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Röske, Kerstin, Alain Blanchard, Isabelle Chambaud, Christine Citti, Jürgen H. Helbig, Marie-Christine Prevost, Renate Rosengarten und Enno Jacobs. „Phase Variation among Major Surface Antigens ofMycoplasma penetrans“. Infection and Immunity 69, Nr. 12 (01.12.2001): 7642–51. http://dx.doi.org/10.1128/iai.69.12.7642-7651.2001.
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ABSTRACT The pathogenicity and prevalence of Mycoplasma penetrans, a Mycoplasma species recently isolated from humans, are still debated. A major P35 antigen, which is used as target epitope in serological assays, was shown to be a phase-variable lipid-associated membrane protein (LAMP). In this study, we performed a comparative analysis of the LAMP patterns from fiveM. penetrans clinical isolates and from the type strain. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles and immunoblots with sera serially collected from an M. penetrans-infected patient indicated that these strains expressed different LAMP repertoires. Furthermore, the intraclonal variation in the expression of LAMPs (P34A, P34B, P35, and P38) was monitored by immunoblot analysis with three specific monoclonal antibodies (MAbs) developed in this study and MAb 7 to P35. The phase variation of these LAMPs occurs in an independent manner, with frequencies of variation ranging from 10−2 to 10−4 per cell per generation. Consistent with their amphipathic nature, the P34B and P38 antigens were found exposed at the cell surface. The DNA sequence encoding the P38 antigen was defined and found to be related to those of the P35 gene and other putative LAMP-encoding genes, suggesting that these variable antigens are encoded by a family of related genes. Finally, the serum samples from an M. penetrans-infected patient contained antibodies that reacted with a P36 antigen expressed in different M. penetrans strains but not in the isolate recovered from this patient. This result suggested that in vivo phase variation of P36 occurred, which would support a role for these LAMP variations in avoiding the host's immune vigilance.
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Beem, J. E., W. B. Clark und A. S. Bleiweis. „Antigenic Variation of Indigenous Streptococci“. Journal of Dental Research 64, Nr. 8 (August 1985): 1039–45. http://dx.doi.org/10.1177/00220345850640080301.
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Isolates of Group D streptococci indigenous to the murine oral cavity were studied to detect the occurrence of antigenic variation. Group D streptococci cultured from molar homogenates of Balb/c mice were randomly selected for study on the basis of distinctive colony morphology. Isolates obtained over a 12-week period were biotyped using the API 20S system, and subjected to Lancefield extraction and rocket immunoelectrophoresis for serotyping. All isolates were compared with an arbitrarily selected standard test strain (W1S-1) isolated the first week of the first experimental series. Four biotypes were encountered during the first week of two experimental series. Two very unusual biotypes detected during the first experimental series persisted throughout that series, as did two more common biotypes throughout the second experimental series. Anti-W1S-1 serum produced three precipitin bands (antigens O, D, and K) against WIS-1 Lancefield extract and against the respective biotypes detected during the first week of the two series. Of the three antigens detected, only the group antigen (D) did not vary during either experimental series. Antigenic variants lacking the O or K antigen and bearing these distinctive phenotypes were repeatedly isolated in subsequent weeks. Ultimately, 16% of 190 strains isolated during the first series and 26% of 167 strains isolated during the second series proved to be antigenic variants of the predominant biotypes detected in both series.
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Li, Qun, Matthew Hobbs und Peter R. Reeves. „The variation of dTDP-l-rhamnose pathway genes in Vibrio cholerae“. Microbiology 149, Nr. 9 (01.09.2003): 2463–74. http://dx.doi.org/10.1099/mic.0.26382-0.
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The genetic variation in the dTDP-l-rhamnose pathway genes (rmlA, rmlB, rmlC and rmlD) in Vibrio cholerae was investigated. The genes are part of the O antigen gene cluster and the aim was to study lateral gene transfer of O antigen gene clusters. The rml genes of an O6 strain were cloned using an Escherichia coli K-12 strain designed for selecting cloned rml genes. Thirty-three strains carrying the known rhamnose-containing O antigens were probed with O6-based rml gene probes, and 19 were positive with from one to all four of the gene probes. Nine rml gene sets from this group were sequenced and found to be in the order rmlBADC, at the 5′ end of the gene clusters. A gradient in the level of variation was observed, with highly similar sequences at the 5′ end rmlB gene, but very divergent and strain-specific sequences at the 3′ end of the rml gene set. The change in level of similarity varied in position, but was always abrupt and coincided with a change in GC content, indicating that the 5′ and 3′ parts are of different origin, and that recombination within rml genes has occurred. The rml gene sets of two of the strains that did not hybridize with any O6 rml gene probes were also cloned and sequenced. Both gene sets were in the middle of the O antigen gene cluster and were very divergent from each other and all other rml gene sets. This supports the hypothesis that presence of rml genes at the end of the O antigen gene cluster facilitates lateral gene transfer of rml-containing O antigen gene clusters in V. cholerae. The sequence relationships make it possible to identify sites of recombination and to distinguish DNA that has long been in V. cholerae and DNA that probably came into the species with the O antigen gene cluster.
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IGLESIAS, R., A. PARAMÁ, M. F. ÁLVAREZ, J. LEIRO, F. M. UBEIRA und M. L. SANMARTÍN. „Philasterides dicentrarchi (Ciliophora: Scuticociliatida) expresses surface immobilization antigens that probably induce protective immune responses in turbot“. Parasitology 126, Nr. 2 (Februar 2003): 125–34. http://dx.doi.org/10.1017/s0031182002002688.
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Philasterides dicentrarchi is a histophagous ciliate causing systemic scuticociliatosis in cultured turbot. This study demonstrates that turbot which survive this disease have serum antibodies that recognize ciliary antigens of this ciliate in ELISA and immobilize/agglutinate the ciliate in vitro. Mouse sera raised against ciliary antigens and integral membrane proteins are likewise capable of immobilizing/agglutinating the ciliates, indicating that P. dicentrarchi, like other ciliates, expresses surface immobilization antigens. Furthermore, the antigen agglutinating reaction induces the parasite to shed its surface antigens rapidly, replacing them with others with different specific serology. This antigen shedding and variation response is similar to that detected in other protozoan parasites. Immunization of turbot with ciliate lysate plus adjuvant or with formalin-fixed ciliates induced synthesis of agglutinating antibodies and conferred a degree of protection against challenge infection, suggesting that the response to surface antigens may play an important role in defence against this pathogen, SDS–PAGE and immunoblotting studies indicated the existence of a predominant polypeptide of about 38 kDa in the ciliary antigen and membrane protein fractions, and this may be the principal surface antigen of P. dicentrarchi.
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Bertina, R. M. „An International Collaborative Study on the Performance of Protein C Antigen Assays“. Thrombosis and Haemostasis 57, Nr. 01 (1987): 112–17. http://dx.doi.org/10.1055/s-0038-1651074.
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SummaryAn international collaborative study was undertaken to evaluate the performance and specificity of protein C antigen (PC) assays. Thirteen lyophilized plasma samples were distributed among 17 laboratories and analysed with 24 methods. No statistically significant results were obtained with the different methods in plasmas containing only the protein C zymogen. ELISA’s, RIA’s and IRMA’s were found to be more sensitive than the electro-immunoassay. In plasmas of patients on oral anticoagulant treatment ELISA methods tend to give lower PC antigen levels than the electro-immunoassay. Complexes between activated protein C (APC) and the protein C inhibitor (PCI), when present in plasma together with PC zymogen, are detected with 100% efficiency in the electro-immunoassay, with 50% efficiency in the ELISA, and with <10% efficiency in the RIA or in assays using monoclonal antibodies against PC.Mean coefficient of variation was calculated to be 22%, and could be reduced - especially in case of the ELISA by normalisation. Within laboratory variation was calculated to be 11.7% and between laboratory variation 17.8%.
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Marcotuillo, Michelle, Vicki Johnson, Richard J. Jenny und Ryan H. Dorfman. „Murine Reagents and Analytical Tools to Enhance the Utility of Mouse Models for the Study of Human Thrombotic Disorders and Disease States.“ Blood 104, Nr. 11 (16.11.2004): 3989. http://dx.doi.org/10.1182/blood.v104.11.3989.3989.
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Abstract The mechanisms that govern blood coagulation and fibrinolysis have been studied extensively by employing a variety of experimental techniques and in-vivo model systems. Murine models are a valuable resource to test new strategies in the areas of thrombosis and haemostasis. Because both normal and genetically altered mice are available, the murine models are ideal for this application. Murine models are in prevalent use and there exists a limited number of reagents available to assist the researcher. Development of murine reagents and analytical tools will enhance the utility of murine coagulation models by allowing researchers to acquire appropriate standards for characterization, validation, and screening of murine model systems. Recently, we have purified and characterized a group of nine mouse plasma proteins including the enzymatic forms of many of these proteins. A total of eight new polyclonal antibodies to murine antigens were generated including sheep a-mouse plasminogen and sheep anti-mouse factor X. Here we report on the development of two competitive based ELISA’s for the quantitation of mouse plasminogen and mouse factor X. The assay format utilizes purified antigen (plasminogen and factor X) standard. A standard curve is generated based upon the competition between fluid-phase antigen and an antigen-biotin conjugate for an immobilized affinity purified polyclonal antibody (sheep a-mouse plasminogen and sheep anti-mouse factor X). Solid-phase antibody/biotinyl-antigen complexes are detected using avidin-peroxidase. A colormetric signal, which is inversely proportional to the amount of antigen in the fluid-phase, is obtained following the addition of chromogenic substrate. Calibration curves are generated for an immobilized antibody ELISA by plotting the absorbance at 490 nm versus the concentration of non-biotin labeled antigen standard. Biotinylated-antigens were used at single concentrations of 50 ng/ml and 10 ng/ml for factor X and plasminogen respectively. In both cases, immobilized antibodies were adsorbed to microtiter plates at 5ug/ ml. Linearity of the factor X assay extends from approximately 10 ng/ml to 1000 ng/ml. The intra-assay coefficient of variation (CV) is ≤1.1 %, while the inter-assay CV is ≤5 %. Linearity of the plasminogen assay extends from approximately 0.1 μg/ml to 10 μg/ml. The intra-assay coefficient of variation (CV) is ≤2.0 %, while the inter-assay CV is ≤6.3 %. Preliminary studies have demonstrated the feasibility of these assays. With further testing and analysis these assays may prove useful to the researcher attempting to quantitate mouse plasma concentrations of these analytes.
