was the same as observed in Cadernos de Saúde Pública, as were qualitative techniques. The most common themes were health services organization (22%), nutrition (22%), health and work (18%), epidemiology (12%), and environmental health (12%).

EXTERIOR VIEW, THE SA WETSIDE BUILDING (ALSO KNOWN AS DISTILLATION ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

EXTERIOR VIEW, THE SA WETSIDE BUILDING (ALSO KNOWN AS DISTILLATION BUILDING OR SOLVAY TOWERS BUILDING) THE HEART OF THE SOLVAY PROCESS. VIEW LOOKING EAST. THIS BUILDING WAS BUILT IN 1954 TO REPLACE A SIMILAR STRUCTURE BUILT CIRCA 1907. - Solvay Process Company, SA Wetside Building, Between Willis & Milton Avenue, Solvay, Onondaga County, NY

77 FR 71491 - Airworthiness Directives; Turbomeca S.A. Turboshaft Engines

Federal Register 2010, 2011, 2012, 2013, 2014

2012-12-03

... Airworthiness Directives; Turbomeca S.A. Turboshaft Engines AGENCY: Federal Aviation Administration (FAA), DOT... Turbomeca S.A. Arriel 1E2, 1S, and 1S1 turboshaft engines. That AD currently requires a one-time inspection... 74 45 15. You may view this service information at the FAA, Engine & Propeller Directorate, 12 New...

78 FR 34284 - Airworthiness Directives; Turbomeca S.A. Turboshaft Engines

Federal Register 2010, 2011, 2012, 2013, 2014

2013-06-07

... Turbomeca S.A. Arriel 2D turboshaft engines. The existing AD currently requires replacing the..., MA. For information on the availability of this material at the FAA, call 781-238-7125. Examining the..., January 31, 2013), for all Turbomeca S.A. Arriel 2D turboshaft engines. That AD requires replacing the HMU...

Detection of peste des petits ruminants virus antigen using immunofiltration and antigen-competition ELISA methods.

PubMed

Raj, G Dhinakar; Rajanathan, T M C; Kumar, C Senthil; Ramathilagam, G; Hiremath, Geetha; Shaila, M S

2008-06-22

Peste des petits ruminants (PPR) is one of the most economically important diseases affecting sheep and goats in India. An immunofiltration-based test has been developed using either mono-specific serum/monoclonal antibodies (mAb) prepared against a recombinant truncated nucleocapsid protein of rinderpest virus (RPV) cross-reactive with PPR virus. This method consists of coating ocular swab eluate from suspected animals onto a nitrocellulose membrane housed in a plastic module, which is allowed to react with suitable dilutions of a mAb or a mono-specific polyclonal antibody. The antigen-antibody complex formed on the membrane is then detected by protein A-colloidal gold conjugate, which forms a pink colour. In the immunofiltration test, concordant results were obtained using either PPRV mAb or mono-specific serum. Another test, an antigen-competition ELISA which relies on the competition between plate-coated recombinant truncated 'N' protein of RPV and the PPRV 'N' protein present in ocular swab eluates (sample) for binding to the mono-specific antibody against N protein of RPV (in liquid phase) was developed. The cut-off value for this test was established using reverse transcription polymerase chain reaction (RT-PCR) positive and negative oculo-nasal swab samples. Linear correlation between percent inhibition (PI) values in antigen-competition ELISA and virus infectivity titres was 0.992. Comparison of the immunofiltration test with the antigen-competition ELISA yielded a sensitivity of 80% and specificity of 100%. These two tests can serve as a screening (immunofiltration) and confirmatory (antigen-competition ELISA) test, respectively, in the diagnosis of PPR in sheep or goats.

Genetic diversity and antigenicity variation of Babesia bovis merozoite surface antigen-1 (MSA-1) in Thailand.

PubMed

Tattiyapong, Muncharee; Sivakumar, Thillaiampalam; Takemae, Hitoshi; Simking, Pacharathon; Jittapalapong, Sathaporn; Igarashi, Ikuo; Yokoyama, Naoaki

2016-07-01

Babesia bovis, an intraerythrocytic protozoan parasite, causes severe clinical disease in cattle worldwide. The genetic diversity of parasite antigens often results in different immune profiles in infected animals, hindering efforts to develop immune control methodologies against the B. bovis infection. In this study, we analyzed the genetic diversity of the merozoite surface antigen-1 (msa-1) gene using 162 B. bovis-positive blood DNA samples sourced from cattle populations reared in different geographical regions of Thailand. The identity scores shared among 93 msa-1 gene sequences isolated by PCR amplification were 43.5-100%, and the similarity values among the translated amino acid sequences were 42.8-100%. Of 23 total clades detected in our phylogenetic analysis, Thai msa-1 gene sequences occurred in 18 clades; seven among them were composed of sequences exclusively from Thailand. To investigate differential antigenicity of isolated MSA-1 proteins, we expressed and purified eight recombinant MSA-1 (rMSA-1) proteins, including an rMSA-1 from B. bovis Texas (T2Bo) strain and seven rMSA-1 proteins based on the Thai msa-1 sequences. When these antigens were analyzed in a western blot assay, anti-T2Bo cattle serum strongly reacted with the rMSA-1 from T2Bo, as well as with three other rMSA-1 proteins that shared 54.9-68.4% sequence similarity with T2Bo MSA-1. In contrast, no or weak reactivity was observed for the remaining rMSA-1 proteins, which shared low sequence similarity (35.0-39.7%) with T2Bo MSA-1. While demonstrating the high genetic diversity of the B. bovis msa-1 gene in Thailand, the present findings suggest that the genetic diversity results in antigenicity variations among the MSA-1 antigens of B. bovis in Thailand. Copyright © 2016 Elsevier B.V. All rights reserved.

21 CFR 660.40 - Hepatitis B Surface Antigen.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 7 2011-04-01 2010-04-01 true 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...

21 CFR 660.40 - Hepatitis B Surface Antigen.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 7 2014-04-01 2014-04-01 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...

21 CFR 660.40 - Hepatitis B Surface Antigen.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 7 2013-04-01 2013-04-01 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...

21 CFR 660.40 - Hepatitis B Surface Antigen.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 7 2012-04-01 2012-04-01 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...

21 CFR 660.40 - Hepatitis B Surface Antigen.

Code of Federal Regulations, 2010 CFR

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...

Cancer-testis antigen expression is shared between epithelial ovarian cancer tumors.

PubMed

Garcia-Soto, Arlene E; Schreiber, Taylor; Strbo, Natasa; Ganjei-Azar, Parvin; Miao, Feng; Koru-Sengul, Tulay; Simpkins, Fiona; Nieves-Neira, Wilberto; Lucci, Joseph; Podack, Eckhard R

2017-06-01

Cancer-testis (CT) antigens have been proposed as potential targets for cancer immunotherapy. Our objective was to evaluate the expression of a panel of CT antigens in epithelial ovarian cancer (EOC) tumor specimens, and to determine if antigen sharing occurs between tumors. RNA was isolated from EOC tumor specimens, EOC cell lines and benign ovarian tissue specimens. Real time-PCR analysis was performed to determine the expression level of 20 CT antigens. A total of 62 EOC specimens, 8 ovarian cancer cell lines and 3 benign ovarian tissues were evaluated for CT antigen expression. The majority of the specimens were: high grade (62%), serous (68%) and advanced stage (74%). 58 (95%) of the EOC tumors analyzed expressed at least one of the CT antigens evaluated. The mean number of CT antigen expressed was 4.5 (0-17). The most frequently expressed CT antigen was MAGE A4 (65%). Antigen sharing analysis showed the following: 9 tumors shared only one antigen with 62% of the evaluated specimens, while 37 tumors shared 4 or more antigens with 82%. 5 tumors expressed over 10 CT antigens, which were shared with 90% of the tumor panel. CT antigens are expressed in 95% of EOC tumor specimens. However, not a single antigen was universally expressed across all samples. The degree of antigen sharing between tumors increased with the total number of antigens expressed. These data suggest a multi-epitope approach for development of immunotherapy for ovarian cancer treatment. Copyright © 2017 Elsevier Inc. All rights reserved.

Preclinical evaluation of a chemically detoxified pneumolysin as pneumococcal vaccine antigen.

PubMed

Hermand, Philippe; Vandercammen, Annick; Mertens, Emmanuel; Di Paolo, Emmanuel; Verlant, Vincent; Denoël, Philippe; Godfroid, Fabrice

2017-01-02

The use of protein antigens able to protect against the majority of Streptococcus pneumoniae serotypes is envisaged as stand-alone and/or complement to the current capsular polysaccharide-based pneumococcal vaccines. Pneumolysin (Ply) is a key virulence factor that is highly conserved in amino acid sequence across pneumococcal serotypes, and therefore may be considered as a vaccine target. However, native Ply cannot be used in vaccines due to its intrinsic cytolytic activity. In the present work a completely, irreversibly detoxified pneumolysin (dPly) has been generated using an optimized formaldehyde treatment. Detoxi-fication was confirmed by dPly challenge in mice and histological analysis of the injection site in rats. Immunization with dPly elicited Ply-specific functional antibodies that were able to inhibit Ply activity in a hemolysis assay. In addition, immunization with dPly protected mice against lethal intranasal challenge with Ply, and intranasal immunization inhibited nasopharyngeal colonization after intranasal challenge with homologous or heterologous pneumococcal strain. Our findings supported dPly as a valid candidate antigen for further pneumococcal vaccine development.

Preclinical evaluation of a chemically detoxified pneumolysin as pneumococcal vaccine antigen

PubMed Central

Hermand, Philippe; Vandercammen, Annick; Mertens, Emmanuel; Di Paolo, Emmanuel; Verlant, Vincent; Denoël, Philippe; Godfroid, Fabrice

2017-01-01

ABSTRACT The use of protein antigens able to protect against the majority of Streptococcus pneumoniae serotypes is envisaged as stand-alone and/or complement to the current capsular polysaccharide-based pneumococcal vaccines. Pneumolysin (Ply) is a key virulence factor that is highly conserved in amino acid sesec-typsecquence across pneumococcal serotypes, and therefore may be considered as a vaccine target. However, native Ply cannot be used in vaccines due to its intrinsic cytolytic activity. In the present work a completely, irreversibly detoxified pneumolysin (dPly) has been generated using an optimized formaldehyde treatment. Detoxi-fication was confirmed by dPly challenge in mice and histological analysis of the injection site in rats. Immunization with dPly elicited Ply-specific functional antibodies that were able to inhibit Ply activity in a hemolysis assay. In addition, immunization with dPly protected mice against lethal intranasal challenge with Ply, and intranasal immunization inhibited nasopharyngeal colonization after intranasal challenge with homologous or heterologous pneumococcal strain. Our findings supported dPly as a valid candidate antigen for further pneumococcal vaccine development. PMID:27768518

Dynamic fracture toughness of ASME SA508 Class 2a ASME SA533 grade A Class 2 base and heat affected zone material and applicable weld metals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Logsdon, W.A.; Begley, J.A.; Gottshall, C.L.

1978-03-01

The ASME Boiler and Pressure Vessel Code, Section III, Article G-2000, requires that dynamic fracture toughness data be developed for materials with specified minimum yield strengths greater than 50 ksi to provide verification and utilization of the ASME specified minimum reference toughness K/sub IR/ curve. In order to qualify ASME SA508 Class 2a and ASME SA533 Grade A Class 2 pressure vessel steels (minimum yield strengths equal 65 kip/in./sup 2/ and 70 kip/in./sup 2/, respectively) per this requirement, dynamic fracture toughness tests were performed on these materials. All dynamic fracture toughness values of SA508 Class 2a base and HAZ material,more » SA533 Grade A Class 2 base and HAZ material, and applicable weld metals exceeded the ASME specified minimum reference toughness K/sub IR/ curve.« less

77 FR 58002 - Airworthiness Directives; Turbomeca S.A. Turboshaft Engines

Federal Register 2010, 2011, 2012, 2013, 2014

2012-09-19

... Airworthiness Directives; Turbomeca S.A. Turboshaft Engines AGENCY: Federal Aviation Administration (FAA), DOT... Turbomeca S.A. Arriel 2B and 2B1 turboshaft engines. That AD currently requires accomplishment of the TU166 modification. This new AD requires adding the Arriel 2S2 and 2C2 engines to the applicability of engines...

A role for mitochondria in antigen processing and presentation

PubMed Central

Bonifaz, Laura C; Cervantes-Silva, Mariana P; Ontiveros-Dotor, Elizabeth; López-Villegas, Edgar O; Sánchez-García, F Javier

2015-01-01

Immune synapse formation is critical for T-lymphocyte activation, and mitochondria have a role in this process, by localizing close to the immune synapse, regulating intracellular calcium concentration, and providing locally required ATP. The interaction between antigen-presenting cells (APCs) and T lymphocytes is a two-way signalling process. However, the role of mitochondria in APCs during this process remains unknown. For APCs to be able to activate T lymphocytes, they must first engage in an antigen-uptake, -processing and -presentation process. Here we show that hen egg white lysozyme (HEL) -loaded B lymphocytes, as a type of APC, undergo a small but significant mitochondrial depolarization by 1–2 hr following antigen exposure, suggesting an increase in their metabolic demands. Inhibition of ATP synthase (oligomycin) or mitochondrial Ca2+ uniporter (MCU) (Ruthenium red) had no effect on antigen uptake. Therefore, antigen processing and antigen presentation were further analysed. Oligomycin treatment reduced the amount of specific MHC–peptide complexes but not total MHC II on the cell membrane of B lymphocytes, which correlated with a decrease in antigen presentation. However, oligomycin also reduced antigen presentation by B lymphocytes, which endogenously express HEL and by B lymphocytes loaded with the HEL48–62 peptide, although to a lesser extent. ATP synthase inhibition and MCU inhibition had a clear inhibitory effect on antigen processing (DQ-OVA). Taken together these results suggest that ATP synthase and MCU are relevant for antigen processing and presentation. Finally, APC mitochondria were found to re-organize towards the APC–T immune synapse. PMID:25251370

Characterization of an antigenic site that contains a dominant, type-specific neutralization determinant on the envelope protein domain III (ED3) of dengue 2 virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gromowski, Gregory D.; Barrett, Alan D.T.

