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- ItemAcesso aberto (Open Access)Analysis of proteins from membrane and soluble fractions of Giardia duodenalis trophozoites of two Brazilian axenic strains(Instituto de Medicina Tropical, 2002-10-01) Guimarães, Semíramis; Sogayar, Maria Inês T. Leme; Franco, Marcello Fabiano de [UNIFESP]; Universidade Estadual Paulista (UNESP); Universidade Federal de São Paulo (UNIFESP)In the present study, we have analyzed by sodium docecyl sulphate - polyacrilamide gel electrophoresis (SDS-PAGE), immunoblotting and Concanavalin A blotting (Con A blotting) proteins of membrane fractions and soluble fractions obtained from Giardia duodenalis trophozoites of two axenic strains isolated in Brazil from a symptomatic (BTU-11) and an asymptomatic patient (BTU-10), as compared to the reference strain Portland 1. Both Brazilian strains showed a complex and homogeneous electrophoretic pattern of proteins, but some differences could be observed. Several glycoproteins were detected, particularly the proteins of 81, 72, 59 kDa and the protein of 62 kDa in the membrane proteins and cytosol, respectively. Many antigenic components were revealed by anti-Giardia rabbit IgG antibodies in the immunoblotting analysis. Among these components, the membrane protein of 32 kDa and the cytosol protein of 30 kDa could be related to giardin, as previously demonstrated.
- ItemAcesso aberto (Open Access)Aspectos imunológicos do diabetes melito tipo 1(Associação Médica Brasileira, 1999-04-01) Balda, C. A. [UNIFESP]; Pacheco-Silva, Alvaro [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)
- ItemSomente MetadadadosCharacteristics of Environmental Paracoccidioides brasiliensis Isolates(Springer, 2010-01-01) Costa, Patricia Fagundes; Fernandes, Geisa Ferreira [UNIFESP]; Santos, Priscila Oliveira dos [UNIFESP]; Amaral, Cristiane Candida [UNIFESP]; Camargo, Zoilo Pires [UNIFESP]; Universidade Federal de São Paulo (UNIFESP); Fed Univ ParaThe ecological niche or exact habitat of the fungus Paracoccidioides brasiliensis is not known, and few isolates have been obtained from the environment. in this study, ten isolates were analyzed with respect to antigenic composition, serology, pathogenicity, and molecular aspects. Gp43 is considered to be the molecular basis for the serodiagnosis of paracoccidioidomycosis; however, in this study only six of the environmental isolates secreted this molecule (four in great amounts and two in small amounts). Other molecules were also produced. When exoantigens from these isolates were tested using immunodiffusion, only four preparations were positive by ID tests. However, when these exoantigens were tested by ELISA, all of them except one were able to detect anti-P. brasiliensis antibodies. in Western blot assays, these exoantigens showed different reactivities. Isolates that secreted gp43 presented positive reactions for this molecule, and isolates that did not secrete gp43 gave positive reactions for other minor molecules. RAPD analysis revealed that there is great genetic variation between these environmental isolates. These isolates were non-pathogenic: no mortality was observed among the inoculated mice during an 18-month follow-up period.
- ItemAcesso aberto (Open Access)Fenotipagem e aloimunização eritrocitária em portadores de anemia falciforme atendidos no Hemocentro de Alagoas(Universidade Federal de São Paulo (UNIFESP), 2010-11-24) Pinto, Patrícia Costa Alves [UNIFESP]; Dos Santos, Amelia Miyashiro [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)Objective: To determine the erythrocyte phenotyping in blood donors and patients with sickle cell anemia (SS) assisted at Hemocentro of Alagoas and also the frequency and factors associated with erythrocyte alloimmunization. Methods: Cross-sectional study with 102 patients with sickle cell anemia (SS) and 100 blood donors. This study was approved by the Ethical Committee of the Federal University of São Paulo and by the Ethical Committee of the University of Science of Health of Alagoas. Patients and blood donors were requested to sign an informed consent form prior to enrolment. The following tests were performed: erythrocyte phenotyping by gel centrifugation, Direct antiglobulin test, Indirect antiglobulin test, and detection of irregular antibodies by panels of phenotyped red blood cells. Variables were compared by Mann-Whitney test and c 2 test or exact Fisher test. Factors associate with alloimmunization was analyzed by univariate and multiple logistic regression analysis. Differences were considered significant if p<0.002. Results: The most frequent antigens found in patients and blood donors were: c (93.1% vs. 83.0%), and (96.1% vs. 94.0%), M (82.4% vs. 82.0%), s (92.2% vs. 79.0%), JK(a) (88.2% vs. 82.0), respectively. Significant differences were observed between the frequency of the phenotype of patients and donors in regard to antigens s, FY(a) and JK(b). From 79 transfused patients, 10 (12.7%) presented positive Indirect Coombs. Thirteen alloantibodies were found, 7 from Rh system, 2 from Kell and 4 were not identified. Factors associated with alloimmunization were the period of time between the last transfusion and the data of the test and being submitted to more than 10 red blood cell transfusions. Patients who received more than 10 transfusions presented a likelihood of presenting alloimmunization 16.39 (2.23-120.59) times higher than who received less than 10 transfusions. All patients presented a negative Direct antiglobulin test. Conclusion: The prevalence of alloimmunization in patients with sickle cell anemia was 12.7% with 70% of antibodies belonging to Rh and Kell systems. This study shows the importance of performing erythrocyte phenotyping in blood donors and receptors in order to decrease the risk of alloimmunization.