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Thornton, Emily E., Mark R. Looney, Oishee Bose, Debasish Sen, Dean Sheppard, Richard Locksley, Xiaozhu Huang und Matthew F. Krummel. „Spatiotemporally separated antigen uptake by alveolar dendritic cells and airway presentation to T cells in the lung“. Journal of Experimental Medicine 209, Nr. 6 (14.05.2012): 1183–99. http://dx.doi.org/10.1084/jem.20112667.
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Asthma pathogenesis is focused around conducting airways. The reasons for this focus have been unclear because it has not been possible to track the sites and timing of antigen uptake or subsequent antigen presentation to effector T cells. In this study, we use two-photon microscopy of the lung parenchyma and note accumulation of CD11b+ dendritic cells (DCs) around the airway after allergen challenge but very limited access of these airway-adjacent DCs to the contents of the airspace. In contrast, we observed prevalent transepithelial uptake of particulate antigens by alveolar DCs. These distinct sites are temporally linked, as early antigen uptake in alveoli gives rise to DC and antigen retention in the airway-adjacent region. Antigen-specific T cells also accumulate in the airway-adjacent region after allergen challenge and are activated by the accumulated DCs. Thus, we propose that later airway hyperreactivity results from selective retention of allergen-presenting DCs and antigen-specific T cells in airway-adjacent interaction zones, not from variation in the abilities of individual DCs to survey the lung.
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Saeed, Mohd, Vikas Kushwaha, Syed Mohd Faisal, Richa Verma, Irfan Ahmad, Huma Mustafa, Magdah Ganash, Mohammad Amjad Kamal und Ghulam Md Ashraf. „A Study on Serological Reactivity Profile of Different Antigen Preparations with Bancroftian filariasis Human Infection Sera“. Protein & Peptide Letters 27, Nr. 9 (15.10.2020): 841–50. http://dx.doi.org/10.2174/0929866527666200225123534.
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Background: Lymphatic Filariasis (LF) is one of the incapacitating and mosquito-borne sicknesses that on progression may prompt a few recognizable types of clutters like extreme lymphedema, hydrocele, and elephantiasis. Methods: Antigenic preparations of B. malayi adult (BmA), S. cervi adult parasites and microfilariae (mf) total parasite extract were used to analyze the serological reactivity profile with human infectious sera collected from endemic areas of Bancroftian filariasis by performing Western blot and ELISA analysis. Sera from healthy human subjects were also included in the study to determine the variation incurred in the reactivity due to the filariasis infection. Gelelectrophoresis analysis of the crude-extract of BmA revealed seven protein bands while more than ten bands were recognized in S. cervi. Results: our results represent a clear variation in protein patterns among the crude-antigens. ELISA results showed highest prevalence of IgG, IgM and IgG4 antibodies against all antigen preparations when recorded among microfilaraemic chronic infected patients. In both the antigenic preparations, the positive reactions were in the order of microfilaraemic>endemic normal>chronic>acute>nonendemic normal subjects. All sera of Mf+ patients were uniformly positive, while sera of both chronic and endemic normal subjects showed less reactivity. Conclusion: In the present study, we endeavoured to establish the extent of cross-reactivity of antigens derived from animal filarial parasites such as B. malayi and S. cervi with W. bancrofti filariasis sera of human patients. Besides, we further analyzed antibody-isotype profile of IgG, IgG4 and IgM in various human infection sera of bancroftian filarial subjects reactive to heterologous parasite antigens derived from adult worms of S. cervi from bovine and B. malayi from bovine and jirds.
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Horino, Atsuko, Yuko Sasaki, Tsuguo Sasaki und Tsuyoshi Kenri. „Multiple Promoter Inversions Generate Surface Antigenic Variation in Mycoplasma penetrans“. Journal of Bacteriology 185, Nr. 1 (01.01.2003): 231–42. http://dx.doi.org/10.1128/jb.185.1.231-242.2003.
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ABSTRACT Mycoplasma penetrans is a newly identified species of the genus Mycoplasma. It was first isolated from a urine sample from a human immunodeficiency virus (HIV)-infected patient. M. penetrans changes its surface antigen profile with high frequency. The changes originate from ON↔OFF phase variations of the P35 family of surface membrane lipoproteins. The P35 family lipoproteins are major antigens recognized by the human immune system during M. penetrans infection and are encoded by the mpl genes. Phase variations of P35 family lipoproteins occur at the transcriptional level of mpl genes; however, the precise genetic mechanisms are unknown. In this study, the molecular mechanisms of surface antigen profile change in M. penetrans were investigated. The focus was on the 46-kDa protein that is present in M. penetrans strain HF-2 but not in the type strain, GTU. The 46-kDa protein was the product of a previously reported mpl gene, pepIMP13, with an amino-terminal sequence identical to that of the P35 family lipoproteins. Nucleotide sequencing analysis of the pepIMP13 gene region revealed that the promoter-containing 135-bp DNA of this gene had the structure of an invertible element that functioned as a switch for gene expression. In addition, all of the mpl genes of M. penetrans HF-2 were identified using the whole-genome sequence data that has recently become available for this bacterium. There are at least 38 mpl genes in the M. penetrans HF-2 genome. Interestingly, most of these mpl genes possess invertible promoter-like sequences, similar to those of the pepIMP13 gene promoter. A model for the generation of surface antigenic variation by multiple promoter inversions is proposed.
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Umezawa, Eufrosina S., Sueli F. Bastos, Mario E. Camargo, Luci M. Yamauchi, Márcia R. Santos, Antonio Gonzalez, Bianca Zingales et al. „Evaluation of Recombinant Antigens for Serodiagnosis of Chagas’ Disease in South and Central America“. Journal of Clinical Microbiology 37, Nr. 5 (1999): 1554–60. http://dx.doi.org/10.1128/jcm.37.5.1554-1560.1999.
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The commercially available diagnostic tests for Chagas’ disease employ whole extracts or semipurified fractions ofTrypanosoma cruzi epimastigotes. Considerable variation in the reproducibility and reliability of these tests has been reported by different research laboratories, mainly due to cross-reactivity with other pathogens and standardization of the reagents. The use of recombinant antigens for the serodiagnosis of Chagas’ disease is recommended to increase the sensitivity and specificity of serological tests. Expressed in Escherichia coli, as fusion products with glutathione S-transferase, six T. cruzirecombinant antigens (H49, JL7, A13, B13, JL8, and 1F8) were evaluated in an enzyme-linked immunosorbent assay for Chagas’ disease. The study was carried out with a panel of 541 serum samples of chagasic and nonchagasic patients from nine countries of Latin America (Argentina, Bolivia, Brazil, Chile, Colombia, El Salvador, Guatemala, Honduras, and Venezuela). The optimal concentration of each recombinant antigen for coating of plates was determined with the help of125I-labelled recombinant proteins. While the specificity of the epimastigote antigen was 84% because of false positives from leishmaniasis cases, for the recombinant antigens it varied from 96.2 to 99.6%. Recombinant antigens reacted with 79 to 100% of serum samples from chronic chagasic patients. In this way, it is proposed that a mixture of a few T. cruzi recombinant antigens should be employed in a diagnostic kit to minimize individual variation and promote high sensitivity in the diagnosis of Chagas’ disease.
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Sharma, Prajna, Pavithra K und Sanchita Shettigar. „Trends in dengue virus infection with seasonal variation at a tertiary care centre, Mangaluru: A retrospective study“. IP International Journal of Medical Microbiology and Tropical Diseases 8, Nr. 4 (15.12.2022): 294–98. http://dx.doi.org/10.18231/j.ijmmtd.2022.057.
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: Dengue is currently the second most prevalent vector borne disease in the world. Dengue fever and its more serious forms, dengue hemorrhagic fever and dengue shock syndrome are becoming important public health problems. Since there is no immunoprophylaxis or specific antiviral therapy available, clinico-microbiological diagnosis plays a vital role in patient management and implementation of control measures. : To identify dengue seropositive patients by NS1 antigen and anti-dengue IgM antibody detection by ELISA and correlate the changes in epidemiology. A retrospective study was performed for a period of 5 years from January 2017 to December 2021 in A.J Institute of Medical Sciences & Research Centre, Mangaluru. 1871 seropositive cases of all age groups admitted in the medical wards were included. Rapid immunochromatography test and ELISA was performed to detect NS1 antigen and IgM antibodies. Platelet count and total leucocyte count were also analysed. : Out of 14656, 1871 samples (12.75%) were positive for dengue. Higher rate of cases were in males and the age group of 21 to 30 years were chiefly affected. 1158(61.89%) were positive for NS1 antigen and total positive cases for IgM antibody was 530(28.32%). About 183 (9.79%) cases were positive for both NS1 antigen and IgM antibody. An increase in the prevalence of dengue was recorded during the month of June to October. The present study indicates a decline in dengue infection from 2020 which may be attributable to preventive and control measures provided by the health care workers besides having increased awareness among people. The decline may also may be due to the outbreak of SARS –Cov-2 in December 2019, which led to under diagnosis of dengue cases.
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Ranjan, Manish, Devanshi Khokhani, Sanjeeva Nayaka, Suchi Srivastava, Zachary P. Keyser und Ashish Ranjan. „Genomic diversity and organization of complex polysaccharide biosynthesis clusters in the genus Dickeya“. PLOS ONE 16, Nr. 2 (11.02.2021): e0245727. http://dx.doi.org/10.1371/journal.pone.0245727.