2007-09-30

The surface of the mature dengue virus (DENV) particle consists of 90 envelope (E) protein dimers that mediate both receptor binding and fusion. The E protein ectodomain can be divided into three structural domains designated ED1, ED2, and ED3, of which ED3 contains the critical and dominant virus-specific neutralization sites. In this study the ED3 epitopes recognized by seven, murine, IgG1 DENV-2 type-specific, monoclonal antibodies (MAbs) were determined using site-directed mutagenesis of a recombinant DENV-2 ED3 (rED3) protein. A total of 41 single amino acid substitutions were introduced into the rED3 at 30 different surface accessible residues. The affinity ofmore » each MAb with the mutant rED3s was assessed by indirect ELISA and the results indicate that all seven MAbs recognize overlapping epitopes with residues K305 and P384 critical for binding. These residues are conserved among DENV-2 strains and cluster together on the upper lateral face of ED3. A linear relationship was observed between relative occupancy of ED3 on the virion by MAb and neutralization of the majority of virus infectivity ({approx} 90%) for all seven MAbs. Depending on the MAb, it is predicted that between 10% and 50% relative occupancy of ED3 on the virion is necessary for virus neutralization and for all seven MAbs occupancy levels approaching saturation were required for 100% neutralization of virus infectivity. Overall, the conserved antigenic site recognized by all seven MAbs is likely to be a dominant DENV-2 type-specific, neutralization determinant.« less

Antigenic relatedness of glucosyltransferase enzymes from streptococcus mutans.

PubMed

Smith, D J; Taubman, M A

1977-01-01

The antigenic relationship of glucosyltransferases (GTF) produced by different serotypes of Streptococcus mutans was studied by using a functional inhibition assay. Rat, rabbit, or hamster immune fluids, directed to cell-associated or supernatant-derived GTF, were tested against ammonium sulfate-precipitated culture supernatants containing GTF from seven strains of S. mutans representing six different serotypes. An antigenic relationship was shown to exist among GTF from serotypes a, d, and g, since both rat and rabbit antisera directed to serotype a or g GTF inhibited GTF of serotypes d and g similarly and both antisera also inhibited serotype a GTF. Furthermore, serum inhibition patterns indicated that GTF of serotypes c and e, and possibly b, are antigenically related to each other, but are antigenically distinct from GTF of serotype a, d, or g. Serum antibody directed to antigens other than enzyme (e.g., serotype-specific antigen or teichoic acid) had little effect on the inhibition assay. Salivas from rats immunized with cell-associated or supernatant-derived GTF exhibited low but consistent inhibition of GTF activity, which generally corresponded to the serum patterns. The sera of two groups of hamsters immunized with GTF (serotype g), enriched either in water-insoluble or water-soluble glucan synthetic activity, gave patterns of inhibition quite similar to those seen with sera from more heterogenous cell-associated or crude supernatant-derived GTF preparations. Both groups of hamster sera also gave virtually identical patterns, suggesting that the two enzyme forms used as antigen share common antigenic determinants. The results from the three animal models suggest that among the cariogenic organisms tested, two (serotypes a, d, g and b, c, e), or perhaps three (serotypes a, d, g; b; and c, e), different subsets of GTF exist that have distinct antigenic determinants within a subset.

Molecular and antigenic traits on hemagglutinin gene of avian influenza H9N2 viruses: Evidence of a new escape mutant in Egypt adapted in quails.

PubMed

Adel, Amany; Arafa, Abdelsatar; Hussein, Hussein A; El-Sanousi, Ahmed A

2017-06-01

The LPAI viruses of H9N2 subtype became widely distributed in Middle Eastern countries, causing great economic losses in poultry industry especially when complicated with other pathogens. The H9N2 viruses in Egypt have a wide spread nature since its first occurrence in 2011. In this study, we collected cloacal and tracheal samples from 19 flocks for detection and propagation of H9N2 virus using real-time RT-PCR and egg inoculation. We studied the molecular evolution of the Hemagglutinin gene of H9N2 viruses by full HA gene sequencing, then the antigenic characterization was implemented using the cross HI assay and analyzed using 3D Bioinformatics cartography software. The phylogenetic analysis of the HA gene of Egyptian H9N2 viruses clearly points out the presence of only one group (Egy/G1) of originally introduced viruses in 2011 related to the G1 lineage within group B, with the presence of multiple minor clusters includes viruses from 2011 to 2015. However, a new variant (Egy/G1var) cluster was detected in quails since 2012. Genetically, Egy/G1var viruses characterized by presence of 20 amino acid substitutions within and adjacent to the antigenic sites in comparison to other Egyptian viruses. In addition, two glycosylation sites at amino acid residues 127 and 189 were determined in close to the receptor binding and antigenic sites. The antigenic analysis based on 3D antigenic mapping showed that the Egy/G1var cluster was clearly distinct from the original Egy/G1 viruses. In conclusion, Egy/G1var is shown to be a new escape mutant variant cluster with an adaptive evolution in quails. Copyright © 2017 Elsevier Ltd. All rights reserved.

Light-chain residue 95 is critical for antigen binding and multispecificity of monoclonal antibody G2.

PubMed

Usui, Daiki; Inaba, Satomi; Kamatari, Yuji O; Ishiguro, Naotaka; Oda, Masayuki

2017-09-02

The monoclonal antibody, G2, specifically binds to the immunogen peptide derived from the chicken prion protein, Pep18mer, and two chicken proteins derived peptides, Pep8 and Pep395; G2 binds with equal affinity to Pep18mer. The amino acid sequences of the three peptides are completely different, and so the recognition mechanism of G2 is unique and interesting. We generated a single-chain Fv (scFv) antibody of G2, and demonstrated its correct folding with an antigen binding function similar to intact G2 antibody. We also generated a Pro containing mutant of G2 scFv at residue 95 of the light chain, and analyzed its antigen binding using a surface plasmon biosensor. The mutant lost its binding ability to Pep18mer, but remained those to Pep8 and Pep395. The results clearly indicate residue 95 as being critical for multispecific antigen binding of G2 at the site generated from the junctional diversity introduced at the joints between the V and J gene segments. Copyright © 2017 Elsevier Inc. All rights reserved.

Bayesian nonparametric clustering in phylogenetics: modeling antigenic evolution in influenza.

PubMed

Cybis, Gabriela B; Sinsheimer, Janet S; Bedford, Trevor; Rambaut, Andrew; Lemey, Philippe; Suchard, Marc A

2018-01-30

Influenza is responsible for up to 500,000 deaths every year, and antigenic variability represents much of its epidemiological burden. To visualize antigenic differences across many viral strains, antigenic cartography methods use multidimensional scaling on binding assay data to map influenza antigenicity onto a low-dimensional space. Analysis of such assay data ideally leads to natural clustering of influenza strains of similar antigenicity that correlate with sequence evolution. To understand the dynamics of these antigenic groups, we present a framework that jointly models genetic and antigenic evolution by combining multidimensional scaling of binding assay data, Bayesian phylogenetic machinery and nonparametric clustering methods. We propose a phylogenetic Chinese restaurant process that extends the current process to incorporate the phylogenetic dependency structure between strains in the modeling of antigenic clusters. With this method, we are able to use the genetic information to better understand the evolution of antigenicity throughout epidemics, as shown in applications of this model to H1N1 influenza. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

A viral, transporter associated with antigen processing (TAP)-independent, high affinity ligand with alternative interactions endogenously presented by the nonclassical human leukocyte antigen E class I molecule.

PubMed

Lorente, Elena; Infantes, Susana; Abia, David; Barnea, Eilon; Beer, Ilan; García, Ruth; Lasala, Fátima; Jiménez, Mercedes; Mir, Carmen; Morreale, Antonio; Admon, Arie; López, Daniel

2012-10-12

The transporter associated with antigen processing (TAP) enables the flow of viral peptides generated in the cytosol by the proteasome and other proteases to the endoplasmic reticulum, where they complex with nascent human leukocyte antigen (HLA) class I. Later, these peptide-HLA class I complexes can be recognized by CD8(+) lymphocytes. Cancerous cells and infected cells in which TAP is blocked, as well as individuals with unusable TAP complexes, are able to present peptides on HLA class I by generating them through TAP-independent processing pathways. Here, we identify a physiologically processed HLA-E ligand derived from the D8L protein in TAP-deficient vaccinia virus-infected cells. This natural high affinity HLA-E class I ligand uses alternative interactions to the anchor motifs previously described to be presented on nonclassical HLA class I molecules. This octameric peptide was also presented on HLA-Cw1 with similar binding affinity on both classical and nonclassical class I molecules. In addition, this viral peptide inhibits HLA-E-mediated cytolysis by natural killer cells. Comparison between the amino acid sequences of the presenting HLA-E and HLA-Cw1 alleles revealed a shared structural motif in both HLA class molecules, which could be related to their observed similar cross-reactivity affinities. This motif consists of several residues located on the floor of the peptide-binding site. These data expand the role of HLA-E as an antigen-presenting molecule.

Plasma membrane vesicles decorated with glycolipid-anchored antigens and adjuvants via protein transfer as an antigen delivery platform for inhibition of tumor growth.

PubMed

Patel, Jaina M; Vartabedian, Vincent F; Bozeman, Erica N; Caoyonan, Brianne E; Srivatsan, Sanjay; Pack, Christopher D; Dey, Paulami; D'Souza, Martin J; Yang, Lily; Selvaraj, Periasamy

2016-01-01

Antigen delivered within particulate materials leads to enhanced antigen-specific immunity compared to soluble administration of antigen. However, current delivery approaches for antigen encapsulated in synthetic particulate materials are limited by the complexity of particle production that affects stability and immunogenicity of the antigen. Herein, we describe a protein delivery system that utilizes plasma membrane vesicles (PMVs) derived from biological materials such as cultured cells or isolated tissues and a simple protein transfer technology. We show that these particulate PMVs can be easily modified within 4 h by a protein transfer process to stably incorporate a glycosylphosphatidylinositol (GPI)-anchored form of the breast cancer antigen HER-2 onto the PMV surface. Immunization of mice with GPI-HER-2-modified-PMVs induced strong HER-2-specific antibody responses and protection from tumor challenge in two different breast cancer models. Further incorporation of the immunostimulatory molecules IL-12 and B7-1 onto the PMVs by protein transfer enhanced tumor protection and induced beneficial Th1 and Th2-type HER-2-specific immune responses. Since protein antigens can be easily converted to GPI-anchored forms, these results demonstrate that isolated plasma membrane vesicles can be modified with desired antigens along with immunostimulatory molecules by protein transfer and used as a vaccine delivery vehicle to elicit potent antigen-specific immunity. Copyright © 2015 Elsevier Ltd. All rights reserved.

Ether modifications to 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(3-phenylpropyl)piperazine (SA4503): effects on binding affinity and selectivity for sigma receptors and monoamine transporters.

PubMed

Xu, Rong; Lord, Sarah A; Peterson, Ryan M; Fergason-Cantrell, Emily A; Lever, John R; Lever, Susan Z

2015-01-01

Two series of novel ether analogs of the sigma (σ) receptor ligand 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(3-phenylpropyl)piperazine (SA4503) have been prepared. In one series, the alkyl portion of the 4-methoxy group was replaced with allyl, propyl, bromoethyl, benzyl, phenethyl, and phenylpropyl moieties. In the second series, the 3,4-dimethoxy was replaced with cyclic methylenedioxy, ethylenedioxy and propylenedioxy groups. These ligands, along with 4-O-des-methyl SA4503, were evaluated for σ1 and σ2 receptor affinity, and compared to SA4503 and several known ether analogs. SA4503 and a subset of ether analogs were also evaluated for dopamine transporter (DAT) and serotonin transporter (SERT) affinity. The highest σ1 receptor affinities, Ki values of 1.75-4.63 nM, were observed for 4-O-des-methyl SA4503, SA4503 and the methylenedioxy analog. As steric bulk increased, σ1 receptor affinity decreased, but only to a point. Allyl, propyl and bromoethyl substitutions gave σ1 receptor Ki values in the 20-30 nM range, while bulkier analogs having phenylalkyl, and Z- and E-iodoallyl, ether substitutions showed higher σ1 affinities, with Ki values in the 13-21 nM range. Most ligands studied exhibited comparable σ1 and σ2 affinities, resulting in little to no subtype selectivity. SA4503, the fluoroethyl analog and the methylenedioxy congener showed modest six- to fourteen-fold selectivity for σ1 sites. DAT and SERT interactions proved much more sensitive than σ receptor interactions to these structural modifications. For example, the benzyl congener (σ1Ki=20.8 nM; σ2Ki=16.4 nM) showed over 100-fold higher DAT affinity (Ki=121 nM) and 6-fold higher SERT affinity (Ki=128nM) than the parent SA4503 (DAT Ki=12650 nM; SERT Ki=760 nM). Thus, ether modifications to the SA4503 scaffold can provide polyfunctional ligands having a broader spectrum of possible pharmacological actions. Copyright © 2014 Elsevier Ltd. All rights reserved.

Thermostability of the coating, antigen and immunostimulator in an adjuvanted oral capsule vaccine formulation.