- ItemAcesso aberto (Open Access)Giardia duodenalis: INTER-STRAIN VARIABILITY OF PROTEINS, ANTIGENS, PROTEASES, ISOENZYMES AND NUCLEIC ACIDS(Instituto de Medicina Tropical, 1999-01-01) Guimarães, Semiramis; Sogayar, Maria Inês Leme; Franco, Marcello Fabiano de [UNIFESP]; Universidade Estadual Paulista (UNESP); Universidade Federal de São Paulo (UNIFESP)Giardia duodenalis isolates from asymptomatic or symptomatic patients and from animals present similarities and differences in the protein composition, antigenic profile, pattern of proteases and isoenzymes, as well as in nucleic acids analysis.In the present overview, these differences and similarities are reviewed with emphasis in the host-parasite interplay and possible mechanisms of virulence of the protozoon.
- ItemAcesso aberto (Open Access)Isolation, cultivation and characterization of CD133+ stem cells from human glioblastoma(Instituto Israelita de Ensino e Pesquisa Albert Einstein, 2012-06-01) Pavon, Lorena Favaro; Marti, Luciana Cavalheiro; Sibov, Tatiana Tais; Miyaki, Liza Aya Mabuchi; Malheiros, Suzana Maria Fleury [UNIFESP]; Mamani, Javier Bustamante; Brandt, Reynaldo Andre; Ribas, Guilherme Carvalhal; Pagura, Jorge Roberto; Joaquim, Marcos Augusto Stavale; Feres Junior, Hallin; Gamarra, Lionel Fernel; Hospital Israelita Albert Einstein Instituto do Cérebro; Hospital Israelita Albert Einstein Centro de Pesquisa Experimental; Hospital Israelita Albert Einstein Faculdade de Enfermagem; Universidade Federal de São Paulo (UNIFESP); Hospital Israelita Albert Einstein Center for Neuro-oncology; Hospital Israelita Albert EinsteinOBJECTIVE: To establish the method of isolation and culture of human glioblastoma neurospheres, and the purification of their stem cells, followed by the process of obtaining tumor subspheres, immunophenotypically characterizing this clonogenic set. METHODS: Through the processing of glioblastoma samples (n=3), the following strategy of action was adopted: (i) establish primary culture of glioblastoma; (ii) isolation and culture of tumor neurospheres; (iii) purify cells that initiate tumors (CD133+) by magnetic separation system (MACS); (iv) obtain tumor subspheres; (v) study the expression of the markers nestin, CD133, and GFAP. RESULTS: The study successfully described the process of isolation and culture of glioblastoma subspheres, which consist of a number of clonogenic cells immunophenotypically characterized as neural, which are able to initiate tumor formation. CONCLUSION: These findings may contribute to a better understanding of the process of gliomagenesis.