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The pectinolytic genus Dickeya (formerly Erwinia chrysanthemi) comprises numerous pathogenic species which cause diseases in various crops and ornamental plants across the globe. Their pathogenicity is governed by complex multi-factorial processes of adaptive virulence gene regulation. Extracellular polysaccharides and lipopolysaccharides present on bacterial envelope surface play a significant role in the virulence of phytopathogenic bacteria. However, very little is known about the genomic location, diversity, and organization of the polysaccharide and lipopolysaccharide biosynthetic gene clusters in Dickeya. In the present study, we report the diversity and structural organization of the group 4 capsule (G4C)/O-antigen capsule, putative O-antigen lipopolysaccharide, enterobacterial common antigen, and core lipopolysaccharide biosynthesis clusters from 54 Dickeya strains. The presence of these clusters suggests that Dickeya has both capsule and lipopolysaccharide carrying O-antigen to their external surface. These gene clusters are key regulatory components in the composition and structure of the outer surface of Dickeya. The O-antigen capsule/group 4 capsule (G4C) coding region shows a variation in gene content and organization. Based on nucleotide sequence homology in these Dickeya strains, two distinct groups, G4C group I and G4C group II, exist. However, comparatively less variation is observed in the putative O-antigen lipopolysaccharide cluster in Dickeya spp. except for in Dickeya zeae. Also, enterobacterial common antigen and core lipopolysaccharide biosynthesis clusters are present mostly as conserved genomic regions. The variation in the O-antigen capsule and putative O-antigen lipopolysaccharide coding region in relation to their phylogeny suggests a role of multiple horizontal gene transfer (HGT) events. These multiple HGT processes might have been manifested into the current heterogeneity of O-antigen capsules and O-antigen lipopolysaccharides in Dickeya strains during its evolution.
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Shi, S. R., R. J. Cote, L. Young, C. Yang, C. Chen, G. D. Grossfeld, D. A. Ginsberg, F. L. Hall und C. R. Taylor. „Development of optimal protocols for antigen retrieval immunohistochemistry based of the effects variation in temperature and pH: use of a ‘test battery’“. Proceedings, annual meeting, Electron Microscopy Society of America 53 (13.08.1995): 828–29. http://dx.doi.org/10.1017/s0424820100140518.
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Following development of the microwave antigen retrieval (AR) technique in 1991 for enhancement of immunohistochemical (IHC) staining, there have been numerous articles published worldwide describing the use of AR-IHC on archival tissue sections. The AR technique provides a method for retrieval of formalin-masked antigens which otherwise yield negative or very weakly positive IHC staining on archival paraffin sections. AR appears to be superior to other unmasking procedures, and serves to reduce detection thresholds of IHC for a wide range of antigen antibody systems. One important result is that AR increases the accuracy of diagnosis when using IHC for pathology. However, certain critical issues remain concerning the optimal protocol, the choice of AR solution, pH and the precise heating conditions. These aspects of AR-IHC require further study in order to improve standardization of AR-IHC, as emphasized by Taylor. Based on our recent study of AR-IHC under the influence of pH, we designed a new study specifically to examine the effect of different microwave heating conditions.
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Wolday, Dawit, Chun Yiu Jordan Fung, Gregory Morgan, Selina Casalino, Erika Frangione, Jennifer Taher und Jordan P. Lerner-Ellis. „HLA Variation and SARS-CoV Specific Antibody Response“. Viruses 15, Nr. 4 (31.03.2023): 906. http://dx.doi.org/10.3390/v15040906.
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Differences in SARS-CoV-2-specific immune responses have been observed between individuals following natural infection or vaccination. In addition to already known factors, such as age, sex, COVID-19 severity, comorbidity, vaccination status, hybrid immunity, and duration of infection, inter-individual variations in SARS-CoV-2 immune responses may, in part, be explained by structural differences brought about by genetic variation in the human leukocyte antigen (HLA) molecules responsible for the presentation of SARS-CoV-2 antigens to T effector cells. While dendritic cells present peptides with HLA class I molecules to CD8+ T cells to induce cytotoxic T lymphocyte responses (CTLs), they present peptides with HLA class II molecules to T follicular helper cells to induce B cell differentiation followed by memory B cell and plasma cell maturation. Plasma cells then produce SARS-CoV-2-specific antibodies. Here, we review published data linking HLA genetic variation or polymorphisms with differences in SARS-CoV-2-specific antibody responses. While there is evidence that heterogeneity in antibody response might be related to HLA variation, there are conflicting findings due in part to differences in study designs. We provide insight into why more research is needed in this area. Elucidating the genetic basis of variability in the SARS-CoV-2 immune response will help to optimize diagnostic tools and lead to the development of new vaccines and therapeutics against SARS-CoV-2 and other infectious diseases.
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Jabbar, Ammar A. R. „HLA and Disease Associations in Iraq“. Disease Markers 11, Nr. 4 (1993): 161–70. http://dx.doi.org/10.1155/1993/192019.
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The HLA system is deeply involved in susceptibility to a variety of diseases. Relationships between HLA and diseases are of considerable interest and importance, as they provide new tools for studying the inheritance, classification, and pathogenesis of these diseases. Studies on the distribution of HLA antigens in different populations have revealed the existence of racial variation and are therefore a prerequisite for studying HLA and disease associations in different racial groups. This study reviews six articles concerning HLA and disease in the Iraqi population. A comparison of these associations and an analysis of overall antigen frequencies among other Arab population and different ethnic groups are included. Some of our HLA-disease associations confirm other studies reported in these racial groups, while other diseases showed different HLA antigen associations from those recorded in other racial groups.
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Lyashchenko, Konstantin P., John M. Pollock, Roberto Colangeli und Maria Laura Gennaro. „Diversity of Antigen Recognition by Serum Antibodies in Experimental Bovine Tuberculosis“. Infection and Immunity 66, Nr. 11 (01.11.1998): 5344–49. http://dx.doi.org/10.1128/iai.66.11.5344-5349.1998.
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ABSTRACT Tuberculosis in cattle remains a major zoonotic and economic problem in many countries. The standard diagnostic assay for bovine tuberculosis, the intradermal tuberculin test, has low accuracy. Therefore, alternative immunodiagnostic methods, such as serological assays, are needed for detection of infected animals. Development of an accurate serodiagnostic test requires a detailed understanding of the humoral immune responses during bovine tuberculosis and, in particular, identification of the key antigens of Mycobacterium bovisinvolved in antibody production. In this study, we characterized antibody responses in cattle experimentally infected with M. bovis. Sequential serum samples were collected every 3 to 4 weeks for up to 27 months postinfection. Circulating immunoglobulin G antibody levels were measured by an enzyme-linked immunosorbent assay using 12 highly purified recombinant proteins of M. bovis. Six proteins, ESAT-6, 14-kDa protein, MPT63, MPT70, MPT51, and MPT32, were identified as major seroreactive antigens in bovine tuberculosis. A remarkable animal-to-animal variation of antigen recognition by serum antibodies was observed. Kinetic analyses of the antibody production to individual antigens during infection revealed that the heterogeneous antigen recognition profile changed markedly in a given infected animal as disease progressed.
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Morelli, Vania, Marieke de Visser, Nico van Tilburg, Hans Vos, Jeroen Eikenboom, Frits Rosendaal und Rogier Bertina. „ABO blood group genotypes, plasma von Willebrand factor levels and loading of von Willebrand factor with A and B antigens“. Thrombosis and Haemostasis 97, Nr. 04 (2007): 534–41. http://dx.doi.org/10.1160/th06-09-0549.
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SummaryABO blood group is a genetic determinant of von Willebrand factor (VWF) levels. We investigated the effect of ABO genotypes on VWF and factor VIII (FVIII) levels and on the degree to which VWF is loaded with A- and B-antigens, expressed as normalized ratios, nA-ratio and nB-ratio, respectively, in the Leiden Thrombophilia Study, a large case-control study on venous thrombosis. We found that the ABO locus had an allele-specific, dosage dependent effect on VWF and FVIII levels and on the loading of VWF with A-antigen and B-antigen. The highest mean nA- and nB-ratios were found in A 1 A 1 and BB genotypes, respectively. Four A1 O carriers had four 43-bp repeats in the minisatellite region of theABO gene in stead of the expected one repeat. All had a reduced nA-ratio compared to A 1 O carriers with one repeat in their A1 allele. The amount of A – and B- antigens expressed onVWF (nA-ratio and nB-ratio) explained about 18% (R2) of the variation in VWF levels.
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Rogan, M. T., I. Marshall, G. D. F. Reid, C. N. L. Macpherson und P. S. Craig. „The potential of vervet monkeys (Cercopithecus aethiops) and baboons (Papio anubis) as models for the study of the immunology of Echinococcus granulosus infections“. Parasitology 106, Nr. 5 (Juni 1993): 511–17. http://dx.doi.org/10.1017/s0031182000076812.
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SUMMARYNine vervet monkeys and nine baboons were infected with eggs of Echinococcus granulosus per as. Six of the vervets and one of the baboons possessed hydatid cysts at autopsy, 15–28 months post-infection. The sequential IgG response to hydatid fluid and protoscolex antigens showed considerable inter-animal variation. Infected vervets and baboons became seropositive after an average of 8 months post-infection. Considerable fluctuation in the IgG response was observed, particularly to the hydatid fluid antigen which, in humans, may contribute to the existence of a significant proportion of seronegative individuals. Vervets, in particular, may be useful to study immunological events associated with exposure, development and resolution of hydatid disease in outbred human populations.
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Ampomah, Paulina, Liz Stevenson, Michael F. Ofori, Lea Barfod und Lars Hviid. „B-Cell Responses to Pregnancy-Restricted and -Unrestricted Plasmodium falciparum Erythrocyte Membrane Protein 1 Antigens in Ghanaian Women Naturally Exposed to Malaria Parasites“. Infection and Immunity 82, Nr. 5 (24.02.2014): 1860–71. http://dx.doi.org/10.1128/iai.01514-13.