PubMed

Longet, Stephanie; Aversa, Vincenzo; O'Donnell, Daire; Tobias, Joshua; Rosa, Monica; Holmgren, Jan; Coulter, Ivan S; Lavelle, Ed C

2017-12-20

Oral vaccines present an attractive alternative to injectable vaccines for enteric diseases due to ease of delivery and the induction of intestinal immunity at the site of infection. However, susceptibility to gastrointestinal proteolysis, limited transepithelial uptake and a lack of clinically acceptable adjuvants present significant challenges. A further challenge to mass vaccination in developing countries is the very expensive requirement to maintain the cold chain. We recently described the effectiveness of a Single Multiple Pill ® (SmPill ® ) adjuvanted capsule approach to enhance the effectiveness of a candidate enterotoxigenic Escherichia coli (ETEC) oral vaccine. Here it was demonstrated that this delivery system maintains the antigenicity of ETEC colonisation factor antigen I (CFA/I) and the immunostimulatory activity of the orally active α-Galactosylceramide (α-GalCer) adjuvant after storage of SmPill ® minispheres under room temperature and extreme storage conditions for several months. In addition, the internal structure of the cores of SmPill ® minispheres and antigen release features at intestinal pH were found to be preserved under all these conditions. However, changes in the surface morphology of SmPill ® minispheres leading to the antigen release at gastric pH were observed after a few weeks of storage under extreme conditions. Those modifications were prevented by the introduction of an Opadry ® White film coating layer between the core of SmPill ® minispheres and the enteric coating. Under these conditions, protection against antigen release at gastric pH was maintained even under high temperature and humidity conditions. These results support the potential of the SmPill ® minisphere approach to maintain the stability of an adjuvanted whole cell killed oral vaccine formulation. Copyright © 2017 Elsevier B.V. All rights reserved.

Does Salicylic Acid (SA) Improve Tolerance to Salt Stress in Plants? A Study of SA Effects On Tomato Plant Growth, Water Dynamics, Photosynthesis, and Biochemical Parameters.

PubMed

Mimouni, Hajer; Wasti, Salma; Manaa, Arafet; Gharbi, Emna; Chalh, Abdellah; Vandoorne, Bertrand; Lutts, Stanley; Ben Ahmed, Hela

2016-03-01

Environmental stresses such as salinity directly impact crop growth, and by extension, world food supply and societal prosperity. It is estimated that over 800 million hectares of land throughout the world are salt-affected. In arid and semi-arid regions, salt concentration can be close to that in the seawater. Hence, there are intensive efforts to improve plant tolerance to salinity and other environmental stressors. Salicylic acid (SA) is an important signal molecule for modulating plant responses to stress. In the present study, we examined, on multiple plant growth related endpoints, whether SA applied through the rooting medium could mitigate the adverse effects of salinity on tomato (Solanum lycopersicum) cv. Marmande. The latter is a hitherto understudied tomato plant from the above perspective; it is a classic variety that produces the large ribbed tomatoes in the Mediterranean and consumed worldwide. We found salt stress negatively affected the growth of cv. Marmande tomato plants. However, the SA-treated plants had greater shoot and root dry mass, leaf area compared to untreated plants when exposed to salt stress. Application of SA restores photosynthetic rates and photosynthetic pigment levels under salt (NaCl) exposure. Leaf water, osmotic potential, stomatal conductance transpiration rate, and biochemical parameters were also ameliorated in SA-treated plants under saline stress conditions. Overall, these data illustrate that SA increases cv. Marmande tomato growth by improving photosynthesis, regulation and balance of osmotic potential, induction of compatible osmolyte metabolism, and alleviating membrane damage. We suggest salicylic acid might be considered as a potential growth regulator to improve tomato plant salinity stress resistance, in the current era of global climate change.

Construction, expression, purification and biotin labeling of a single recombinant multi-epitope antigen for double-antigen sandwich ELISA to detect hepatitis C virus antibody.

PubMed

He, Jing; Xiu, Bingshui; Wang, Guohua; Chen, Kun; Feng, Xiaoyan; Song, Xiaoguo; Zhu, Cuixia; Yang, Xiqin; Bai, Guanzhong; Ling, Shigan; Zhang, Heqiu

2011-08-01

Based on B cell epitope predictions, a recombinant antigen with multiple epitopes from four Hepatitis C Virus fragments (C, NS3, NS4 and NS5) were engineered. The recombinant gene was then highly expressed in E. coli. The non-modified and C-terminal-modified recombinant proteins were used for coating and biotin labeling, respectively, to establish the double-antigen sandwich ELISA. Ten positive reference samples confirmed by the CHIRON RIBA HCV 3.0 SIA kit were detected positive, Forty one plasma samples were positive among samples from 441 volunteers, which indicated that the recombinant antigen could readily react well with plasma HCV antibody. As critical reagents of double-antigen sandwich ELISA, the recombinant multi-epitope antigen and the C-terminal-modified and biotin-conjugated antigen show good antigenicity. In this study, we provide a simple approach to produce multiple epitopes within one recombinant protein in order to avoid the costly expression of less-effective pools of multiple proteins, which is the conventional strategy of diagnostic antigen production for HCV antibody detection.

Vaginal type-II mucosa is an inductive site for primary CD8+ T-cell mucosal immunity

PubMed Central

Wang, Yichuan; Sui, Yongjun; Kato, Shingo; Hogg, Alison E.; Steel, Jason C.; Morris, John C.; Berzofsky, Jay A.

2014-01-01

The structured lymphoid tissues are considered the only inductive sites where primary T cell immune responses occur. The naïve T cells in structured lymphoid tissues, once being primed by antigen -bearing dendritic cells, differentiate into memory T cells and traffic back to the mucosal sites through the bloodstream. Contrary to this belief, here we show that the vaginal type-II mucosa itself, despite lack of structured lymphoid tissues, can act as an inductive site during primary CD8+ T cell immune responses. We provide evidence that the vaginal mucosa supports both the local immune priming of naïve CD8+ T cells and the local expansion of antigen-specific CD8+ T cells, thereby demonstrating a different paradigm for primary mucosal T cell immune induction. PMID:25600442

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

PubMed Central

Gao, Jie; Ochyl, Lukasz J; Yang, Ellen; Moon, James J

2017-01-01

Cationic liposomes (CLs) have been widely examined as vaccine delivery nanoparticles since they can form complexes with biomacromolecules, promote delivery of antigens and adjuvant molecules to antigen-presenting cells (APCs), and mediate cellular uptake of vaccine components. CLs are also known to trigger antigen cross-presentation – the process by which APCs internalize extracellular protein antigens, degrade them into minimal CD8+ T-cell epitopes, and present them in the context of major histocompatibility complex-I (MHC-I). However, the precise mechanisms behind CL-mediated induction of cross-presentation and cross-priming of CD8+ T-cells remain to be elucidated. In this study, we have developed two distinct CL systems and examined their impact on the lysosomal pH in dendritic cells (DCs), antigen degradation, and presentation of peptide:MHC-I complexes to antigen-specific CD8+ T-cells. To achieve this, we have used 3β-[N-(N′,N′-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) as the prototypical components of CLs with tertiary amine groups and compared the effect of CLs and anionic liposomes on lysosomal pH, antigen degradation, and cross-presentation by DCs. Our results showed that CLs, but not anionic liposomes, elevated the lysosomal pH in DCs and reduced antigen degradation, thereby promoting cross-presentation and cross-priming of CD8+ T-cell responses. These studies shed new light on CL-mediated cross-presentation and suggest that intracellular fate of vaccine components and subsequent immunological responses can be controlled by rational design of nanomaterials. PMID:28243087

Sarcocystis neurona Merozoites Express a Family of Immunogenic Surface Antigens That Are Orthologues of the Toxoplasma gondii Surface Antigens (SAGs) and SAG-Related Sequences†

PubMed Central

Howe, Daniel K.; Gaji, Rajshekhar Y.; Mroz-Barrett, Meaghan; Gubbels, Marc-Jan; Striepen, Boris; Stamper, Shelby

2005-01-01

Sarcocystis neurona is a member of the Apicomplexa that causes myelitis and encephalitis in horses but normally cycles between the opossum and small mammals. Analysis of an S. neurona expressed sequence tag (EST) database revealed four paralogous proteins that exhibit clear homology to the family of surface antigens (SAGs) and SAG-related sequences of Toxoplasma gondii. The primary peptide sequences of the S. neurona proteins are consistent with the two-domain structure that has been described for the T. gondii SAGs, and each was predicted to have an amino-terminal signal peptide and a carboxyl-terminal glycolipid anchor addition site, suggesting surface localization. All four proteins were confirmed to be membrane associated and displayed on the surface of S. neurona merozoites. Due to their surface localization and homology to T. gondii surface antigens, these S. neurona proteins were designated SnSAG1, SnSAG2, SnSAG3, and SnSAG4. Consistent with their homology, the SnSAGs elicited a robust immune response in infected and immunized animals, and their conserved structure further suggests that the SnSAGs similarly serve as adhesins for attachment to host cells. Whether the S. neurona SAG family is as extensive as the T. gondii SAG family remains unresolved, but it is probable that additional SnSAGs will be revealed as more S. neurona ESTs are generated. The existence of an SnSAG family in S. neurona indicates that expression of multiple related surface antigens is not unique to the ubiquitous organism T. gondii. Instead, the SAG gene family is a common trait that presumably has an essential, conserved function(s). PMID:15664946

Sarcocystis neurona merozoites express a family of immunogenic surface antigens that are orthologues of the Toxoplasma gondii surface antigens (SAGs) and SAG-related sequences.

PubMed

Howe, Daniel K; Gaji, Rajshekhar Y; Mroz-Barrett, Meaghan; Gubbels, Marc-Jan; Striepen, Boris; Stamper, Shelby

2005-02-01

Sarcocystis neurona is a member of the Apicomplexa that causes myelitis and encephalitis in horses but normally cycles between the opossum and small mammals. Analysis of an S. neurona expressed sequence tag (EST) database revealed four paralogous proteins that exhibit clear homology to the family of surface antigens (SAGs) and SAG-related sequences of Toxoplasma gondii. The primary peptide sequences of the S. neurona proteins are consistent with the two-domain structure that has been described for the T. gondii SAGs, and each was predicted to have an amino-terminal signal peptide and a carboxyl-terminal glycolipid anchor addition site, suggesting surface localization. All four proteins were confirmed to be membrane associated and displayed on the surface of S. neurona merozoites. Due to their surface localization and homology to T. gondii surface antigens, these S. neurona proteins were designated SnSAG1, SnSAG2, SnSAG3, and SnSAG4. Consistent with their homology, the SnSAGs elicited a robust immune response in infected and immunized animals, and their conserved structure further suggests that the SnSAGs similarly serve as adhesins for attachment to host cells. Whether the S. neurona SAG family is as extensive as the T. gondii SAG family remains unresolved, but it is probable that additional SnSAGs will be revealed as more S. neurona ESTs are generated. The existence of an SnSAG family in S. neurona indicates that expression of multiple related surface antigens is not unique to the ubiquitous organism T. gondii. Instead, the SAG gene family is a common trait that presumably has an essential, conserved function(s).

Comparative characteristics of the VP7 and VP4 antigenic epitopes of the rotaviruses circulating in Russia (Nizhny Novgorod) and the Rotarix and RotaTeq vaccines.

PubMed

Morozova, O V; Sashina, T A; Fomina, S G; Novikova, N A

2015-07-01

Two live, attenuated rotavirus A (RVA) vaccines, Rotarix and RotaTeq, have been successfully introduced into national immunization programs worldwide. The parent strains of both vaccines were obtained more than 30 years ago. Nonetheless, only very limited data are available on the molecular similarity of the vaccine strains and their genetic relationships to the wild-type strains circulating within the territory of Russian Federation. In this study, we have determined the nucleotide sequences of the genes encoding the viral proteins VP7 and VP4 (the globular domain VP8*) of vaccine strains and natural isolates of rotaviruses in Nizhny Novgorod, Russia. The VP7 and VP4 proteins contain antigenic sites that are the main targets of neutralizing antibodies. Phylogenetic analysis based on VP4 and VP7 showed that the majority of the natural RVA isolates from Nizhny Novgorod and the vaccine strains belong to different clusters. Four amino acids within the VP7 antigenic sites were common in both the wild-type and vaccine strains. The largest number of amino acid differences was found between the vaccine strain Rotarix and the Nizhny Novgorod G2 strains (19 residues out of 29). From 3 to 5 amino acid differences per strain were identified in the antigenic sites of VP4 (domain VP8*) between wild-type strains and the vaccine RotaTeq, and 6-8 substitutions were found when they were compared with the vaccine strain Rotarix. For the first time, immunodominant T-cell epitopes of VP7 were analyzed, and differences in the sequences between the vaccine and the wild-type strains were found. The accumulation of amino acid substitutions in the VP7 and VP4 antigenic sites may potentially reduce the immune protection of vaccinated children from wild-type strains of rotavirus.

Why do proteases mess up with antigen presentation by re-shuffling antigen sequences?

PubMed

Liepe, Juliane; Ovaa, Huib; Mishto, Michele

2018-04-30

The sequence of a large number of MHC-presented epitopes is not present as such in the original antigen because it has been re-shuffled by the proteasome or other proteases. Why do proteases throw a spanner in the works of our model of antigen tagging and immune recognition? We describe in this review what we know about the immunological relevance of post-translationally spliced epitopes and why proteases seem to have a second (dark) personality, which is keen to create new peptide bonds. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Original antigenic sin: A comprehensive review.

PubMed

Vatti, Anup; Monsalve, Diana M; Pacheco, Yovana; Chang, Christopher; Anaya, Juan-Manuel; Gershwin, M Eric

2017-09-01

The concept of "original antigenic sin" was first proposed by Thomas Francis, Jr. in 1960. This phenomenon has the potential to rewrite what we understand about how the immune system responds to infections and its mechanistic implications on how vaccines should be designed. Antigenic sin has been demonstrated to occur in several infectious diseases in both animals and humans, including human influenza infection and dengue fever. The basis of "original antigenic sin" requires immunological memory, and our immune system ability to autocorrect. In the context of viral infections, it is expected that if we are exposed to a native strain of a pathogen, we should be able to mount a secondary immune response on subsequent exposure to the same pathogen. "Original antigenic sin" will not contradict this well-established immunological process, as long as the subsequent infectious antigen is identical to the original one. But "original antigenic sin" implies that when the epitope varies slightly, then the immune system relies on memory of the earlier infection, rather than mount another primary or secondary response to the new epitope which would allow faster and stronger responses. The result is that the immunological response may be inadequate against the new strain, because the immune system does not adapt and instead relies on its memory to mount a response. In the case of vaccines, if we only immunize to a single strain or epitope, and if that strain/epitope changes over time, then the immune system is unable to mount an accurate secondary response. In addition, depending of the first viral exposure the secondary immune response can result in an antibody-dependent enhancement of the disease or at the opposite, it could induce anergy. Both of them triggering loss of pathogen control and inducing aberrant clinical consequences. Copyright © 2017 Elsevier Ltd. All rights reserved.