- ItemAcesso aberto (Open Access)Lymphocyte transformation assay for C neoformans antigen is not reliable for detecting cellular impairment in patients with Neurocryptococcosis(Biomed Central Ltd, 2012-10-30) Rocha, Katya C.; Pinhal, Cinthia; Cavalcanti, Sonia; Vidal, Monica S. M.; Toscano, Matheus; Moraes-Vasconcelos, Dewton; Duarte, Alberto J. S.; Fonseca, Fernando Luiz Affonso [UNIFESP]; Abreu, Luiz Carlos de; Valenti, Vitor E.; Grumach, Anete S. G.; Universidade de São Paulo (USP); Fac Med ABC; Universidade Federal de São Paulo (UNIFESP); Univ Estadual PaulistaBackground: Cryptococcus neoformans causes meningitis and disseminated infection in healthy individuals, but more commonly in hosts with defective immune responses. Cell-mediated immunity is an important component of the immune response to a great variety of infections, including yeast infections. We aimed to evaluate a specific lymphocyte transformation assay to Cryptococcus neoformans in order to identify immunodeficiency associated to neurocryptococcosis (NCC) as primary cause of the mycosis.Methods: Healthy volunteers, poultry growers, and HIV-seronegative patients with neurocryptococcosis were tested for cellular immune response. Cryptococcal meningitis was diagnosed by India ink staining of cerebrospinal fluid and cryptococcal antigen test (Immunomycol-Inc, SP, Brazil). Isolated peripheral blood mononuclear cells were stimulated with C. neoformans antigen, C. albicans antigen, and pokeweed mitogen. the amount of H-3-thymidine incorporated was assessed, and the results were expressed as stimulation index (SI) and log SI, sensitivity, specificity, and cut-off value (receiver operating characteristics curve). We applied unpaired Student t tests to compare data and considered significant differences for p<0.05.Results: the lymphotoxin alpha showed a low capacity with all the stimuli for classifying patients as responders and non-responders. Lymphotoxin alpha stimulated by heated-killed antigen from patients with neurocryptococcosis was not affected by TCD4+ cell count, and the intensity of response did not correlate with the clinical evolution of neurocryptococcosis.Conclusion: Response to lymphocyte transformation assay should be analyzed based on a normal range and using more than one stimulator. the use of a cut-off value to classify patients with neurocryptococcosis is inadequate. Statistical analysis should be based on the log transformation of SI. A more purified antigen for evaluating specific response to C. neoformans is needed.
- ItemSomente MetadadadosMapeamento do epitopo de células T do antígeno principal de diagnóstico do paracoccidioides brasilienses, indutor de resposta imune protetora do tipo Th1 contra a infecção experimental em camundongo(Universidade Federal de São Paulo (UNIFESP), 1998) Taborda, Carlos Pelleschi [UNIFESP]; Travassos, Luiz Rodolpho [UNIFESP]A glicoproteína de 43 kDa do Paracoccidioides brasiliensis é o principalantígeno diagnóstico da paracoccidioidomicose, micose sistêmica prevalente na América Latina. Além de induzir altos títulos de anticorpos, a gp43 é também imunodominante nas reações de hipersensibilidade do tipo tardio em animais e humanos infectados. A resposta imune em camundongos imunizados com gp43 na presença de adjuvante completo de Freund envolve linfócitos T CD4+ Th1, secretando INF-y e IL-2 mas não IL-4 e IL-10. O epitopo de célula T desse antígeno foi mapeado a partir de um peptídio de 15 amino-ácidos obtido através de ensaios de linfoproliferação com células de camundongo de três diferentes haplótipos e análise proteíca por computador. A estrutura mínima exigida para o epitopo de células T foi determinada pelo teste de uma série de peptídios análogos ou truncados do P10. Somente peptídios com 12 ou mais amino ácidos eram ativos, confirmando a apresentação pelo sistema de MHC classe II. o "core" interno formado pelos amino ácidos HTLAIR é essencial para o epítopo, podendo ser utilizadas várias regiões flanqueadoras. A imunização de camundonggos com a gp43 ou P10 induziu a uma significativa proteção em camundongos Balb/c, desafiados intra-traquealmente com leveduras virulentas de P. brasiliensis, apresentando um número 200 vezes menor de CFUs no pulmão em relação aos animais não induzidos. O efeito protetor do P10 é principalmente atribuído á resposta imune celular medida pelo IFN-y. Somente a gp43 era capaz de induzir uma resposta com anticorpos compatível com ativação de células Th1 e Th2 em camundongos Balb/c infectados, enquanto que P10 não induziu resposta humoral. A proteção induzida pela gp43 ou P10 era caracterizada por poucas leveduras viáveis em granulomas epitelióides bem delimitados nos pulmões ou por lesões resolvidas com fúngicas não detectáveis. Camundongos INF-y-/- eram muito susceptíveis à infecção intratraqueal pelo P. brasiliensis com alta mortalidade e morbidade, caracterizada pela intensa proliferação fúngica e disseminação para o fígado e baço. Contrariamente ao tipo selvagem, todos os animais Kos morreram após 5 semanas de infecção pelo fungo.
- ItemAcesso aberto (Open Access)Tolerância oral: uma nova perspectiva no tratamento de doenças autoimunes(Associação Médica Brasileira, 1999-03-01) Bueno, Valquiria [UNIFESP]; Pacheco-Silva, Alvaro [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)