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ABSTRACTProtective immunity toPlasmodium falciparummalaria acquired after natural exposure is largely antibody mediated. IgG-specificP. falciparumEMP1 (PfEMP1) proteins on the infected erythrocyte surface are particularly important. The transient antibody responses and the slowly acquired protective immunity probably reflect the clonal antigenic variation and allelic polymorphism of PfEMP1. However, it is likely that other immune-evasive mechanisms are also involved, such as interference with formation and maintenance of immunological memory. We measured PfEMP1-specific antibody levels by enzyme-linked immunosorbent assay (ELISA) and memory B-cell frequencies by enzyme-linked immunosorbent spot (ELISPOT) assay in a cohort ofP. falciparum-exposed nonpregnant Ghanaian women. The antigens used were a VAR2CSA-type PfEMP1 (IT4VAR04) with expression restricted to parasites infecting the placenta, as well as two commonly recognized PfEMP1 proteins (HB3VAR06 and IT4VAR60) implicated in rosetting and not pregnancy restricted. This enabled, for the first time, a direct comparison in the same individuals of immune responses specific for a clinically important parasite antigen expressed only during well-defined periods (pregnancy) to responses specific for comparable antigens expressed independent of pregnancy. Our data indicate that PfEMP1-specific B-cell memory is adequately acquired even when antigen exposure is infrequent (e.g., VAR2CSA-type PfEMP1). Furthermore, immunological memory specific for VAR2CSA-type PfEMP1 can be maintained for many years without antigen reexposure and after circulating antigen-specific IgG has disappeared. The study provides evidence that natural exposure toP. falciparumleads to formation of durable B-cell immunity to clinically important PfEMP1 antigens. This has encouraging implications for current efforts to develop PfEMP1-based vaccines.
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Margolis, D. J., N. Mitra, J. L. Duke, R. Berna, J. D. Margolis, O. Hoffstad, B. S. Kim et al. „Human leukocyte antigen class-I variation is associated with atopic dermatitis: A case-control study“. Human Immunology 82, Nr. 8 (August 2021): 593–99. http://dx.doi.org/10.1016/j.humimm.2021.04.001.
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Hjertholm, Peter, Morten Fenger-Grøn, Mogens Vestergaard, Morten B. Christensen, Michael Borre, Henrik Møller und Peter Vedsted. „Variation in general practice prostate-specific antigen testing and prostate cancer outcomes: An ecological study“. International Journal of Cancer 136, Nr. 2 (12.06.2014): 435–42. http://dx.doi.org/10.1002/ijc.29008.
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Carobene, Anna, Elena Guerra, Massimo Locatelli, Vito Cucchiara, Alberto Briganti, Aasne K. Aarsand, Abdurrahman Coşkun et al. „Biological variation estimates for prostate specific antigen from the European Biological Variation Study; consequences for diagnosis and monitoring of prostate cancer“. Clinica Chimica Acta 486 (November 2018): 185–91. http://dx.doi.org/10.1016/j.cca.2018.07.043.
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Takahashi, Satoshi, Jun Ooi, Nobuhiro Tsukada, Seiko Kato, Aki Sato, Toshiro Kawakita, Yasuyuki Nagata et al. „The Impact of HLA Haplotype Matching for Mismatched Cord Blood Transplantation.“ Blood 114, Nr. 22 (20.11.2009): 1206. http://dx.doi.org/10.1182/blood.v114.22.1206.1206.
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Abstract Abstract 1206 Poster Board I-228 [Study purpose] With the increased number of cord blood transplantation (CBT) for adults, more human leukocyte antigen (HLA)-mismatched grafts are selected as alternative donor. In Japan, we have performed more than 5,500 CBT and almost two-third of them has been using 2-loci HLA-mismatched grafts. Slow engraftment and high risk of graft failure should be solved to improve the clinical results. In general, the degree of HLA disparity is known to be associated with risks of poor graft function and of graft-versus-host disease (GVHD). On the other hand, those risks of transplant-related complications are not equivalent even in the donor-recipient pairs who have same number of mismatched HLA antigens. There might be “haplotype matching effect” because novel undetected major histocompatiblility antigen (MHC) resident variation encoded on HLA haplotypes and those mismatching could be responsible for post-transplant risks. In this study, we have analyzed the impact of HLA haplotype matching in HLA-mismatched CBT using the same method in the single institute. [Patients and Methods] We studied the clinical outcomes of 149 consecutive adult patients who received unrelated CBT between August 1998 and June 2009 in the institute of medical Science, University of Tokyo. Patients received previous allogeneic tranplants were excluded from this study. All patients received myeloablative regimens including 12 Gy of total body irradiation, cyclosporine plus short term methotraxate for GVHD prophylaxis and almost the same supportive care. By low-resolution typing method for HLA-A, -B and –DR loci, 9 patients received matched grafts, 46 received 1 antigen-mismatched and 94 received 2 antigens-mismatched grafts in the host-versus-graft (HvG) direction. In the graft-versus-host (GvH) direction, 7 patients received matched grafts, 51 received 1 antigen-mismatched and 91 received 2 antigens-mismatched grafts. When we looked at the maximum number of mismatched antigens for both directions, 38 patients received 1 antigen-mismatched and 111 received 2 antigens-mismatched grafts respectively, but there was no matched pair. Common haplotypes in Japanese population were referred from the 11th International Histocompatibility Workshop and other previous reports. Median numbers of leukocytes and CD34+ progenitor cells before freezing of cord blood grafts were 2.4×107/kg and 0.9×105/kg, respectively. Median follow-up was 39 months. We evaluated the impact of haplotype matching on cumulative incidences of hematopoietic recovery, of GVHD, of relapse and of non-relapse mortality (NRM) using the Pepe and Mori's test. Estimates of overall survival were calculated using the Kaplan-Meier method and analyzed by the log-rank test. [Results] Thirty (11 of 38 one antigen-mismatched and 19 of 111 two antigens-mismatched) among all 149 pairs were defined as the haplotype-matched pairs sharing same haplotypes in both grafts and recipients. The age, sex, cytomegalovirus serological status, diagnosis, risk of the disease at the transplant, numbers of total nucleated cells and CD34+ cells at the cryopreserved were not significantly different between both groups with and without matched haplotypes. Among the 1 antigen-mismatched pairs in the HvG direction, early engraftment of neutrophil after CBT occurred in haplotype-matched group compared with control group (median: 20.5 days versus 23 days, P=0.01). The haplotype matched group had better platelet engraftment in the 1 antigen-mismatched pairs (cumulative incidence on day 120: 86% versus 62%; median: 41 days versus 53 days), but this is not significant (P=0.29). Such correlation between engraftment and haplotype matching was not observed in 2 antigens-mismatched pairs. The cumulative incidences of grades II to IV acute GVHD were not significantly different between haplotype matched and control groups, however, those of grades III and IV in patients with matched-haplotype tended to be lower among 1 antigen-mismatched pairs in GvH direction (P=0.10) and were significantly lower among 2 antigens-mismatched pairs (P=0.02). Those haplotype matching effects were not observed in survival rates, cumulative incidences of relapse and NRM among any HLA mismatched pairs. [Conclusion] Those data suggest that untyped variation carried on the HLA haplotytpe might be better to be matched and the haplotype matching might effect on better engraftment and lower risk of sever acute GVHD after HLA-mismatched CBT. Disclosures: No relevant conflicts of interest to declare.
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Huang, X., S. Chen und E. I. Tietz. „Immunocytochemical detection of regional protein changes in rat brain sections using computer-assisted image analysis.“ Journal of Histochemistry & Cytochemistry 44, Nr. 9 (September 1996): 981–87. http://dx.doi.org/10.1177/44.9.8773563.
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We used several approaches to assess the reliability and sensitivity of computer-assisted densitometry to detect regional changes in tissue antigen content as a function of immunohistochemical staining density. We designed a model system to mimic variations in antigen concentration in postfixed, slide-mounted rat brain sections by varying the ratios of conjugated (biotinylated) to unconjugated secondary antibody. Antigen concentration was also varied in tissue discs made from mixing rat brain homogenate with increasing amounts of tissue embedding compound. The monoclonal antibody bd-17 to the beta2/3 subunit of the GABAA receptor was used as the primary antibody. Immunostaining density was visualized with diaminobenzidine (DAB). There was a significant, positive linear relationship (r = 0.97-0.99) between immunostaining intensity and antigen concentration. With this approach, changes in antigen content of less than 10%, as reflected in immunostaining intensity, were detectable in brain sections. The low degree of variability in measures of regional variation in immunostaining in sections from naive rats (n = 7) suggested that the method was suitable for quantitative analysis and indicated the reliability of the method. This systematic study of the utility of computer-assisted image analysis for semiquantitative immunohistochemical analysis found the method to be both reliable and sensitive.
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Spencer, David H., Arnold Kas, Eric E. Smith, Christopher K. Raymond, Elizabeth H. Sims, Michele Hastings, Jane L. Burns, Rajinder Kaul und Maynard V. Olson. „Whole-Genome Sequence Variation among Multiple Isolates of Pseudomonas aeruginosa“. Journal of Bacteriology 185, Nr. 4 (15.02.2003): 1316–25. http://dx.doi.org/10.1128/jb.185.4.1316-1325.2003.
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ABSTRACT Whole-genome shotgun sequencing was used to study the sequence variation of three Pseudomonas aeruginosa isolates, two from clonal infections of cystic fibrosis patients and one from an aquatic environment, relative to the genomic sequence of reference strain PAO1. The majority of the PAO1 genome is represented in these strains; however, at least three prominent islands of PAO1-specific sequence are apparent. Conversely, ∼10% of the sequencing reads derived from each isolate fail to align with the PAO1 backbone. While average sequence variation among all strains is roughly 0.5%, regions of pronounced differences were evident in whole-genome scans of nucleotide diversity. We analyzed two such divergent loci, the pyoverdine and O-antigen biosynthesis regions, by complete resequencing. A thorough analysis of isolates collected over time from one of the cystic fibrosis patients revealed independent mutations resulting in the loss of O-antigen synthesis alternating with a mucoid phenotype. Overall, we conclude that most of the PAO1 genome represents a core P. aeruginosa backbone sequence while the strains addressed in this study possess additional genetic material that accounts for at least 10% of their genomes. Approximately half of these additional sequences are novel.