76 FR 71622 - Proposed Collection; Comment Request for Form 1099-SA

Federal Register 2010, 2011, 2012, 2013, 2014

2011-11-18

... comments concerning Form 1099-SA, Distributions From an HSA, Archer MSA or Medical Advantage MSA. DATES... HSA, Archer MSA or Medical Advantage MSA. OMB Number: 1545-1517. Form Number: 1099-SA. Abstract: This... Review: Extension of a currently approved collection. Affected Public: Business or other for-profit...

Indium 111 ZCE-025 immunoscintigraphy in occult recurrent colorectal cancer with elevated carcinoembryonic antigen level

DOE Office of Scientific and Technical Information (OSTI.GOV)

Doerr, R.J.; Abdel-Nabi, H.; Merchant, B.

1990-02-01

We investigated the utility of scanning with indium 111 labeled to monoclonal antibody in 13 patients after curative resection of colorectal cancer who had elevated carcinoembryonic antigen levels and negative results of clinical workup. Each patient received 1 mg of anti-carcinoembryonic antigen monoclonal antibody type ZCE 025 labeled with 5.5 mCi of {sup 111}In, plus 9 to 39 mg of the same antibody unlabeled. Patients underwent scanning 3 to 7 days after infusion by planar and emission computed tomography. ZCE-025 monoclonal antibody imaging detected tumor recurrence or metastasis in 11 of 13 patients. In one patient the monoclonal antibody scanmore » gave a true-negative result, and in one patient the monoclonal antibody scan failed to disclose a metachronous cecal primary. Tumor sites identified were the pelvis (2 patients), abdominal wall (2), retroperitoneum (1), lymph nodes (3); liver (2), bone (2), and lung (1). The accurate localization of colorectal carcinoma recurrences by means of {sup 111}In ZCE-025 monoclonal antibody demonstrates the usefulness of this diagnostic agent in the setting of elevated carcinoembryonic antigen level and negative results of clinical and radiologic workup.« less

A Modeling and Experimental Investigation of the Effects of Antigen Density, Binding Affinity, and Antigen Expression Ratio on Bispecific Antibody Binding to Cell Surface Targets*

PubMed Central

Rhoden, John J.; Dyas, Gregory L.

2016-01-01

Despite the increasing number of multivalent antibodies, bispecific antibodies, fusion proteins, and targeted nanoparticles that have been generated and studied, the mechanism of multivalent binding to cell surface targets is not well understood. Here, we describe a conceptual and mathematical model of multivalent antibody binding to cell surface antigens. Our model predicts that properties beyond 1:1 antibody:antigen affinity to target antigens have a strong influence on multivalent binding. Predicted crucial properties include the structure and flexibility of the antibody construct, the target antigen(s) and binding epitope(s), and the density of antigens on the cell surface. For bispecific antibodies, the ratio of the expression levels of the two target antigens is predicted to be critical to target binding, particularly for the lower expressed of the antigens. Using bispecific antibodies of different valencies to cell surface antigens including MET and EGF receptor, we have experimentally validated our modeling approach and its predictions and observed several nonintuitive effects of avidity related to antigen density, target ratio, and antibody affinity. In some biological circumstances, the effect we have predicted and measured varied from the monovalent binding interaction by several orders of magnitude. Moreover, our mathematical framework affords us a mechanistic interpretation of our observations and suggests strategies to achieve the desired antibody-antigen binding goals. These mechanistic insights have implications in antibody engineering and structure/activity relationship determination in a variety of biological contexts. PMID:27022022

A Modeling and Experimental Investigation of the Effects of Antigen Density, Binding Affinity, and Antigen Expression Ratio on Bispecific Antibody Binding to Cell Surface Targets.

PubMed

Rhoden, John J; Dyas, Gregory L; Wroblewski, Victor J

2016-05-20

Despite the increasing number of multivalent antibodies, bispecific antibodies, fusion proteins, and targeted nanoparticles that have been generated and studied, the mechanism of multivalent binding to cell surface targets is not well understood. Here, we describe a conceptual and mathematical model of multivalent antibody binding to cell surface antigens. Our model predicts that properties beyond 1:1 antibody:antigen affinity to target antigens have a strong influence on multivalent binding. Predicted crucial properties include the structure and flexibility of the antibody construct, the target antigen(s) and binding epitope(s), and the density of antigens on the cell surface. For bispecific antibodies, the ratio of the expression levels of the two target antigens is predicted to be critical to target binding, particularly for the lower expressed of the antigens. Using bispecific antibodies of different valencies to cell surface antigens including MET and EGF receptor, we have experimentally validated our modeling approach and its predictions and observed several nonintuitive effects of avidity related to antigen density, target ratio, and antibody affinity. In some biological circumstances, the effect we have predicted and measured varied from the monovalent binding interaction by several orders of magnitude. Moreover, our mathematical framework affords us a mechanistic interpretation of our observations and suggests strategies to achieve the desired antibody-antigen binding goals. These mechanistic insights have implications in antibody engineering and structure/activity relationship determination in a variety of biological contexts. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

SA-Search: a web tool for protein structure mining based on a Structural Alphabet

PubMed Central

Guyon, Frédéric; Camproux, Anne-Claude; Hochez, Joëlle; Tufféry, Pierre

2004-01-01

SA-Search is a web tool that can be used to mine for protein structures and extract structural similarities. It is based on a hidden Markov model derived Structural Alphabet (SA) that allows the compression of three-dimensional (3D) protein conformations into a one-dimensional (1D) representation using a limited number of prototype conformations. Using such a representation, classical methods developed for amino acid sequences can be employed. Currently, SA-Search permits the performance of fast 3D similarity searches such as the extraction of exact words using a suffix tree approach, and the search for fuzzy words viewed as a simple 1D sequence alignment problem. SA-Search is available at http://bioserv.rpbs.jussieu.fr/cgi-bin/SA-Search. PMID:15215446

SA-Search: a web tool for protein structure mining based on a Structural Alphabet.

PubMed

Guyon, Frédéric; Camproux, Anne-Claude; Hochez, Joëlle; Tufféry, Pierre

2004-07-01

SA-Search is a web tool that can be used to mine for protein structures and extract structural similarities. It is based on a hidden Markov model derived Structural Alphabet (SA) that allows the compression of three-dimensional (3D) protein conformations into a one-dimensional (1D) representation using a limited number of prototype conformations. Using such a representation, classical methods developed for amino acid sequences can be employed. Currently, SA-Search permits the performance of fast 3D similarity searches such as the extraction of exact words using a suffix tree approach, and the search for fuzzy words viewed as a simple 1D sequence alignment problem. SA-Search is available at http://bioserv.rpbs.jussieu.fr/cgi-bin/SA-Search.

Multiple slips effects on MHD SA-Al2O3 and SA-Cu non-Newtonian nanofluids flow over a stretching cylinder in porous medium with radiation and chemical reaction

NASA Astrophysics Data System (ADS)

Tlili, I.; Khan, W. A.; Khan, I.

2018-03-01

The purpose of this communication is to examine the collective influence of velocity, and thermal slips on magnetohydrodyanmics (MHD) SA-Al2O3 and SA-Cu non-Newtonian nanofluids flow over a stretching cylinder in porous medium together with thermal radiation and chemical reaction effects. Sodium Alginate (SA-NaAlg) is taken as non-Newtonian base fluid. Two types of nanoparticles alumina or aluminum oxide (Al2O3) and copper (Cu) are suspended in sodium alginate (SA) which is taken as base fluid, an example of non-Newtonian Casson fluid. The formulated nonlinear partial differential equations with auxiliary boundary conditions are transformed into non-dimensional form by applying suitable similarity transformations. The resulting dimensionless problem is solved numerically using shooting and fourth order Runge-Kutta method. The impacts of various thermophysical parameters on local skin-friction, local Nusselt number, temperature and velocity are analyzed through graphs as well as in tabular form and discussed in detail. A comparison between SA-Al2O3 and SA-Cu nanofluids is clearly shown and in limiting sense the present results are compared with published results from literature. The results show that with magnetic parameter, skin-friction and Nusselt number both decreased, and Nusselt numbers are the highest in case of Al2O3 than Cu nanoparticles.

[The isolation and evaluation of Aspergillus fumigatus antigens].

PubMed

Lirio, V de S; de Assis, C M; Cano, M I; Lacaz, C da S

1992-01-01

Antigens from three strains of Aspergillus fumigatus (354, 356, and JIG) and an antiserum against the mixing of these antigens have been produced, and evaluated immunochemically. The antigens were obtained through a modified Coleman & Kaufman technique (culture filtrate concentrated by acetone). Analysis by the immunodiffusion test (ID) against homologous serum has yielded 100% sensitivity (with the studied sera). Concerning heterologous sera we found reactivity with a serum of a patient of candidiasis and another with histoplasmosis. The same result was obtained with a reference antigen in immunodiffusion, showing similar standards of response. Titration of the antiserum by ID and counterimmunoelectrophoresis showed a title of 1:32, and by complement fixation (micro-technique) a title of 1:128. Using immunoelectrophoresis (IEF), the produced antiserum yielded 8 lines of precipitation (5 in the anodic pole and 3 in the cathodic one). In SDS-PAGE at 12.5% the antigen has presented a rather complex electrophoretic profile (26 proteic subunits with a molecular weight ranging from 18 a > 100 kDa). Immunogenicity of the antigen was observed in all fractions of SDS-PAGE when the immunoblotting against the antiserum was carried out.

Personalized Therapy: Tumor Antigen Discovery for Adoptive Cellular Therapy.

PubMed

Yee, Cassian; Lizee, Gregory A

Adoptive cell therapy using endogenous T cells involves the ex vivo isolation and expansion of antigen-specific T cells from the peripheral blood and is uniquely suited for validating and translating antigen discovery. Endogenous T-cell therapy does not require accessible tumor as a source of infiltrating T cells and is free of regulatory and logistical constraints associated with engineering T cells. Candidate epitope peptides identified through antigen discovery may be rapidly implemented as targets in clinical trials of endogenous T-cell therapy and even incorporated as an "ad hoc" approach to personalized treatment when autologous tumor is available. Several first-in-human studies using a uniform population of antigen-specific T cells defined by phenotype and specificity have provided a means to confirm candidate antigens as potential tumor rejection antigens and to evaluate the reasons for success or failure using as a "transferrable cellular biomarker" the adoptively transferred T cells.

Immunogenicity of a Japanese encephalitis chimeric virus vaccine as a booster dose after primary vaccination with SA14-14-2 vaccine in Thai children.

PubMed

Janewongwirot, Pakpoom; Puthanakit, Thanyawee; Anugulruengkitt, Suvaporn; Jantarabenjakul, Watsamon; Phasomsap, Chayapa; Chumket, Sompong; Yoksan, Sutee; Pancharoen, Chitsanu

2016-10-17

Japanese Encephalitis chimeric virus vaccine (JE-CV) and SA14-14-2 vaccine are live-attenuated JE vaccines produced from the same virus strain. Data on interchangeability is limited. To evaluate the immunogenicity and safety of JE-CV booster after primary vaccination with SA14-14-2 vaccine. This study was an open-label clinical trial in Thai children who had received a primary SA14-14-2 vaccination at 12-24monthsbefore enrollment (ClinicalTrials.gov NCT02602652). JE-CV was administered. A 50% plaque reduction neutralization test (PRNT 50 ) against three virus strains; JE-CV, SA-14-14-2andwild-type JE virus was measured before and 28-days post vaccination. The laboratory was performed at PRNT 50 titers ⩾10 (1/dil) were considered seroprotective against JE. Geometric mean titer (GMT) of PRNT 50 was calculated. Adverse events were observed for 28days. From March 2014 to June 2015, 50 children (64% male) were enrolled. Mean age and duration after primary vaccination was 26.9 (SD 4.6) and 12.8 (SD 2.7) months, respectively. The proportion of participants who had PRNT 50 pre and post-booster vaccination were 92% and 96% against JE-CV virus, 56% and 98% against SA-14-14-2 strain and 70% and 98% against wild-type JE virus, respectively. Solicited injection site reactions including erythema, pain and swelling occurred in 18%, 10% and 4% of subjects, respectively. Four children (8%) had fever (⩾37.7Celsius). Eight children (16%) had adverse events, which were not related to the vaccine. AJE-CV booster dose is highly immunogenic and safe among children who previously received SA14-14-2 vaccine. Copyright © 2016 Elsevier Ltd. All rights reserved.

Multifunctional SA-PProDOT Binder for Lithium Ion Batteries.

PubMed

Ling, Min; Qiu, Jingxia; Li, Sheng; Yan, Cheng; Kiefel, Milton J; Liu, Gao; Zhang, Shanqing

2015-07-08

An environmentally benign, highly conductive, and mechanically strong binder system can overcome the dilemma of low conductivity and insufficient mechanical stability of the electrodes to achieve high performance lithium ion batteries (LIBs) at a low cost and in a sustainable way. In this work, the naturally occurring binder sodium alginate (SA) is functionalized with 3,4-propylenedioxythiophene-2,5-dicarboxylic acid (ProDOT) via a one-step esterification reaction in a cyclohexane/dodecyl benzenesulfonic acid (DBSA)/water microemulsion system, resulting in a multifunctional polymer binder, that is, SA-PProDOT. With the synergetic effects of the functional groups (e.g., carboxyl, hydroxyl, and ester groups), the resultant SA-PProDOT polymer not only maintains the outstanding binding capabilities of sodium alginate but also enhances the mechanical integrity and lithium ion diffusion coefficient in the LiFePO4 (LFP) electrode during the operation of the batteries. Because of the conjugated network of the PProDOT and the lithium doping under the battery environment, the SA-PProDOT becomes conductive and matches the conductivity needed for LiFePO4 LIBs. Without the need of conductive additives such as carbon black, the resultant batteries have achieved the theoretical specific capacity of LiFePO4 cathode (ca. 170 mAh/g) at C/10 and ca. 120 mAh/g at 1C for more than 400 cycles.

Interleukin 18 secretion and its effect in improving Chimeric Antigen Receptors efficiency

NASA Astrophysics Data System (ADS)

Kim, Jae-Kun

Clinical trials have shown that chimeric antigen receptor T cells modified to target cancer cells expressing a surface antigen found on immature B-cells. The purpose of this experiment is to take a pro-inflammatory cytokine, and analyze its effect in improving the efficiency of the T cells. IL-18 has been previously shown to recruit T cells to the tumor site and improve their secretion of cytotoxic cytokines. A human model of the proposed armored T cell has been created and has shown success in combating cancer cells in vitro. The next step is to design and produce a murine model to test in vivo in immunocompetent mice. This research project aimed to create two models: one utilizing 2A peptides and another utilizing IRES elements as a multicistronic vector. Both models would require the insertion of the desired genes into SFG backbones. IRES, a DNA element which acts as a binding site for the transcriptional machinery to recognize which part of the DNA to transcribe, commonly found in bicistronic vectors, is large with 500-600 base pairs, and has a lower transgene expression rate. P2A is smaller, only consisting of about 20 amino acids, and typically has a higher transgene expression rate, which may or may not result in higher effectiveness of the model. I would like to thank Dr. Renier Brentjens for being a mentor who cared about giving his interns as much educational value as possible.

[Biochemical characteristics and antigenic structures of Chlamydia].

PubMed

Puy, H; Fuentes, V; Eb, F; Orfila, J

1989-01-01

New biotechnology in immunology and molecular biology has enabled the identification and definition of the structure of glycolipids and especially membrane proteins of Chlamydia. Chlamydia antigen lipopolysaccharide, major outer membrane protein, protein 74 kDa, eukaryotic cell binding protein and cysteine rich proteins are all carriers of antigenic determinants, genus, species or type specific. They are very usefull for diagnosis of Chlamydial infections and epidemiological studies. These membranous antigens have an important role in the pathogenesis of these bacteries. Finally these studies have contributed to the isolation of a new species: C. pneumoniae (TWAR strains).

21 CFR 660.1 - Antibody to Hepatitis B Surface Antigen.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 7 2013-04-01 2013-04-01 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...

21 CFR 660.1 - Antibody to Hepatitis B Surface Antigen.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 7 2014-04-01 2014-04-01 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...

21 CFR 660.1 - Antibody to Hepatitis B Surface Antigen.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 7 2012-04-01 2012-04-01 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...

21 CFR 660.1 - Antibody to Hepatitis B Surface Antigen.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 7 2011-04-01 2010-04-01 true 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...

21 CFR 660.1 - Antibody to Hepatitis B Surface Antigen.

Code of Federal Regulations, 2010 CFR

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...

Lipid antigens in immunity

PubMed Central

Dowds, C. Marie; Kornell, Sabin-Christin

2014-01-01

Lipids are not only a central part of human metabolism but also play diverse and critical roles in the immune system. As such, they can act as ligands of lipid-activated nuclear receptors, control inflammatory signaling through bioactive lipids such as prostaglandins, leukotrienes, lipoxins, resolvins, and protectins, and modulate immunity as intracellular phospholipid- or sphingolipid-derived signaling mediators. In addition, lipids can serve as antigens and regulate immunity through the activation of lipid-reactive T cells, which is the topic of this review. We will provide an overview of the mechanisms of lipid antigen presentation, the biology of lipid-reactive T cells, and their contribution to immunity. PMID:23999493

Engineering antigens for in situ erythrocyte binding induces T-cell deletion.

PubMed

Kontos, Stephan; Kourtis, Iraklis C; Dane, Karen Y; Hubbell, Jeffrey A

2013-01-02

Antigens derived from apoptotic cell debris can drive clonal T-cell deletion or anergy, and antigens chemically coupled ex vivo to apoptotic cell surfaces have been shown correspondingly to induce tolerance on infusion. Reasoning that a large number of erythrocytes become apoptotic (eryptotic) and are cleared each day, we engineered two different antigen constructs to target the antigen to erythrocyte cell surfaces after i.v. injection, one using a conjugate with an erythrocyte-binding peptide and another using a fusion with an antibody fragment, both targeting the erythrocyte-specific cell surface marker glycophorin A. Here, we show that erythrocyte-binding antigen is collected much more efficiently than free antigen by splenic and hepatic immune cell populations and hepatocytes, and that it induces antigen-specific deletional responses in CD4(+) and CD8(+) T cells. We further validated T-cell deletion driven by erythrocyte-binding antigens using a transgenic islet β cell-reactive CD4(+) T-cell adoptive transfer model of autoimmune type 1 diabetes: Treatment with the peptide antigen fused to an erythrocyte-binding antibody fragment completely prevented diabetes onset induced by the activated, autoreactive CD4(+) T cells. Thus, we report a translatable modular biomolecular approach with which to engineer antigens for targeted binding to erythrocyte cell surfaces to induce antigen-specific CD4(+) and CD8(+) T-cell deletion toward exogenous antigens and autoantigens.

A sarcoidosis clinician's perspective of MHC functional elements outside the antigen binding site.

PubMed

Judson, Marc A

2018-05-30

Sarcoidosis is a multisystem granulomatous disease of unknown cause. Evidence supports an integral role for interactions at the MHC binding site in the development of sarcoidosis. However, despite this evidence, there are clinical data that suggest that additional mechanisms are involved in the immunopathogenesis of this disease. This manuscript provides a brief clinical description of sarcoidosis, and a clinician's perspective of the immunopathogenesis of sarcoidosis in terms of the MHC binding site, MHC functional elements beyond the binding site, and other possible alternative mechanisms. Input from clinicians will be essential in establishing the immunologic cause of sarcoidosis as a detailed phenotypic characterization of disease will be required. Copyright © 2018. Published by Elsevier Inc.

A New MIC1-MAG1 Recombinant Chimeric Antigen Can Be Used Instead of the Toxoplasma gondii Lysate Antigen in Serodiagnosis of Human Toxoplasmosis

PubMed Central

Holec-Gąsior, Lucyna; Ferra, Bartłomiej; Drapała, Dorota; Lautenbach, Dariusz

2012-01-01

This study presents an evaluation of the MIC1 (microneme protein 1)-MAG1 (matrix antigen 1) Toxoplasma gondii recombinant chimeric antigen for the serodiagnosis of human toxoplasmosis for the first time. The recombinant MIC1-MAG1 antigen was obtained as a fusion protein containing His tags at the N- and C-terminal ends using an Escherichia coli expression system. After purification by metal affinity chromatography, the chimeric protein was tested for usefulness in an enzyme-linked immunosorbent assay (ELISA) for the detection of anti-T. gondii immunoglobulin G (IgG). One hundred ten sera from patients at different stages of infection and 40 sera from seronegative patients were examined. The results obtained for the MIC1-MAG1 chimeric antigen were compared with those of IgG ELISAs using a Toxoplasma lysate antigen (TLA), a combination of recombinant antigens (rMIC1ex2-rMAG1) and single recombinant proteins (rMIC1ex2 and rMAG1). The sensitivity of the IgG ELISA calculated from all of the positive serum samples was similar for the MIC1-MAG1 chimeric antigen (90.8%) and the TLA (91.8%), whereas the sensitivities of the other antigenic samples used were definitely lower, at 69.1% for the mixture of antigens, 75.5% for the rMIC1ex2, and 60% for rMAG1. This study demonstrates that the MIC1-MAG1 recombinant chimeric antigen can be used instead of the TLA in the serodiagnosis of human toxoplasmosis. PMID:22116686

Evaluation of cytokine responses against novel Mtb antigens as diagnostic markers for TB disease.

PubMed

Awoniyi, Dolapo O; Teuchert, Andrea; Sutherland, Jayne S; Mayanja-Kizza, Harriet; Howe, Rawleigh; Mihret, Adane; Loxton, Andre G; Sheehama, Jacob; Kassa, Desta; Crampin, Amelia C; Dockrell, Hazel M; Kidd, Martin; Rosenkrands, Ida; Geluk, Annemieke; Ottenhoff, Tom H M; Corstjens, P L A M; Chegou, Novel N; Walzl, Gerhard

2016-09-01

We investigated the accuracy of host markers detected in Mtb antigen-stimulated whole blood culture supernatant in the diagnosis of TB. Prospectively, blood from 322 individuals with presumed TB disease from six African sites was stimulated with four different Mtb antigens (Rv0081, Rv1284, ESAT-6/CFP-10, and Rv2034) in a 24 h whole blood stimulation assay (WBA). The concentrations of 42 host markers in the supernatants were measured using the Luminex multiplex platform. Diagnostic biosignatures were investigated through the use of multivariate analysis techniques. 17% of the participants were HIV infected, 106 had active TB disease and in 216 TB was excluded. Unstimulated concentrations of CRP, SAA, ferritin and IP-10 had better discriminating ability than markers from stimulated samples. Accuracy of marker combinations by general discriminant analysis (GDA) identified a six analyte model with 77% accuracy for TB cases and 84% for non TB cases, with a better performance in HIV uninfected patients. A biosignature of 6 cytokines obtained after stimulation with four Mtb antigens has moderate potential as a diagnostic tool for pulmonary TB disease individuals and stimulated marker expression had no added value to unstimulated marker performance. Copyright © 2016 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Correlates of protection, antigen delivery and molecular epidemiology: basics for designing an HIV vaccine.

PubMed

Wagner, R; Shao, Y; Wolf, H

1999-03-26

Major obstacles in the development of HIV vaccines are the high variability of the virus and its complex interaction with the immune system. Recent studies demonstrated, that CTLs recognizing highly conserved epitopes in the group-specific antigen are capable of controlling HIV-replication in long-term nonprogressors. Necessary consequences for novel vaccine concepts are the presentation of a large repertoire of antigenic sites as well as the stimulation of different effectors of the immune system. Accordingly, different types of recombinant HIV-1 virus-like particles (VLPs) have been constructed stimulating the induction of neutralizing antibodies and HIV-specific CD8-positive CTL responses in preclinical studies. With respect to future vaccine trials, HIV vaccine formulations may need to be tailored to the local strains circulating within a geographical region. The expert group of the joint United Nations Programme on AIDS recently identified Yunnan, a southwestern province of China, as a region, in which the HIV epidemic is starting to gain speed, resembling to the situation in Thailand 10 years ago. A molecular clone of a representative virus strain is now available for the development of innovative antigen delivery systems aiming to be evaluated in future clinical vaccine trials throughout this area.

SA1 and TRF1 synergistically bind to telomeric DNA and promote DNA-DNA pairing

NASA Astrophysics Data System (ADS)

Wang, Hong; Lin, Jiangguo; Countryman, Preston; Pan, Hai; Parminder Kaur Team; Robert Riehn Team; Patricia Opresko Team; Jane Tao Team; Susan Smith Team

Impaired telomere cohesion leads to increased aneuploidy and early onset of tumorigenesis. Cohesion is thought to occur through the entrapment of two DNA strands within tripartite cohesin ring(s), along with a fourth subunit (SA1/SA2). Surprisingly, cohesion rings are not essential for telomere cohesion, which instead requires SA1 and shelterin proteins including TRF1. However, neither this unique cohesion mechanism at telomeres or DNA-binding properties of SA1 is understood. Here, using single-molecule fluorescence imaging of quantum dot-labeled proteins on DNA we discover that while SA1 diffuses across multiple telomeric and non-telomeric regions, the diffusion mediated through its N-terminal domain is slower at telomeric regions. However, addition of TRF1 traps SA1 within telomeric regions, which form longer DNA-DNA pairing tracts than with TRF1 alone, as revealed by atomic force microscopy. Together, these experimental results and coarse-grained molecular dynamics simulations suggest that TRF1 and SA1 synergistically interact with DNA to support telomere cohesion without cohesin rings.

[Detection of antigen structures in blood cells in various prepared plasma transfusions].

PubMed

Barz, D

1994-01-01

We investigated the content of antigen-bearing cells and cell fragments in Fresh Frozen Plasma (FFP) from blood centers, in Octaplas (virus-inactivated fresh plasma produced with the solvent/detergent technique by the Octapharma Company) and in MB-plasma (virus-inactivated fresh plasma after photodynamic treatment with methylen blue coming from the German Red Cross in Springe, Lower Saxony). With the aid of an immunoassay (MAIPA-test) these plasmas were tested regarding Rhesus-D-antigen, HLA-class-I- and HLA-class-II-antigens, platelet specific antigens HPA-1a/HPA-1b and granulocyte specific antigens NA1/NA2. In Octaplas (n = 10) we did not find cells or cell fragments and no antigen-bearing blood cell structures. In FFP (n = 28) there were platelet specific antigens in 27 cases (96.4%) and HLA-class-I-antigens in 4 cases (14.3%). In MB-plasma (n = 14) we found platelet specific antigens in all cases, HLA-class-I-antigens in 4 cases (18.6%), HLA-class-II-antigens and granulocyte specific antigens in 1 case (7.1%) and Rhesus-D-antigen in 3 cases (21.4%). Plasma derived from whole blood showed lower levels of cells and antigens than plasma which was produced with the aid of the cell separator.

Prediction of protein-protein interaction sites using electrostatic desolvation profiles.