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Mendelsohn, A. C., L. M. Sanmarco, R. G. Spallanzani, D. Brown, F. J. Quintana, S. Breton und M. A. Battistone. „From initial segment to cauda: a regional characterization of mouse epididymal CD11c+ mononuclear phagocytes based on immune phenotype and function“. American Journal of Physiology-Cell Physiology 319, Nr. 6 (01.12.2020): C997—C1010. http://dx.doi.org/10.1152/ajpcell.00392.2020.
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Successful sperm maturation and storage rely on a unique immunological balance that protects the male reproductive organs from invading pathogens and spermatozoa from a destructive autoimmune response. We previously characterized one subset of mononuclear phagocytes (MPs) in the murine epididymis, CX3CR1+ cells, emphasizing their different functional properties. This population partially overlaps with another subset of understudied heterogeneous MPs, the CD11c+ cells. In the present study, we analyzed the CD11c+ MPs for their immune phenotype, morphology, and antigen capturing and presenting abilities. Epididymides from CD11c-EYFP mice, which express enhanced yellow fluorescent protein (EYFP) in CD11c+ MPs, were divided into initial segment (IS), caput/corpus, and cauda regions. Flow cytometry analysis showed that CD11c+ MPs with a macrophage phenotype (CD64+ and F4/80+) were the most abundant in the IS, whereas those with a dendritic cell signature [CD64− major histocompatibility complex class II (MHCII)+] were more frequent in the cauda. Immunofluorescence revealed morphological and phenotypic differences between CD11c+ MPs in the regions examined. To assess the ability of CD11c+ cells to take up antigens, CD11c-EYFP mice were injected intravenously with ovalbumin. In the IS, MPs expressing macrophage markers were most active in taking up the antigens. A functional antigen-presenting coculture study was performed, whereby CD4+ T cells were activated after ovalbumin presentation by CD11c+ epididymal MPs. The results demonstrated that CD11c+ MPs in all regions were capable of capturing and presenting antigens. Together, this study defines a marked regional variation in epididymal antigen-presenting cells that could help us understand fertility and contraception but also has larger implications in inflammation and disease pathology.
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Wendland, Lori D., Paul A. Klein, Elliott R. Jacobson und Mary B. Brown. „Mycoplasma agassizii Strain Variation and Distinct Host Antibody Responses Explain Differences between Enzyme-Linked Immunosorbent Assays and Western Blot Assays“. Clinical and Vaccine Immunology 17, Nr. 11 (01.09.2010): 1739–45. http://dx.doi.org/10.1128/cvi.00215-10.
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ABSTRACT The precarious status of desert (Gopherus agassizii) and gopher (G. polyphemus) tortoises has resulted in conservation efforts that now include health assessment as an important component of management decision-making. Mycoplasmal upper respiratory tract disease (URTD) is one of very few diseases in chelonians for which comprehensive and rigorously validated diagnostic tests exist. In this study, serum samples obtained from eight Gopherus tortoises documented at necropsy to (i) be enzyme-linked immunosorbent assay (ELISA) seropositive using the PS6 antigen, (ii) be infected with Mycoplasma agassizii as indicated by direct isolation of the pathogen from the respiratory surfaces, and (iii) have histological lesions of mycoplasmal URTD were used to evaluate four distinct clinical isolates of M. agassizii as antigens for ELISA and Western blot analyses. Each animal sample reacted in the Western blot with its homologous M. agassizii strain, but recognition of heterologous M. agassizii strains was variable. Further, individual animals varied significantly with respect to the specific proteins recognized by the humoral immune response. An additional 114 Gopherus serum samples were evaluated using ELISA antigens prepared from the four distinct M. agassizii strains; A 405 values were significantly correlated (r 2 goodness of fit range, 0.708 to 0.771; P < 0.0001) for all antigens tested. The results confirm that strain variation is responsible for the observed differences between Western blot binding patterns. Thus, reliance on a single M. agassizii strain as an antigen in Western blot assays may provide false-negative results. This could have adverse consequences for the well-being of these environmentally sensitive hosts if false-negative animals were relocated to sites consisting of true-negative populations.
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Petersen, F. C., S. Assev, H. C. van der Mei, H. J. Busscher und A. A. Scheie. „Functional Variation of the Antigen I/II Surface Protein in Streptococcus mutans and Streptococcus intermedius“. Infection and Immunity 70, Nr. 1 (Januar 2002): 249–56. http://dx.doi.org/10.1128/iai.70.1.249-256.2002.
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ABSTRACT Although Streptococcus intermedius and Streptococcus mutans are regarded as members of the commensal microflora of the body, S. intermedius is often associated with deep-seated purulent infections, whereas S. mutans is frequently associated with dental caries. In this study, we investigated the roles of the S. mutans and S. intermedius antigen I/II proteins in adhesion and modulation of cell surface characteristics. By using isogenic mutants, we show that the antigen I/II in S. mutans, but not in S. intermedius, was involved in adhesion to a salivary film under flowing conditions, as well as in binding to rat collagen type I. Binding to human fibronectin was a common function associated with the S. mutans and S. intermedius antigen I/II. Adhesion of S. mutans or S. intermedius to human collagen types I or IV was negligible. Hydrophobicity, as measured by water contact angles, and zeta potentials were unaltered in the S. intermedius mutant. The S. mutans isogenic mutants, on the other hand, exhibited more positive zeta potentials at physiological pH values than did the wild type. The results indicate common and species-specific roles for the antigen I/II in mediating the attachment of S. mutans and S. intermedius to host components and in determining cell surface properties.
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van der Bom, Johanna G., Michiel L. Bots, Frits Haverkate, Cornelis Kluft und Diederick E. Grobbee. „The 4G5G polymorphism in the gene for PAI-1 and the circadian oscillation of plasma PAI-1“. Blood 101, Nr. 5 (01.03.2003): 1841–44. http://dx.doi.org/10.1182/blood-2002-07-2181.
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Plasminogen activator inhibitor type I (PAI-1) antigen concentrations follow a circadian oscillation peaking in the morning. Some individuals show no apparent circadian rhythm, while others show up to a 10-fold variation in PAI-1 over 24 hours. Results from experimental studies suggest that a polymorphism in the promoter of the gene for PAI-1 (4G5G) directly influences the circadian expression of the PAI-1 gene. We studied whether the diurnal variation of PAI-1 antigen differs for the genotypes of the4G5G polymorphism. A population-based, cross-sectional study was performed among 263 subjects selected from the Rotterdam Study, a population-based cohort of 7983 men and women aged 55 years and older. The 4G allele was associated with a more pronounced circadian expression of PAI-1 antigen. Morning PAI-1 antigen concentrations were 79 ng/mL (95% confidence interval [CI], 68-92) in subjects homozygous for 4G, 62 ng/mL (95% CI, 54-72) in heterozygous subjects, and 59 ng/mL (95% CI, 49-71) in subjects homozygous for 5G. While respective PAI-1 antigen concentrations in the afternoon were 40 ng/mL (95% CI, 33-49), 41 ng/mL (95% CI, 37-47), and 40 ng/mL (95% CI, 49-71). These findings suggest that the morning increase in PAI-1 antigen concentration is more pronounced among subjects homozygous for the4G allele compared with the morning increase among the other genotypes. Additionally, these findings show that homozygosity for the 4G allele is associated with increased PAI-1 levels during the morning only.
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Hubbard, Anthony R., und Man Yu Wong. „A Collaborative Study on the Measurement of Protein S Antigen in Plasma“. Thrombosis and Haemostasis 68, Nr. 02 (1992): 115–18. http://dx.doi.org/10.1055/s-0038-1656334.
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SummaryA collaborative study on the measurement of protein S (PS) antigen (total and free) in a freeze-dried ampouled test plasma by assay against local house standard plasmas was carried out in eleven laboratories. Potency estimates of total PS showed good agreement between laboratories with a geometric coefficient of variation (gcv) of 5.9% and an overall combined potency of 0.84 units per ml. Potency estimates of free PS antigen in the test sample were associated with increased variability between laboratories resulting from the polyethylene glycol (PEG) precipitation step which is used to separate free PS from PS bound to the C4b binding protein. Free PS in the test could be expressed relative to either total PS in the house standards (e. g. 0.28 units per ml) or relative to free PS in the house standards following PEG precipitation (e. g. 0.71 units free PS per ml).
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Morgan, Rhian M., Robert J. C. Steele, Ghulam Nabi und Colin McCowan. „Socioeconomic Variation and Prostate Specific Antigen Testing in the Community: A United Kingdom Based Population Study“. Journal of Urology 190, Nr. 4 (Oktober 2013): 1207–12. http://dx.doi.org/10.1016/j.juro.2013.04.044.
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Tonouchi, Keisuke, Yu Adachi, Saya Moriyama, Kaori Sano, Koshiro Tabata, Keigo Ide, Haruko Takeyama, Tadaki Suzuki und Yoshimasa Takahashi. „Stereotyped B-cell response that counteracts antigenic variation of influenza viruses“. International Immunology 32, Nr. 9 (06.06.2020): 613–21. http://dx.doi.org/10.1093/intimm/dxaa038.
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Abstract Influenza A subtypes are categorized into group 1 and group 2 based on the hemagglutinin (HA) sequence. Owing to the phylogenetic distance of HAs in different groups, antibodies that bind multiple HA subtypes across different groups are extremely rare. In this study, we demonstrated that an immunization with acid-treated HA antigen elicits germinal center (GC) B cells that bind multiple HA subtypes in both group 1 and group 2. The cross-group GC B cells utilized mostly one VH gene (1S56) and exhibited a sign of clonal evolution within GCs. The 1S56-lineage IgGs derived from GC B cells were able to bind to HA protein on the infected cell surface but not to the native form of HA protein, suggesting the cryptic nature of the 1S56 epitope and its exposure in infected cells. Finally, the 1S56-lineage IgGs provided protection against lethal infection in an Fc-dependent manner, independent of the virus-neutralizing activity. Thus, we identified 1S56-lineage antibodies as a unique stereotype for achieving cross-group influenza specificity. The antigens exposing the 1S56 epitope may be good candidates for broadly protective immunogens.