PubMed

Fiorucci, Sébastien; Zacharias, Martin

2010-05-19

Protein-protein complex formation involves removal of water from the interface region. Surface regions with a small free energy penalty for water removal or desolvation may correspond to preferred interaction sites. A method to calculate the electrostatic free energy of placing a neutral low-dielectric probe at various protein surface positions has been designed and applied to characterize putative interaction sites. Based on solutions of the finite-difference Poisson equation, this method also includes long-range electrostatic contributions and the protein solvent boundary shape in contrast to accessible-surface-area-based solvation energies. Calculations on a large set of proteins indicate that in many cases (>90%), the known binding site overlaps with one of the six regions of lowest electrostatic desolvation penalty (overlap with the lowest desolvation region for 48% of proteins). Since the onset of electrostatic desolvation occurs even before direct protein-protein contact formation, it may help guide proteins toward the binding region in the final stage of complex formation. It is interesting that the probe desolvation properties associated with residue types were found to depend to some degree on whether the residue was outside of or part of a binding site. The probe desolvation penalty was on average smaller if the residue was part of a binding site compared to other surface locations. Applications to several antigen-antibody complexes demonstrated that the approach might be useful not only to predict protein interaction sites in general but to map potential antigenic epitopes on protein surfaces. Copyright (c) 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Prediction of Protein-Protein Interaction Sites Using Electrostatic Desolvation Profiles

PubMed Central

Fiorucci, Sébastien; Zacharias, Martin

2010-01-01

Abstract Protein-protein complex formation involves removal of water from the interface region. Surface regions with a small free energy penalty for water removal or desolvation may correspond to preferred interaction sites. A method to calculate the electrostatic free energy of placing a neutral low-dielectric probe at various protein surface positions has been designed and applied to characterize putative interaction sites. Based on solutions of the finite-difference Poisson equation, this method also includes long-range electrostatic contributions and the protein solvent boundary shape in contrast to accessible-surface-area-based solvation energies. Calculations on a large set of proteins indicate that in many cases (>90%), the known binding site overlaps with one of the six regions of lowest electrostatic desolvation penalty (overlap with the lowest desolvation region for 48% of proteins). Since the onset of electrostatic desolvation occurs even before direct protein-protein contact formation, it may help guide proteins toward the binding region in the final stage of complex formation. It is interesting that the probe desolvation properties associated with residue types were found to depend to some degree on whether the residue was outside of or part of a binding site. The probe desolvation penalty was on average smaller if the residue was part of a binding site compared to other surface locations. Applications to several antigen-antibody complexes demonstrated that the approach might be useful not only to predict protein interaction sites in general but to map potential antigenic epitopes on protein surfaces. PMID:20441756

Antigen-specific T cell responses to BK polyomavirus antigens identify functional anti-viral immunity and may help to guide immunosuppression following renal transplantation

PubMed Central

Chakera, A; Bennett, S; Lawrence, S; Morteau, O; Mason, P D; O'Callaghan, C A; Cornall, R J

2011-01-01

Infection with the polyoma virus BK (BKV) is a major cause of morbidity following renal transplantation. Limited understanding of the anti-viral immune response has prevented the design of a strategy that balances treatment with the preservation of graft function. The proven utility of interferon-gamma enzyme-linked immunospot (ELISPOT) assays to measure T cell responses in immunocompetent hosts was the basis for trying to develop a rational approach to the management of BKV following renal transplantation. In a sample of transplant recipients and healthy controls, comparisons were made between T cell responses to the complete panel of BKV antigens, the Epstein–Barr virus (EBV) antigens, BZLF1 and EBNA1, and the mitogen phytohaemagglutinin (PHA). Correlations between responses to individual antigens and immunosuppressive regimens were also analysed. Antigen-specific T cell responses were a specific indicator of recent or ongoing recovery from BKV infection (P < 0·05), with responses to different BKV antigens being highly heterogeneous. Significant BKV immunity was undetectable in transplant patients with persistent viral replication or no history of BKV reactivation. Responses to EBV antigens and mitogen were reduced in patients with BKV reactivation, but these differences were not statistically significant. The T cell response to BKV antigens is a useful and specific guide to recovery from BKV reactivation in renal transplant recipients, provided that the full range of antigenic responses is measured. PMID:21671906

Probing the Energetics of Antigen-Antibody Recognition by Titration Microcalorimetry

PubMed

Jelesarov; Leder; Bosshard

1996-06-01

Our understanding of the energetics that govern antigen-antibody recognition lags behind the increasingly rapid accumulation of structural information on antigen-antibody complexes. Thanks to the development of highly sensitive microcalorimeters, the thermodynamic parameters of antigen-antibody interactions can now be measured with precision and using only nanomole quantities of protein. The method of choice is isothermal titration calorimetry, in which a solution of the antibody (or antigen) is titrated with small aliquots of the antigen (or antibody) and the heat change accompanying the formation of the antigen-antibody complex is measured with a sensitivity as high as 0.1 μcal s-1. The free energy of binding (DeltaG), the binding enthalpy (DeltaH), and the binding entropy (DeltaS) are usually obtained from a single experiment, and no spectroscopic or radioactive label must be introduced into the antigen or antibody. The often large and negative change in heat capacity (DeltaCp) accompanying the formation of an antigen-antibody complex is obtained from DeltaH measured at different temperatures. The basic theory and the principle of the measurements are reviewed and illustrated by examples. The thermodynamic parameters relate to the dynamic physical forces that govern the association of the freely moving antigen and antibody into a well-structured and unique complex. This information complements the static picture of the antigen-antibody complex that results from X-ray diffraction analysis. Attempts to correlate dynamic and static aspects are discussed briefly.

Controlled Release of Antigens for One Dose Immunization

DTIC Science & Technology

1983-01-01

microencapsulation of antigen coated alum or by microencapsulating clusters of smaller (᝺ microns) microcapsules . Microcapsules under 10 microns in... microencapsulation were studied to determine what criteria must be satisfied to provide a protective immune response to hepatitis B surface antigen... microencapsulated in poly (DL-lactide-co- glycolide) in a form that was too large to be phagocytized and had an antigen release profile similar to that achieved with

Solving Fuzzy Optimization Problem Using Hybrid Ls-Sa Method

NASA Astrophysics Data System (ADS)

Vasant, Pandian

2011-06-01

Fuzzy optimization problem has been one of the most and prominent topics inside the broad area of computational intelligent. It's especially relevant in the filed of fuzzy non-linear programming. It's application as well as practical realization can been seen in all the real world problems. In this paper a large scale non-linear fuzzy programming problem has been solved by hybrid optimization techniques of Line Search (LS), Simulated Annealing (SA) and Pattern Search (PS). As industrial production planning problem with cubic objective function, 8 decision variables and 29 constraints has been solved successfully using LS-SA-PS hybrid optimization techniques. The computational results for the objective function respect to vagueness factor and level of satisfaction has been provided in the form of 2D and 3D plots. The outcome is very promising and strongly suggests that the hybrid LS-SA-PS algorithm is very efficient and productive in solving the large scale non-linear fuzzy programming problem.

Expression of Hepatitis C Virus Core and E2 antigenic recombinant proteins and their use for development of diagnostic assays.

PubMed

Ali, Amjad; Nisar, Muhammad; Idrees, Muhammad; Rafique, Shazia; Iqbal, Muhammad

2015-05-01

Early diagnosis of HCV infection is based on detection of antibodies against HCV proteins using recombinant viral antigens. The present study was designed to select, clone and express the antigenic regions of Core and E2 genes from local HCV-3a genotype and to utilize the antigenic recombinant proteins (Core & E2) to develop highly sensitive, specific and economical diagnostic assays for detection of HCV infection. The antigenic sites were determined within Core and E2 genes and were then cloned in pET-28a expression vector. The right orientation of the desired inserted fragments of Core and E2 were confirmed via sequencing prior to expression and were then transformed in BL21 (DE3) pLysS strains of E. coli and induced with 0.5mM Isopropyl-b-D-thiogalactopyranoside (IPTG) for the production of antigenic recombinant proteins. The produced truncated antigens were then purified by Nickel affinity chromatography and were confirmed by western blotting, immunoblotting and enzyme-linked immunosorbent assay (ELISA). The expressed Core and E2 recombinant antigens were used to develop immunoblotting assay for the detection of anti-HCV antibodies in sera. With immunoblotting, a total of 93-HCV infected sera and 35-HCV negative individuals were tested for the presence of anti-HCV antibodies to the Core and E2 antigens. Recombinant antigen showed 100% reactivity against HCV infected sera, with no cross reactivity against HCV-negative sera. The immunoblot assay mixture of recombinant antigens (Core+E2) showed a strong reaction intensity in the test area (TA) as compared to the individual truncated Core and E2 recombinant antigens. In the in-house ELISA assay, mixed Core and E2 recombinant antigens showed 100% reactivity against a standardized panel of 150-HCV-positive sera and non reactivity against a standardized panel of 150 HCV-negative sera while also being non reactive to sera positive for other viral infections. The antigenic recombinant antigens also were tested for the

Induction of human antigen-specific suppressor factors in vitro.

PubMed Central

Kontiainen, S; Woody, J N; Rees, A; Feldmann, M

1981-01-01

Based on methods used for the in vitro induction of antigen-specific suppressor cells in the mouse, we have cultured Ficoll-Isopaque-separated human blood cells with high dose of antigen (100 microgram/ml) in Marbrook culture vessels for 4 days. The resulting cells, when further recultured for 24 hr with a low dose of antigen (1 microgram/ml), released into the supernatant material, termed 'suppressor factor', which inhibited, in an antigen-specific manner, the antibody response of mouse spleen cells in culture. The suppressor factor was analysed using immunoabsorbents, and was bound to and eluted from specific antigen, concanavalin A and lentil lectin, anti-human Ia antibodies, and anti-mouse suppressor factor antibodies, but was not bound to antibodies against human IgG. PMID:6169475

78 FR 79295 - Airworthiness Directives; CFM International S.A. Turbofan Engines

Federal Register 2010, 2011, 2012, 2013, 2014

2013-12-30

... Airworthiness Directives; CFM International S.A. Turbofan Engines AGENCY: Federal Aviation Administration (FAA... International (CFM) S.A. CFM56-3 and CFM56-7B series turbofan engines with certain accessory gearboxes (AGBs... of total loss of engine oil from CFM56 series turbofan engines while in flight. This AD requires an...

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

PubMed Central

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

1986-01-01

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

Identification and Characterization of Tumor Antigens Associated with Breast Cancer

DTIC Science & Technology

1999-08-01

antigens resulted in strong production of anti-envelope and anti- ATRX antibodies (Hampton et al., figure 5). Isotype analysis of the antibody response...antigens Key Words: tumor antigen, endogenous retrovirus, antibody , adenocarcinoma, ATRX ABSTRACT Evaluation of immunotherapy strategies in mouse models...individuals often develop a limited immune response to their tumor. The production of antibodies directed against tumor antigens has been described for

Embryonal carcinoma antigen and the T/t locus of the mouse.

PubMed Central

Kemler, R; Babinet, C; Condamine, H; Gachelin, G; Guenet, J L; Jacob, F

1976-01-01

The presence of the F9 antigen and of four other antigens related to the T/t locus of the mouse was investigated by immunofluorescence on preimplantation embryos. In morulae heterozygous for any of these t haplotypes, both the appropriate t antigen and the F9 antigen are expressed. The F9 antigen segregates among the progeny of crosses producing embryos homozygous for some (tw32 and tw5) but not for other haplotypes. It is concluded that (i) whatever the time of action of a t haplotype, its corresponding antigen is expressed during cleavage and (ii) the F9 antigen is specified by a gene(s) in the region of the T/t locus. Images PMID:1069295

The Gab1 protein is a docking site for multiple proteins involved in signaling by the B cell antigen receptor.

PubMed

Ingham, R J; Holgado-Madruga, M; Siu, C; Wong, A J; Gold, M R

1998-11-13

Gab1 is a member of the docking/scaffolding protein family which includes IRS-1, IRS-2, c-Cbl, p130(cas), and p62(dok). These proteins contain a variety of protein-protein interaction motifs including multiple tyrosine residues that when phosphorylated can act as binding sites for Src homology 2 (SH2) domain-containing signaling proteins. We show in the RAMOS human B cell line that Gab1 is tyrosine-phosphorylated in response to B cell antigen receptor (BCR) engagement. Moreover, tyrosine phosphorylation of Gab1 correlated with the binding of several SH2-containing signaling proteins to Gab1 including Shc, Grb2, phosphatidylinositol 3-kinase, and the SHP-2 tyrosine phosphatase. Far Western analysis showed that the SH2 domains of Shc, SHP-2, and the p85 subunit of phosphatidylinositol 3-kinase could bind directly to tyrosine-phosphorylated Gab1 isolated from activated RAMOS cells. In contrast, the Grb2 SH2 domain did not bind directly to Gab1 but instead to the Shc and SHP-2 associated with Gab1. We also show that Gab1 is present in the membrane-enriched particulate fraction of RAMOS cells and that Gab1/signaling protein complexes are found in this fraction after BCR engagement. Thus, tyrosine-phosphorylated Gab1 may recruit cytosolic signaling proteins to cellular membranes where they can act on membrane-bound targets. This may be a critical step in the activation of multiple BCR signaling pathways.

[Research advances of genomic GYP coding MNS blood group antigens].

PubMed

Liu, Chang-Li; Zhao, Wei-Jun

2012-02-01

The MNS blood group system includes more than 40 antigens, and the M, N, S and s antigens are the most significant ones in the system. The antigenic determinants of M and N antigens lie on the top of GPA on the surface of red blood cells, while the antigenic determinants of S and s antigens lie on the top of GPB on the surface of red blood cells. The GYPA gene coding GPA and the GYPB gene coding GPB locate at the longarm of chromosome 4 and display 95% homologus sequence, meanwhile both genes locate closely to GYPE gene that did not express product. These three genes formed "GYPA-GYPB-GYPE" structure called GYP genome. This review focuses on the molecular basis of genomic GYP and the variety of GYP genome in the expression of diversity MNS blood group antigens. The molecular basis of Miltenberger hybrid glycophorin polymorphism is specifically expounded.

Molecular recognition of microbial lipid-based antigens by T cells.