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Nijen Twilhaar, Maarten K., Lucas Czentner, Joanna Grabowska, Alsya J. Affandi, Chun Yin Jerry Lau, Katarzyna Olesek, Hakan Kalay et al. „Optimization of Liposomes for Antigen Targeting to Splenic CD169+ Macrophages“. Pharmaceutics 12, Nr. 12 (25.11.2020): 1138. http://dx.doi.org/10.3390/pharmaceutics12121138.
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Despite promising progress in cancer vaccination, therapeutic effectiveness is often insufficient. Cancer vaccine effectiveness could be enhanced by targeting vaccine antigens to antigen-presenting cells, thereby increasing T-cell activation. CD169-expressing splenic macrophages efficiently capture particulate antigens from the blood and transfer these antigens to dendritic cells for the activation of CD8+ T cells. In this study, we incorporated a physiological ligand for CD169, the ganglioside GM3, into liposomes to enhance liposome uptake by CD169+ macrophages. We assessed how variation in the amount of GM3, surface-attached PEG and liposomal size affected the binding to, and uptake by, CD169+ macrophages in vitro and in vivo. As a proof of concept, we prepared GM3-targeted liposomes containing a long synthetic ovalbumin peptide and tested the capacity of these liposomes to induce CD8+ and CD4+ T-cell responses compared to control liposomes or soluble peptide. The data indicate that the delivery of liposomes to splenic CD169+ macrophages can be optimized by the selection of liposomal constituents and liposomal size. Moreover, optimized GM3-mediated liposomal targeting to CD169+ macrophages induces potent immune responses and therefore presents as an interesting delivery strategy for cancer vaccination.
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Bento, Dulce, Sandra Jesus, Filipa Lebre, Teresa Gonçalves und Olga Borges. „Chitosan Plus Compound 48/80: Formulation and Preliminary Evaluation as a Hepatitis B Vaccine Adjuvant“. Pharmaceutics 11, Nr. 2 (09.02.2019): 72. http://dx.doi.org/10.3390/pharmaceutics11020072.
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Current vaccine research is mostly based on subunit antigens. Despite the better toxicity profile of these antigens they are often poorly immunogenic, so adjuvant association has been explored as a strategy to obtain a potent vaccine formulation. Recently, mast cell activators were recognized as a new class of vaccine adjuvants capable of potentiating mucosal and systemic immune responses. In this study, a co-adjuvanted delivery system was developed and characterized, combining the mast cell activator C48/80 with chitosan nanoparticles (Chi-C48/80 NPs), and the results were compared with plain chitosan nanoparticles. The adsorption of model antigens onto the NP surface as well as the biocompatibility of the system was not affected by the incorporation of C48/80 in the formulation. The stability of the nanoparticles was demonstrated by studying the variation of size and zeta potential at different times, and the ability to be internalized by antigen presenting cells was confirmed by confocal microscopy. Vaccination studies with hepatitis B surface antigen loaded Chi-C48/80 NPs validated the adjuvanticity of the delivery system, demonstrating for the first time a successful association between a mast cell activator and chitosan nanoparticles as a vaccine adjuvant for hepatitis B virus, applied to a nasal vaccination strategy.
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van der Bom, Johanna, Michiel Bots, Frits Haverkate, Piet Meyer, Albert Hofman, Cornelis Kluft und Diederick Grobbee. „Fibrinolytic Activity in Peripheral Atherosclerosis in the Elderly“. Thrombosis and Haemostasis 81, Nr. 02 (1999): 275–80. http://dx.doi.org/10.1055/s-0037-1614457.
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SummaryIncreased concentrations of plasminogen activator inhibitor type 1 (PAI-1) and of D-dimer have jointly been found in subjects with cardiovascular disease. To understand this apparent paradox of increased inhibition of fibrinolysis (high PAI-1) combined with increased fibrinolytic activity (high D-dimer), we examined the relation between D-dimer, PAI-1 and the activator of fibrinolysis, tissue type plasminogen activator (t-PA) in subjects with varying severity of peripheral atherosclerosis. In 325 subjects selected from the Rotterdam Study, a cohort of 7983 men and women aged 55 years and over, the ankle to brachial systolic blood pressure ratio, t-PA antigen and activity, PAI-1 antigen and D-dimer were measured.T-PA antigen and t-PA activity were, independent from each other, increased with degree of atherosclerosis; t-PA antigen increased with 3.5 ng/ml (SE 1.7, p = 0.04) and t-PA activity with 0.46 IU/ml (0.20, p = 0.02) per unit decrease in ankle to brachial pressure ratio (i.e. increase in atherosclerosis). PAI-1 antigen was not related to atherosclerosis. More marked atherosclerosis was associated with increased D-dimer, mainly in subgroups with PAI-1 antigen below 50 ng/ml, t-PA antigen below 10 ng/ml, or t-PA activity above 1.5 IU/ml. In contrast to current beliefs, we found that only a fraction of the variation of t-PA antigen was due to the variation in circulating PAI-1 antigen. A slight positive association was observed between t-PA antigen and D-dimer. PAI-1 and t-PA activity were not associated with D-dimer concentration.In conclusion, in subjects with peripheral atherosclerosis PAI-1 antigen is not increased, but low PAI-1 levels (and possibly also low levels of t-PA antigen and high levels of t-PA activity) appear to be required to increase circulating D-dimer. This suggests that increased D-dimer levels in subjects with atherosclerosis do not reflect increased inhibition, but rather reflect increased fibrinolysis.
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Santander, Javier, Soo-Young Wanda, Cheryl A. Nickerson und Roy Curtiss. „Role of RpoS in Fine-Tuning the Synthesis of Vi Capsular Polysaccharide in Salmonella enterica Serotype Typhi“. Infection and Immunity 75, Nr. 3 (18.12.2006): 1382–92. http://dx.doi.org/10.1128/iai.00888-06.
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ABSTRACT Regulation of the synthesis of Vi polysaccharide, a major virulence determinant in Salmonella enterica serotype Typhi, is under the control of two regulatory systems, ompR-envZ and rscB-rscC, which respond to changes in osmolarity. Some serotype Typhi strains exhibit overexpression of Vi polysaccharide, which masks clinical detection of lipopolysaccharide O antigen. This variation in Vi polysaccharide and O antigen display (VW variation) has been observed since the initial studies of serotype Typhi. In this study, we report that rpoS plays a role in this increased expression in Vi polysaccharide. We constructed a variety of isogenic serotype Typhi mutants that differed in their expression levels of RpoS and examined the role of the rpoS product in synthesis of Vi polysaccharide under different osmolarity conditions. Vi polysaccharide synthesis was also examined in serotype Typhi mutants in which the native promoter of the rpoS was replaced by an araCPBAD cassette, so that the expression of rpoS was arabinose dependent. The RpoS− strains showed increased syntheses of Vi polysaccharide, which at low and medium osmolarities masked O antigen detection. In contrast, RpoS+ strains showed lower syntheses of Vi polysaccharide, and an increased detection of O antigen was observed. During exponential growth, when rpoS is unstable or present at low levels, serotype Typhi RpoS+ strains overexpress the Vi polysaccharide at levels comparable to those for RpoS− strains. Our results show that RpoS is another regulator of Vi polysaccharide synthesis and contributes to VW variation in serotype Typhi, which has implications for the development of recombinant attenuated Salmonella vaccines in humans.
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Sun, Yamin, Min Wang, Quan Wang, Boyang Cao, Xin He, Kun Li, Lu Feng und Lei Wang. „Genetic Analysis of the Cronobacter sakazakii O4 to O7 O-Antigen Gene Clusters and Development of a PCR Assay for Identification of All C. sakazakii O Serotypes“. Applied and Environmental Microbiology 78, Nr. 11 (23.03.2012): 3966–74. http://dx.doi.org/10.1128/aem.07825-11.
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ABSTRACTThe Gram-negative bacteriumCronobacter sakazakiiis an emerging food-borne pathogen that causes severe invasive infections in neonates. Variation in the O-antigen lipopolysaccharide in the outer membrane provides the basis for Gram-negative bacteria serotyping. The O-antigen serotyping scheme forC. sakazakii, which includes seven serotypes (O1 to O7), has been recently established, and the O-antigen gene clusters and specific primers for threeC. sakazakiiserotypes (O1, O2, and O3) have been characterized. In this study, theC. sakazakiiO4, O5, O6, and O7 O-antigen gene clusters were sequenced, and gene functions were predicted on the basis of homology.C. sakazakiiO4 shared a similar O-antigen gene cluster withEscherichia coliO103. The general features and anomalies of all sevenC. sakazakiiO-antigen gene clusters were evaluated and the relationship between O-antigen structures and their gene clusters were investigated. Serotype-specific genes for O4 to O7 were identified, and a molecular serotyping method for allC. sakazakiiO serotypes, a multiplex PCR assay, was developed by screening against 136 strains ofC. sakazakiiand closely related species. The sensitivity of PCR-based serotyping method was determined to be 0.01 ng of genomic DNA and 103CFU of each strain/ml. This study completes the elucidation ofC. sakazakiiO-antigen genetics and provides a molecular method suitable for the identification ofC. sakazakiiO1 to O7 strains.
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Wu, Yuxiang, Qi Fan, Yinuo Chen, Xia Sun und Guoqing Shi. „Production and Selection of Antibody–Antigen Pairs for the Development of Immunoenzyme Assay and Lateral Flow Immunoassay Methods for Carbofuran and Its Analogues“. Biosensors 12, Nr. 8 (24.07.2022): 560. http://dx.doi.org/10.3390/bios12080560.
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To produce a sensitive monoclonal antibody (mAb) for the simultaneous detection of carbofuran, benfuracarb, carbosulfan and 3-hydroxy-carbofuran, 2,3-dihydro-2,2-dimethyl-7-benzofuranmethanamine (DDB) was conjugated to bovine serum albumin (BSA) to prepare the immunogen DDB-BSA and mice were immunized. Coating antigens were prepared by conjugating DDB and 5-methoxy-2,3-dihydrobenzofuran-3-acetic acid (MDA) to BSA and ovalbumin (OVA), respectively. Furthermore, the effect of different antibody–antigen pairs on the sensitivity of ELISA and LFIA methods for the detection of carbofuran was investigated. After the immunization, a high-affinity mAb 13C8 was obtained. The ability of the coating antigen to compete with carbofuran for binding antibodies was found to be significantly different between ELISA and LFIA methods. With the antibody–antigen pair 13C8-MDA-OVA, the IC50 values of the ELISA and QD-LFIA methods for carbofuran were 0.18 ng/mL and 0.67 ng/mL, respectively. The cross-reactivity (CR) values of the two methods for benfuracarb, carbosulfan and 3-hydroxy-carbofuran ranged from 72.0% to 83.7%, while, for other carbamate pesticides, the CR values were less than 1%. The spiked recoveries of carbofuran in vegetables by the QD-LFIA method were 83–111%, with a coefficient of variation below 10%, and the test results of the actual samples were consistent with the HPLC-MS method. Overall, this study provides key materials for the development of immunoassays for carbofuran and its analogues, and the antibody–antigen pair selection strategy established in this study provides useful insights for the development of sensitive immunoassays for other compounds.
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BEHAL, R., R. JAIN, K. K. BEHAL und T. N. DHOLE. „Variation in the host ABO blood group may be associated with susceptibility to hepatitis C virus infection“. Epidemiology and Infection 138, Nr. 8 (14.12.2009): 1096–99. http://dx.doi.org/10.1017/s0950268809991117.
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SUMMARYThis study aimed to determine the relationship between hepatitis C virus (HCV) infection and ABO/Rhesus blood groups, age and sex. A total of 20 000 patients who came to donate blood in the blood bank of GSVM Medical College, Kanpur were enrolled in the study. Demographic data recorded for each patient included age, sex and blood group. Blood samples were tested for anti-HCV antibodies and ABO/Rhesus blood group antigen typing was performed. The overall positive rate of anti-HCV was 0·34%. We found that seropositivity for HCV increased with age. Anti-HCV antibodies were detected in 1/765 women (0·13%), compared to 67/19 235 men (0·35%). Seroprevalence of HCV was found to be higher in blood group O individuals (0·42%) and lowest in blood group AB individuals (0·04%). The results of this study demonstrate that that HCV infection may not be related to age and sex but the possible association of blood group antigens with HCV infection cannot be ruled out.
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Zhou, Lei, Jinxing Song, Mengxiang Wang, Zhuoya Sun, Junru Sun, Panpan Tian, Guoqing Zhuang, Angke Zhang, Yanan Wu und Gaiping Zhang. „Establishment of a Dual-Antigen Indirect ELISA Based on p30 and pB602L to Detect Antibodies against African Swine Fever Virus“. Viruses 15, Nr. 9 (30.08.2023): 1845. http://dx.doi.org/10.3390/v15091845.
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African swine fever (ASF) is an acute, virulent, and highly fatal infectious disease caused by the African swine fever virus (ASFV). There is no effective vaccine or diagnostic method to prevent and control this disease currently, which highlights the significance of ASF early detection. In this study, we chose an early antigen and a late-expressed antigen to co-detect the target antibody, which not only helps in early detection but also improves accuracy and sensitivity. CP204L and B602L were successfully expressed as soluble proteins in an Escherichia coli vector system. By optimizing various conditions, a dual-antigen indirect ELISA for ASFV antibodies was established. The assay was non-cross-reactive with antibodies against the porcine reproductive and respiratory syndrome virus, classical swine fever virus, porcine circovirus type 2, and pseudorabies virus. The maximum serum dilution for detection of ASFV-positive sera was 1:1600. The intra-batch reproducibility coefficient of variation was <5% and the inter-batch reproducibility coefficient of variation was <10%. Compared with commercial kits, the dual-antigen indirect ELISA had good detection performance. In conclusion, we established a detection method with low cost, streamlined production process, and fewer instruments. It provides a new method for the serological diagnosis of ASF.
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Klinkenberg, Lieke JJ, Eef GWM Lentjes und Arjen-Kars Boer. „Clinical interpretation of prostate-specific antigen values: Type of applied cut-off value exceeds methods bias as the major source of variation“. Annals of Clinical Biochemistry: International Journal of Laboratory Medicine 56, Nr. 2 (24.02.2019): 259–65. http://dx.doi.org/10.1177/0004563218822665.
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Background Prostate-specific antigen is the biochemical gold standard for the (early) detection and monitoring of prostate cancer. Interpretation of prostate-specific antigen is both dependent on the method and cut-off. The aim of this study was to examine the effect of method-specific differences and cut-off values in a national external quality assessment scheme (EQAS). Methods The Dutch EQAS for prostate-specific antigen comprised an annual distribution of 12 control materials. The results of two distributions were combined with the corresponding cut-off value. Differences between methods were quantified by simple linear regression based on the all laboratory trimmed mean. To assess the clinical consequence of method-specific differences and cut-off values, a clinical data-set of 1040 patients with an initial prostate-specific antigen measurement and concomitant conclusive prostate biopsy was retrospectively collected. Sensitivity and specificity for prostate cancer were calculated for all EQAS participants individually. Results In the Netherlands, seven different prostate-specific antigen methods are used. Interestingly, 67% of these laboratories apply age-specific cut-off values. Methods showed a maximal relative difference of 26%, which were not reflected in the cut-off values. The largest differences were caused by the type of cut-off, for example in the Roche group the cut-off value differed maximal 217%. Clinically, a fixed prostate-specific antigen cut-off has a higher sensitivity than an age-specific cut-off (mean 89% range 86–93% versus 79% range 63–95%, respectively). Conclusions This study shows that the differences in cut-off values exceed the method-specific differences. These results emphasize the need for (inter)national harmonization/standardization programmes including cut-off values to allow for laboratory-independent clinical decision-making.
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Down, Liz, Chris Metcalfe, Richard M. Martin, David E. Neal, Freddie C. Hamdy, Jenny L. Donovan und J. Athene Lane. „Seasonal variation in prostate-specific antigen levels: a large cross-sectional study of men in the UK“. BJU International 108, Nr. 9 (31.03.2011): 1409–14. http://dx.doi.org/10.1111/j.1464-410x.2011.10174.x.
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Saarinen, Niila V. V., Jussi Lehtonen, Riitta Veijola, Johanna Lempainen, Mikael Knip, Heikki Hyöty, Olli H. Laitinen und Vesa P. Hytönen. „Multiplexed High-Throughput Serological Assay for Human Enteroviruses“. Microorganisms 8, Nr. 6 (26.06.2020): 963. http://dx.doi.org/10.3390/microorganisms8060963.
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Immunological assays detecting antibodies against enteroviruses typically use a single enterovirus serotype as antigen. This limits the ability of such assays to detect antibodies against different enterovirus types and to detect possible type-specific variation in antibody responses. We set out to develop a multiplexed assay for simultaneous detection of antibodies against multiple enterovirus and rhinovirus types encompassing all human infecting species. Seven recombinant VP1 proteins from enteroviruses EV-A to EV-D and rhinoviruses RV-A to RV-C species were produced. Using Meso Scale Diagnostics U-PLEX platform we were able to study antibody reactions against these proteins as well as non-structural enterovirus proteins in a single well with 140 human serum samples. Adults had on average 33-fold stronger antibody responses to these antigens (p < 10−11) compared to children, but children had less cross-reactivity between different enterovirus types. The results suggest that this new high-throughput assay offers clear benefits in the evaluation of humoral enterovirus immunity in children, giving more exact information than assays that are based on a single enterovirus type as antigen.
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Wagner, D. K., J. York-Jolley, T. R. Malek, J. A. Berzofsky und D. L. Nelson. „Antigen-specific murine T cell clones produce soluble interleukin 2 receptor on stimulation with specific antigens.“ Journal of Immunology 137, Nr. 2 (15.07.1986): 592–96. http://dx.doi.org/10.4049/jimmunol.137.2.592.
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Abstract In this study, we used monoclonal antibodies to the murine IL 2 receptor (IL 2R) termed 3C7 and 7D4, which bind to different epitopes on the murine IL 2R, to develop an ELISA to measure soluble murine IL 2R. Surprisingly, stimulated murine spleen cells not only expressed cell-associated IL 2R, but also produced a considerable level of cell free IL 2R in the culture supernatant fluid. To assess the fine specificity of this response, myoglobin-immune murine T cell clones were stimulated with appropriate or inappropriate antigen and syngeneic or allogeneic presenting cells. Proliferation, measured by [3H] thymidine incorporation, and levels of soluble IL 2R were determined at day 4. The production of soluble IL 2R displayed the same epitope fine specificity, genetic restriction, and antigen dose-response as the proliferative response. Indeed, in some cases there was sharper discrimination of epitope specificity and genetic restriction with the soluble IL 2R levels. There was also reproducible clone-to-clone variation in the amount of soluble receptor produced in response to antigen among 12 T cell clones and lines tested. In time course experiments, proliferation was greatest at day 3, whereas soluble IL 2R levels continued to rise in subsequent days. To our knowledge, this is the first demonstration of release or secretion of soluble IL 2R by murine T cells, and the first demonstration of the fine specificity and genetic restriction of the induction of soluble IL 2R by specific antigen.
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Wallnöfer, A. E., J. M. T. van Griensven, H. C. Schoemaker, A. F. Cohen, W. Lambert, C. Kluft, P. Meijer und T. Kooistra. „Effect of Isotretinoin on Endogenous Tissue-Type Plasminogen Activator (t-PA) and Plasminogen Activator Inhibitor 1 (PAI-1) in Humans“. Thrombosis and Haemostasis 70, Nr. 06 (1993): 1005–8. http://dx.doi.org/10.1055/s-0038-1649715.