PubMed

Gras, Stephanie; Van Rhijn, Ildiko; Shahine, Adam; Le Nours, Jérôme

2018-05-01

The immune system has evolved to protect hosts from pathogens. T cells represent a critical component of the immune system by their engagement in host defence mechanisms against microbial infections. Our knowledge of the molecular recognition by T cells of pathogen-derived peptidic antigens that are presented by the major histocompatibility complex glycoproteins is now well established. However, lipids represent an additional, distinct chemical class of molecules that when presented by the family of CD1 antigen-presenting molecules can serve as antigens, and be recognized by specialized subsets of T cells leading to antigen-specific activation. Over the past decades, numerous CD1-presented self- and bacterial lipid-based antigens have been isolated and characterized. However, our understanding at the molecular level of T cell immunity to CD1 molecules presenting microbial lipid-based antigens is still largely unexplored. Here, we review the insights and the molecular basis underpinning the recognition of microbial lipid-based antigens by T cells.

Evaluating the B-cell density with various activation functions using White Noise Path Integral Approach

NASA Astrophysics Data System (ADS)

Aban, C. J. G.; Bacolod, R. O.; Confesor, M. N. P.

2015-06-01

A The White Noise Path Integral Approach is used in evaluating the B-cell density or the number of B-cell per unit volume for a basic type of immune system response based on the modeling done by Perelson and Wiegel. From the scaling principles of Perelson [1], the B- cell density is obtained where antigens and antibodies mutates and activation function f(|S-SA|) is defined describing the interaction between a specific antigen and a B-cell. If the activation function f(|S-SA|) is held constant, the major form of the B-cell density evaluated using white noise analysis is similar to the form of the B-cell density obtained by Perelson and Wiegel using a differential approach.A piecewise linear functionis also used to describe the activation f(|S-SA|). If f(|S-SA|) is zero, the density decreases exponentially. If f(|S-SA|) = S-SA-SB, the B- cell density increases exponentially until it reaches a certain maximum value. For f(|S-SA|) = 2SA-SB-S, the behavior of B-cell density is oscillating and remains to be in small values.

Antigenic Drift of the Pandemic 2009 A(H1N1) Influenza Virus in a Ferret Model

PubMed Central

Guarnaccia, Teagan; Carolan, Louise A.; Maurer-Stroh, Sebastian; Lee, Raphael T. C.; Job, Emma; Reading, Patrick C.; Petrie, Stephen; McCaw, James M.; McVernon, Jodie; Hurt, Aeron C.; Kelso, Anne; Mosse, Jennifer; Barr, Ian G.; Laurie, Karen L.

2013-01-01

Surveillance data indicate that most circulating A(H1N1)pdm09 influenza viruses have remained antigenically similar since they emerged in humans in 2009. However, antigenic drift is likely to occur in the future in response to increasing population immunity induced by infection or vaccination. In this study, sequential passaging of A(H1N1)pdm09 virus by contact transmission through two independent series of suboptimally vaccinated ferrets resulted in selection of variant viruses with an amino acid substitution (N156K, H1 numbering without signal peptide; N159K, H3 numbering without signal peptide; N173K, H1 numbering from first methionine) in a known antigenic site of the viral HA. The N156K HA variant replicated and transmitted efficiently between naïve ferrets and outgrew wildtype virus in vivo in ferrets in the presence and absence of immune pressure. In vitro, in a range of cell culture systems, the N156K variant rapidly adapted, acquiring additional mutations in the viral HA that also potentially affected antigenic properties. The N156K escape mutant was antigenically distinct from wildtype virus as shown by binding of HA-specific antibodies. Glycan binding assays demonstrated the N156K escape mutant had altered receptor binding preferences compared to wildtype virus, which was supported by computational modeling predictions. The N156K substitution, and culture adaptations, have been detected in human A(H1N1)pdm09 viruses with N156K preferentially reported in sequences from original clinical samples rather than cultured isolates. This study demonstrates the ability of the A(H1N1)pdm09 virus to undergo rapid antigenic change to evade a low level vaccine response, while remaining fit in a ferret transmission model of immunization and infection. Furthermore, the potential changes in receptor binding properties that accompany antigenic changes highlight the importance of routine characterization of clinical samples in human A(H1N1)pdm09 influenza surveillance

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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

1985-12-01

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

Management of Men with Prostate-specific Antigen Failure After Prostate Radiotherapy: The Case Against Early Androgen Deprivation.

PubMed

Brand, Douglas; Parker, Chris

2018-04-01

In men with prostate-specific antigen failure after radical radiotherapy, androgen deprivation therapy should be delayed until the site of recurrence is known to allow consideration of curative treatment options, to delay androgen deprivation therapy-related morbidity, and to enable earlier access to abiraterone and docetaxel. Copyright © 2017 European Association of Urology. Published by Elsevier B.V. All rights reserved.

Original antigenic sin responses to influenza viruses.

PubMed

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

2009-09-01

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

Do On-site Mental Health Professionals Change Pediatricians’ Responses to Children’s Mental Health Problems?

PubMed Central

Horwitz, Sarah McCue; Storfer-Isser, Amy; Kerker, Bonnie D.; Szilagyi, Moira; Garner, Andrew S.; O’Connor, Karen G.; Hoagwood, Kimberly E.; Green, Cori M.; Foy, Jane M.; Stein, Ruth E.K.

2016-01-01

Objective The objectives were to: assess the availability of on-site mental health professionals (MHP) in primary care; examine practice/pediatrician characteristics associated with on-site MHPs; and determine whether presence of on-site MHPs is related to pediatricians’ co-managing or more frequently identifying, treat/managing or referring MH problems. Methods Analyses included AAP members who participated in an AAP Periodic Survey in 2013 and who practiced general pediatrics (N=321). Measures included socio-demographics, practice characteristics, questions on about on-site MHPs, co-management of MH problems and pediatricians’ behaviors in response to 5 prevalent MH problems. Weighted univariate, bivariate and multivariable analyses were performed. Results Thirty-five percent reported on-site MHPs. Practice characteristics (medical schools/universities/HMOs, <100 visits/week, <80% of patients privately insured), and interactions of practice location (urban) with visits and patient insurance, were associated with on-site MHPs. There was no overall association between co-location and co-management or whether pediatricians usually identified, treat/managed or referred 5 common child MH problems. Among the subset of pediatricians who reported co-managing there was an association with co-management when the on-site MHP was a child psychiatrist, SA counselor, or social worker. Conclusions On-site MHPs are more frequent in settings where low-income children are served and where pediatricians train. Pediatricians who co-manage MH problems are more likely to do so when the on-site MHP is a child psychiatrist, SA counselor, or social worker. Overall, on-site MHPs were not associated with co-management or increased likelihood of pediatricians identifying, treating/managing, or referring children with 5 common child MH problems. PMID:27064141

Inhibiting DNA methylation activates cancer testis antigens and expression of the antigen processing and presentation machinery in colon and ovarian cancer cells.

PubMed

Siebenkäs, Cornelia; Chiappinelli, Katherine B; Guzzetta, Angela A; Sharma, Anup; Jeschke, Jana; Vatapalli, Rajita; Baylin, Stephen B; Ahuja, Nita

2017-01-01

Innovative therapies for solid tumors are urgently needed. Recently, therapies that harness the host immune system to fight cancer cells have successfully treated a subset of patients with solid tumors. These responses have been strong and durable but observed in subsets of patients. Work from our group and others has shown that epigenetic therapy, specifically inhibiting the silencing DNA methylation mark, activates immune signaling in tumor cells and can sensitize to immune therapy in murine models. Here we show that colon and ovarian cancer cell lines exhibit lower expression of transcripts involved in antigen processing and presentation to immune cells compared to normal tissues. In addition, treatment with clinically relevant low doses of DNMT inhibitors (that remove DNA methylation) increases expression of both antigen processing and presentation and Cancer Testis Antigens in these cell lines. We confirm that treatment with DNMT inhibitors upregulates expression of the antigen processing and presentation molecules B2M, CALR, CD58, PSMB8, PSMB9 at the RNA and protein level in a wider range of colon and ovarian cancer cell lines and treatment time points than had been described previously. In addition, we show that DNMTi treatment upregulates many Cancer Testis Antigens common to both colon and ovarian cancer. This increase of both antigens and antigen presentation by epigenetic therapy may be one mechanism to sensitize patients to immune therapies.

Inhibition of CD1 antigen presentation by human cytomegalovirus.

PubMed

Raftery, Martin J; Hitzler, Manuel; Winau, Florian; Giese, Thomas; Plachter, Bodo; Kaufmann, Stefan H E; Schönrich, Günther

2008-05-01

The betaherpesvirus human cytomegalovirus (HCMV) encodes several molecules that block antigen presentation by the major histocompatibility complex (MHC) proteins. Humans also possess one other family of antigen-presenting molecules, the CD1 family; however, the effect of HCMV on CD1 expression is unknown. The majority of CD1 molecules are classified on the basis of homology as group 1 CD1 and are present almost exclusively on professional antigen-presenting cells such as dendritic cells, which are a major target for HCMV infection and latency. We have determined that HCMV encodes multiple blocking strategies targeting group 1 CD1 molecules. CD1 transcription is strongly inhibited by the HCMV interleukin-10 homologue cmvIL-10. HCMV also blocks CD1 antigen presentation posttranscriptionally by the inhibition of CD1 localization to the cell surface. This function is not performed by a known HCMV MHC class I-blocking molecule and is substantially stronger than the blockage induced by herpes simplex virus type 1. Antigen presentation by CD1 is important for the development of the antiviral immune response and the generation of mature antigen-presenting cells. HCMV present in antigen-presenting cells thus blunts the immune response by the blockage of CD1 molecules.

Ammonia Offgassing from SA9T

NASA Technical Reports Server (NTRS)

Monje, Oscar; Nolek, Sara D.; Wheeler, Raymond M.

2011-01-01

NH3 is a degradation product of SA9T, a solid-amine sorbent developed by Hamilton Sundstrand, that is continually emitted into the gas stream being conditioned by this sorbent. NH3 offgassing rates were measured using FTIR spectroscopy using a packed bed at similar contact times as offgassing tests conducted at Hamilton Sundstrand and at the Ames Research Center. The bed was challenged with moist air at several flow rates and humidities and NH3 concentration of the effluent was measured for several hours. The NH3 offgassing rates in open-loop testing were calculated from the steady state outlet NH3 concentration and flow rate. NH3 offgassing rates from SA9T were found to be influenced by the contact time with the adsorbent (flow rate) and by the humidity of the inlet gas stream, which are consistent with previous studies. Closed-loop vacuum-swing adsorption cycling rates verified that NH3 offgassing continues when a constant source of water vapor is present.

Impact of obesity on the predictive accuracy of prostate-specific antigen density and prostate-specific antigen in native Korean men undergoing prostate biopsy.

PubMed

Kim, Jae Heon; Doo, Seung Whan; Yang, Won Jae; Lee, Kwang Woo; Lee, Chang Ho; Song, Yun Seob; Jeon, Yoon Su; Kim, Min Eui; Kwon, Soon-Sun

2014-10-01

To evaluate the impact of obesity on the biopsy detection of prostate cancer. We retrospectively reviewed data of 1182 consecutive Korean patients (≥50 years) with serum prostate-specific antigen levels of 3-10 ng/mL who underwent initial extended 12-cores biopsy from September 2009 to March 2013. Patients who took medications that were likely to influence the prostate-specific antigen level were excluded. Receiver operating characteristic curves were plotted for prostate-specific antigen and prostate-specific antigen density predicting cancer status among non-obese and obese men. A total of 1062 patients (mean age 67.1 years) were enrolled in the analysis. A total of 230 men (21.7%) had a positive biopsy. In the overall study sample, the area under the receiver operator characteristic curve of serum prostate-specific antigen for predicting prostate cancer on biopsy were 0.584 and 0.633 for non-obese and obese men, respectively (P = 0.234). However, the area under the curve for prostate-specific antigen density in predicting cancer status showed a significant difference (non-obese 0.696, obese 0.784; P = 0.017). There seems to be a significant difference in the ability of prostate-specific antigen density to predict biopsy results between non-obese and obese men. Obesity positively influenced the overall ability of prostate-specific antigen density to predict prostate cancer. © 2014 The Japanese Urological Association.

Ride through SaFIRES : lessons learned from SaFIRES - an APTS operational test in Prince William County, Virginia.

DOT National Transportation Integrated Search

2002-06-01

This interim report documents the lessons learned to date from the Smart Flex-route Integrated Real-time Enhancement System (SaFIRES) operational test in Prince William County, Virginia. This route deviation service has proven to be popular with Coun...

Prostate-Specific Antigen (PSA) Test: MedlinePlus Lab Test Information

MedlinePlus

... gov/labtests/prostatespecificantigenpsatest.html Prostate-Specific Antigen (PSA) Test To use the sharing features on this page, ... JavaScript. What is a prostate-specific antigen (PSA) test? A prostate-specific antigen (PSA) test measures the ...

Conjugating influenza a (H1N1) antigen to n-trimethylaminoethylmethacrylate chitosan nanoparticles improves the immunogenicity of the antigen after nasal administration.

PubMed

Liu, Qingfeng; Zheng, Xiaoyao; Zhang, Chi; Shao, Xiayan; Zhang, Xi; Zhang, Qizhi; Jiang, Xinguo

2015-11-01

As one of the most serious infectious respiratory diseases, influenza A (H1N1) is a great threat to human health, and it has created an urgent demand for effective vaccines. Nasal immunization can induce both systemic and mucosal immune responses against viruses, and it can serve as an ideal route for vaccination. However, the low immunogenicity of antigens on nasal mucosa is a high barrier for the development of nasal vaccines. In this study, we covalently conjugated an influenza A (H1N1) antigen to the surface of N-trimethylaminoethylmethacrylate chitosan (TMC) nanoparticles (H1N1-TMC/NP) through thioester bonds to increase the immunogenicity of the antigen after nasal administration. SDS-PAGE revealed that most of the antigen was conjugated on TMC nanoparticles, and an in vitro biological activity assay confirmed the stability of the antigen after conjugation. After three nasal immunizations, the H1N1-TMC/NP induced significantly higher levels of serum IgG and mucosal sIgA compared with free antigen. A hemagglutination inhibition assay showed that H1N1-TMC/NP induced much more protective antibodies than antigen-encapsulated nanoparticles or alum-precipitated antigen (I.M.). In the mechanistic study, H1N1-TMC/NP was shown to stimulate macrophages to produce IL-1β and IL-6 and to stimulate spleen lymphocytes to produce IL-2 and IFN-γ. These results indicated that H1N1-TMC/NP may be an effective vaccine against influenza A (H1N1) viruses for use in nasal immunization. © 2015 Wiley Periodicals, Inc.

Mannose-pepstatin conjugates as targeted inhibitors of antigen processing.

PubMed

Free, Paul; Hurley, Christopher A; Kageyama, Takashi; Chain, Benjamin M; Tabor, Alethea B

2006-05-07

The molecular details of antigen processing, including the identity of the enzymes involved, their intracellular location and their substrate specificity, are still incompletely understood. Selective inhibition of proteolytic antigen processing enzymes such as cathepsins D and E, using small molecular inhibitors such as pepstatin, has proven to be a valuable tool in investigating these pathways. However, pepstatin is poorly soluble in water and has limited access to the antigen processing compartment in antigen presenting cells. We have synthesised mannose-pepstatin conjugates, and neomannosylated BSA-pepstatin conjugates, as tools for the in vivo study of the antigen processing pathway. Conjugation to mannose and to neomannosylated BSA substantially improved the solubility of the conjugates relative to pepstatin. The mannose-pepstatin conjugates showed no reduction in inhibition of cathepsin E, whereas the neomannosylated BSA-pepstatin conjugates showed some loss of inhibition, probably due to steric factors. However, a neomannosylated BSA-pepstatin conjugate incorporating a cleavable disulfide linkage between the pepstatin and the BSA showed the best uptake to dendritic cells and the best inhibition of antigen processing.

The three-dimensional structure of a T-cell antigen receptor V alpha V beta heterodimer reveals a novel arrangement of the V beta domain.

PubMed Central

Housset, D; Mazza, G; Grégoire, C; Piras, C; Malissen, B; Fontecilla-Camps, J C

1997-01-01

The crystal structure of a mouse T-cell antigen receptor (TCR) Fv fragment complexed to the Fab fragment of a specific anti-clonotypic antibody has been determined to 2.6 A resolution. The polypeptide backbone of the TCR V alpha domain is very similar to those of other crystallographically determined V alphas, whereas the V beta structure is so far unique among TCR V beta domains in that it displays a switch of the c" strand from the inner to the outer beta-sheet. The beta chain variable region of this TCR antigen-binding site is characterized by a rather elongated third complementarity-determining region (CDR3beta) that packs tightly against the CDR3 loop of the alpha chain, without leaving any intervening hydrophobic pocket. Thus, the conformation of the CDR loops with the highest potential diversity distinguishes the structure of this TCR antigen-binding site from those for which crystallographic data are available. On the basis of all these results, we infer that a significant conformational change of the CDR3beta loop found in our TCR is required for binding to its cognate peptide-MHC ligand. PMID:9250664

Experimental infection of octopus vulgaris (Cuvier, 1797) with Photobacterium damsela subsp. piscicida. Immunohistochemical tracking of antigen and tissue responses.

PubMed

Bakopoulos, Vasileios; White, Daniella; Valsamidis, Michail-Aggelos; Vasilaki, Feli

2017-03-01

Adult common octopus individuals were intramuscularly infected with Photobacterium damsela subsp. piscicida in order to investigate if this species is sensitive to this common and important fish pathogen. The fate of the bacterial antigens and the tissue responses of Octopus vulgaris were studied employing immunohistochemical techniques. Strong reaction at the site of injection was evident from day 2 post-infection that continued until day 14. Great numbers of hemocytes that were attracted at the site of infection were involved in phagocytosis of bacteria. Very early in the infection, a transition of cells to fibroblasts and an effort to isolate the infection was observed. During the course of the study, very large necrotic cells were seen at the site of infection, whereas during the later stages hemocytes with phagocytosed bacteria were observed in well-defined pockets inside the muscle tissue. None of the internal organs tested for the presence of the bacterium were positive with the exception of the digestive gland where antigen staining was observed which was not associated with hemocyte infiltration. The high doses of bacterial cells used in this experimental infection and the lack of disease signs from Octopus vulgaris suggest that, under normal conditions, octopus is resistant to Photobacterium damsela subsp. piscicida. Copyright © 2017 Elsevier Inc. All rights reserved.

75 FR 53857 - Airworthiness Directives; Eurocopter France Model SA330J Helicopters

Federal Register 2010, 2011, 2012, 2013, 2014

2010-09-02

... Airworthiness Directives; Eurocopter France Model SA330J Helicopters AGENCY: Federal Aviation Administration... known U.S. owners and operators of Eurocopter France (Eurocopter) Model SA330J helicopters by individual...'' rather than checking for ``play.'' This helicopter model is manufactured in France and is type...

A Molecular Simulation Study of Antibody-Antigen Interactions on Surfaces for the Rational Design of Next-Generation Antibody Microarrays

NASA Astrophysics Data System (ADS)

Bush, Derek B.

Antibody microarrays constitute a next-generation sensing platform that has the potential to revolutionize the way that molecular detection is conducted in many scientific fields. Unfortunately, current technologies have not found mainstream use because of reliability problems that undermine trust in their results. Although several factors are involved, it is believed that undesirable protein interactions with the array surface are a fundamental source of problems where little detail about the molecular-level biophysics are known. A better understanding of antibody stability and antibody-antigen binding on the array surface is needed to improve microarray technology. Despite the availability of many laboratory methods for studying protein stability and binding, these methods either do not work when the protein is attached to a surface or they do not provide the atomistic structural information that is needed to better understand protein behavior on the surface. As a result, molecular simulation has emerged as the primary method for studying proteins on surfaces because it can provide metrics and views of atomistic structures and molecular motion. Using an advanced, coarse-grain, protein-surface model this study investigated how antibodies react to and function on different types of surfaces. Three topics were addressed: (1) the stability of individual antibodies on surfaces, (2) antibody binding to small antigens while on a surface, and (3) antibody binding to large antigens while on a surface. The results indicate that immobilizing antibodies or antibody fragments in an upright orientation on a hydrophilic surface can provide the molecules with thermal stability similar to their native aqueous stability, enhance antigen binding strength, and minimize the entropic cost of binding. Furthermore, the results indicate that it is more difficult for large antigens to approach the surface than small antigens, that multiple binding sites can aid antigen binding, and that

Antigenic evolution of H9N2 chicken influenza viruses isolated in China during 2009–2013 and selection of a candidate vaccine strain with broad cross-reactivity

PubMed Central

Wei, Yandi; Xu, Guanlong; Zhang, Guozhong; Wen, Chu; Anwar, Furkat; Wang, Shuoguo; Lemmon, Gordon; Wang, Jinliang; Carter, Robert; Wang, Min; Sun, Honglei; Sun, Yipeng; Zhao, Jixun; Wu, Gang; Webster, Robert G.; Liu, Jinhua; Pu, Juan

2016-01-01

We previously demonstrated that H9N2 subtype avian influenza viruses (AIVs) isolated from 1994 to 2008 evolved into distinct antigenic groups (C, D, and E) and then underwent antigenic drift from commercial vaccines, causing a country-wide outbreak during 2010–2013. In this study, H9N2 AIVs isolated from chickens during 2009–2013 were antigenically analyzed by performing hemagglutination inhibition and neutralization assays using a panel of polyclonal antibodies. Our findings confirmed the antigenic drift of recent H9N2 viruses from the commercial vaccine and showed that most of these antigenic variants form a novel HI antigenic group, F, with a few belonging to groups D and E. Slight antigenic variation was observed in group F viruses. Genetic analysis of amino acid sequences deduced from hemagglutinin (HA) gene sequences indicated that 9 of 15 mutations predominant in the 2009–2013 viruses can be mapped to known antigenic sites, which might be responsible for the novel antigenicity of group F. These antigenic changes make it necessary to modify the influenza vaccine to ensure efficient protection. A vaccine candidate, Ck/HeB/YT/10, was selected and provided significant protection against viruses from different antigenic groups in terms of reduction in virus shedding, suggesting broad cross-reactivity. Taken together, our results indicate that the H9N2 chicken influenza viruses in China have evolved from distinct antigenic groups into a novel group F that became dominant during the country-wide outbreak and now seems to be undergoing new antigenic divergence. Systematic surveillance and timely updating of vaccine strains are important for viral prevention and control in the future. PMID:26711021

Antigen recognition by H-2-restricted T cells. I. Cell-free antigen processing

PubMed Central

1983-01-01

We examined the ability of a set of cloned chicken ovalbumin (cOVA)- specific, Id-restricted, T cell hybridomas to produce interleukin-2 in response to cOVA presented by the Ia+ B cell lymphoma line, A20-2J. Although viable A20-2J cells presented native, denatured, and fragmented cOVA more or less equally well, A20-2J cells that were glutaraldehyde-fixed could present only enzymatically or chemically fragmented cOVA. These results suggest that antigen fragmentation may be both necessary and sufficient to define accessory cell processing of soluble antigens so that they may be recognized in association with I- region molecules by T cells. PMID:6193218

Use of antibodies specific to defined regions of scorpion. cap alpha. -toxin to study its interaction with its receptor site on the sodium channel

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ayeb, M.E.; Bahraoui, E.M.; Granier, C.

1986-10-21

Five antibody populations selected by immunoaffinity chromatography for the specificity toward various regions of toxin II of the scorpion Androctonus australis Hector were used to probe the interaction of this protein with its receptor site on the sodium channel. These studies indicate that two antigenic sites, one located around the disulfide bridge 12-63 and one encompassing residues 50-59, are involved in the molecular mechanisms of toxicity neutralization. Fab fragments specific to the region around disulfide bridge 12-63 inhibit binding of the /sup 125/I-labeled toxin to its receptor site. Also, these two antigenic regions are inaccessible to the antibodies when themore » toxin is bound to its receptor site. In contrast, the two other antigenic sites encompassing the only ..cap alpha..-helix region (residues 23-32) and a ..beta..-turn structure (residues 32-35) are accessible to the respective antibodies when the toxin is bound to its receptor. Together, these data support the recent proposal that a region made of residues that are conserved in the scorpion toxin family is involved in the binding of the toxin to the receptor.« less

Imaging of blood antigen distribution on blood cells by thermal lens microscopy

NASA Astrophysics Data System (ADS)

Kimura, Hiroko; Sekiguchi, Kazuya; Nagao, Fumiko; Mukaida, Masahiro; Kitamori, Takehiko; Sawada, Tsuguo

2000-05-01

Blood group antigens on a cell were measured by a new microscopic method, i.e. thermal lens microscopy which involves spectrometry using a laser-induced thermal-lens effect. The blood group antigen was immunologically stained using antibody labeled with colloidal gold. Human leukocyte antigens (HLA) on lymphocytes and mononuclear leukocytes were observed by the thermal lens microscope, and Lewis blood group antigens on erythrocytes and polymorphonuclear leukocytes were also observed. The antigen distribution on each cell-surface was imaged using this technique. In spite of convex surface of living cells, colloidal gold was correctly quantified by adjusting the deviation of the focal point of the probe laser by the phase of the signal. In the measurement of leukocyte antigens, antigens of HLA-A, -B, -C loci on the lymphocytes were identified and quantitated by using a single cell. The image of HLA-A, -B, -C antigen distribution on a mononuclear leukocyte was obtained. In the measurement of erythrocyte antigens, a small quantity of Lewis antigens was detected on the cord erythrocytes. Localized small quantities of membrane antigens are better quantitated without extraction or cytolysis. Our thermal lens microscope is a powerful and highly sensitive analytical tool for detecting and quantitating localized antigens in single cells and/or cell-surface-associated molecules.

Comprehensive definition of human immunodominant CD8 antigens in tuberculosis.

PubMed

Lewinsohn, Deborah A; Swarbrick, Gwendolyn M; Park, Byung; Cansler, Meghan E; Null, Megan D; Toren, Katelynne G; Baseke, Joy; Zalwango, Sarah; Mayanja-Kizza, Harriet; Malone, LaShaunda L; Nyendak, Melissa; Wu, Guanming; Guinn, Kristi; McWeeney, Shannon; Mori, Tomi; Chervenak, Keith A; Sherman, David R; Boom, W Henry; Lewinsohn, David M

2017-01-01

Despite widespread use of the Bacillus Calmette-Guerin vaccine, tuberculosis, caused by infection with Mycobacterium tuberculosis , remains a leading cause of morbidity and mortality worldwide. As CD8 + T cells are critical to tuberculosis host defense and a phase 2b vaccine trial of modified vaccinia Ankara expressing Ag85a that failed to demonstrate efficacy, also failed to induce a CD8 + T cell response, an effective tuberculosis vaccine may need to induce CD8 + T cells. However, little is known about CD8, as compared to CD4, antigens in tuberculosis. Herein, we report the results of the first ever HLA allele independent genome-wide CD8 antigen discovery program. Using CD8 + T cells derived from humans with latent tuberculosis infection or tuberculosis and an interferon-γ ELISPOT assay, we screened a synthetic peptide library representing 10% of the Mycobacterium tuberculosis proteome, selected to be enriched for Mycobacterium tuberculosis antigens. We defined a set of immunodominant CD8 antigens including part or all of 74 Mycobacterium tuberculosis proteins, only 16 of which are previously known CD8 antigens. Immunogenicity was associated with the degree of expression of mRNA and protein. Immunodominant antigens were enriched in cell wall proteins with preferential recognition of Esx protein family members, and within proteins comprising the Mycobacterium tuberculosis secretome. A validation study of immunodominant antigens demonstrated that these antigens were strongly recognized in Mycobacterium tuberculosis -infected individuals from a tuberculosis endemic region in Africa. The tuberculosis vaccine field will likely benefit from this greatly increased known repertoire of CD8 immunodominant antigens and definition of properties of Mycobacterium tuberculosis proteins important for CD8 antigenicity.