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SummaryThe effect of isotretinoin on fibrinolysis was investigated in 10 healthy, male volunteers in a randomized, double-blind, crossover-designed study. Isotretinoin (40 mg) was administered in the morning and in the evening for 5 days. t-PA, u-PA and PAI-1 antigen and activity in plasma were measured every morning at 9 a.m. on days 1 to 4 and every 3 hours over 24 hours on day 5. Isotretinoin treatment had no significant stimulatory effect on endogenous t-PA antigen and activity in morning plasma samples nor on their circadian variation. Also, u-PA antigen levels did not change after isotretinoin treatment. Mean PAI-1 antigen and PAI activity in 9 a.m. plasma samples were non-significantly higher during isotretinoin than during placebo treatment. After treatment with isotretinoin a significant rise of fasting triglyceride plasma levels was observed as compared to placebo. The study shows that isotretinoin has no clinically significant effect on endogenous fibrinolysis.
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Craig, Vanessa J., Isabelle Arnold, Christiane Gerke, Minh Q. Huynh, Thomas Wündisch, Andreas Neubauer, Christoph Renner, Stanley Falkow und Anne Müller. „Gastric MALT lymphoma B cells express polyreactive, somatically mutated immunoglobulins“. Blood 115, Nr. 3 (21.01.2010): 581–91. http://dx.doi.org/10.1182/blood-2009-06-228015.
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Abstract Gastric B-cell lymphoma of mucosa-associated lymphoid tissue (MALT) arises against a background of chronic inflammation caused by persistent Helicobacter pylori infection. The clinical and histopathologic features of the human tumor can be reproduced by Helicobacter infection of BALB/c mice. In this study, we have analyzed the antibody sequences and antigen specificity of a panel of murine and human MALT lymphoma–derived antibodies. We find that a majority of tumors in patients as well as experimentally infected mice are monoclonal. The tumor immunoglobulin heavy chain genes have undergone somatic hypermutation, and approximately half of all tumors show evidence of intraclonal variation and positive and/or negative selective pressure. Recombinantly expressed MALT lymphoma antibodies bind with intermediate affinity to various unrelated self- and foreign antigens, including Helicobacter sonicate, immunoglobulin G (IgG), DNA, and stomach extract; antigen binding is blocked in a dose-dependent manner in competitive enzyme-linked immunosorbent assays. A strong bias toward the use of VH gene segments previously linked to autoantibodies and/or polyreactive antibodies in B-cell malignancies or autoimmune pathologies supports the experimental finding of polyreactivity. Our results suggest that MALT lymphoma development may be facilitated by an array of local self- and foreign antigens, providing direct antigenic stimulation of the tumor cells via their B-cell receptor.
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Lagattuta, Kaitlyn, Mujin Kwun und Soumya Raychaudhuri. „Quantifying how TCR sequence variation affects T cell fate at single-cell resolution“. Journal of Immunology 210, Nr. 1_Supplement (01.05.2023): 76.19. http://dx.doi.org/10.4049/jimmunol.210.supp.76.19.
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Abstract Mucosal-Associated Invariant T (MAIT) and Natural Killer T (NKT) cells demonstrate that some T cell transcriptional fates are driven by the T cell antigen receptor (TCR). Though activation through the TCR is integral to T cell differentiation, the contribution of TCR sequence features to other T cell transcriptional fates remains yet to be defined. In this study, we identify how αβ V(D)J recombination affects T cell differentiation at single-cell resolution. By applying Canonical Correlation Analysis (CCA) to 340,557 TCR clones collected from 256 individuals, we define a set of TCR scoring functions that quantify transcriptional fate predispositions conferred by the TCR. Unsurprisingly, the strongest fate predispositions correspond to cognate peptide presentation molecules: MR1- or CD1d-restricted PLZFhigh innate-like (MAIT/NKT) transcriptional fate (72.6-fold increase in fate likelihood for top percentile compared to bottom percentile TCRs, AUC = 0.85) and MHC class I-restricted CD8 fate versus MHC class II-restricted CD4 fate (18.7-fold increase, AUC = 0.75). However, our results also uncover that hydrophobic CDR3 residues promote regulatory T cell transcriptional states in both the CD8 and CD4 lineages, and that TCR sequence features preferred by thymic positive selection continue to promote memory formation in the periphery. Applying our TCR scoring functions to 10× dextramer-labeled T cells reveals that not all antigen-specific TCRs are equally capable of mounting an effector response. Even among T cells that recognize the same antigen, biophysical variation in the TCR sequence directs which T cells undergo transcriptional reprogramming to form immunological memory. Supported by grant from NIH (T32GM007753)
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Plikaytis, Brian D., Maria Stella, Giuseppe Boccadifuoco, Lisa M. DeTora, Mauro Agnusdei, Laura Santini, Brunella Brunelli et al. „Interlaboratory Standardization of the Sandwich Enzyme-Linked Immunosorbent Assay Designed for MATS, a Rapid, Reproducible Method for Estimating the Strain Coverage of Investigational Vaccines“. Clinical and Vaccine Immunology 19, Nr. 10 (08.08.2012): 1609–17. http://dx.doi.org/10.1128/cvi.00202-12.
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ABSTRACTThe meningococcal antigen typing system (MATS) sandwich enzyme-linked immunosorbent assay (ELISA) was designed to measure the immunologic cross-reactivity and quantity of antigens in target strains of a pathogen. It was first used to measure the factor H-binding protein (fHbp), neisserial adhesin A (NadA), and neisserial heparin-binding antigen (NHBA) content of serogroup B meningococcal (MenB) isolates relative to a reference strain, or “relative potency” (RP). With the PorA genotype, the RPs were then used to assess strain coverage by 4CMenB, a multicomponent MenB vaccine. In preliminary studies, MATS accurately predicted killing in the serum bactericidal assay using human complement, an accepted correlate of protection for meningococcal vaccines. A study across seven laboratories assessed the reproducibility of RPs for fHbp, NadA, and NHBA and established qualification parameters for new laboratories. RPs were determined in replicate for 17 MenB reference strains at laboratories A to G. The reproducibility of RPs among laboratories and against consensus values across laboratories was evaluated using a mixed-model analysis of variance. Interlaboratory agreement was very good; the Pearson correlation coefficients, coefficients of accuracy, and concordance correlation coefficients exceeded 99%. The summary measures of reproducibility, expressed as between-laboratory coefficients of variation, were 7.85% (fHbp), 16.51% (NadA), and 12.60% (NHBA). The overall within-laboratory measures of variation adjusted for strain and laboratory were 19.8% (fHbp), 28.8% (NHBA), and 38.3% (NadA). The MATS ELISA was successfully transferred to six laboratories, and a further laboratory was successfully qualified.
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Price, Lance B., Martin Hugh-Jones, Paul J. Jackson und Paul Keim. „Genetic Diversity in the Protective Antigen Gene ofBacillus anthracis“. Journal of Bacteriology 181, Nr. 8 (15.04.1999): 2358–62. http://dx.doi.org/10.1128/jb.181.8.2358-2362.1999.
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ABSTRACT Bacillus anthracis is a gram-positive spore-forming bacterium that causes the disease anthrax. The anthrax toxin contains three components, including the protective antigen (PA), which binds to eucaryotic cell surface receptors and mediates the transport of toxins into the cell. In this study, the entire 2,294-nucleotide protective antigen gene (pag) was sequenced from 26 of the most diverse B. anthracis strains to identify potential variation in the toxin and to further our understanding of B. anthracis evolution. Five point mutations, three synonymous and two missense, were identified. These differences correspond to six different haploid types, which translate into three different amino acid sequences. The two amino acid changes were shown to be located in an area near a highly antigenic region critical to lethal factor binding. Nested primers were used to amplify and sequence this same region of pag from necropsy samples taken from victims of the 1979 Sverdlovsk incident. This investigation uncovered five different alleles among the strains present in the tissues, including two not seen in the 26-sample survey. One of these two alleles included a novel missense mutation, again located just adjacent to the highly antigenic region. Phylogenetic (cladistic) analysis of thepag corresponded with previous strain grouping based on chromosomal variation, suggesting that plasmid evolution in B. anthracis has occurred with little or no horizontal transfer between the different strains.
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Nowak-Göttl, Ulrike, Birgit Fröhlich, Sabine Thedieck, Andreas Huge und Monika Stoll. „Association of the protein Z ATG haplotype with symptomatic nonvascular stroke or thromboembolism in white children: a family-based cohort study“. Blood 113, Nr. 10 (05.03.2009): 2336–41. http://dx.doi.org/10.1182/blood-2008-10-181461.
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Abstract To clarify the role of protein Z (PZ) in children with stroke/thromboembolism (TE), the present haplotype (HT)–based family study was performed. We genotyped 365 pediatric stroke/TE families (stroke n = 216; TE n = 149) for 4 single nucleotide polymorphisms (SNPs; rs3024718, rs3024731, rs3024772, and rs3024778) to assess the association between genetic variation within a conserved block of linkage disequilibrium harboring the PZ gene and pediatric TE. Association was assessed with use of the transmission disequilibrium test (TDT), corrected for multiple testing (permutation testing: HAPLOVIEW). In addition, PZ antigen was determined and correlated with carriership of PZ haplotypes and the FV G1691A mutation. Rs3024718, rs3024731, and rs3024772 are in tight linkage disequilibrium (LD) and define 4 haplotypes, capturing 97% of the genetic variation for this LD block. HT1 (ATG) was significantly overtransmitted from parents to affected offspring (HT frequency 73.5%, T:U 122:80, χ2 = 8.791, P = .003). The ATG risk haplotype was significantly correlated with greater PZ antigen levels. Multivariate analysis adjusted for age, sex, established thrombophilias, smoking, fibrinogen, and PZ levels revealed a significant association of the ATG haplotype and TE in children (odds ratio [OR] 1.4; 95% confidence interval [95% CI] 1.08-1.93). Our results suggest that the ATG haplotype of the PZ gene is a genetic marker for symptomatic TE in white German children.