Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana
Autor(a) principal: | |
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Data de Publicação: | 2008 |
Tipo de documento: | Tese |
Idioma: | por |
Título da fonte: | Repositório Institucional da UFU |
Texto Completo: | https://repositorio.ufu.br/handle/123456789/16620 |
Resumo: | Neurocysticercosis (NC) caused by Taenia solium metacestodes usually affects the central nervous system and can be confused with other brain pathologies. The serological diagnosis of NC is of great importance in routine clinical management of patients because it supports the diagnosis in patients that have clinical and radiological findings compatible with this disease. The use of purified antigens has allowed the development of more sensitive and specific tests, resulting in reliable seroepidemiological and screening studies. Purification procedures using Triton X-114 (TX-114) and affinity chromatography are simple and low cost methods. The aim of this study was to characterize and to compare different antigenic fractions from crude saline extract of T. solium metacestodes obtained by phase partitioning using TX-114 and affinity chromatography for the serodiagnosis of human NC. A total of 132 serum samples were analyzed, from which 40 were of patients with NC, 62 were of patients with other parasitic diseases and 30 were from apparently healthy individuals. The crude saline extract (S) and the cyst vesicular fluid (VF) were submitted to phase partitioning using TX-114, resulting in following fractions: detergent (DS and DVF, respectively) and aqueous (AS and AVF, respectively) phases. The extract S was purified in jacalin or N-acetyl-glucosamine columns, but only the jacalin-unbound fraction was then submitted to Concanavalin A column or TX-114 partitioning. All antigens were analyzed in SDS-PAGE 12% and antigen bands were visualized by silver staining. Serum samples were evaluated for IgG detection at 1:200 dilution for ELISA and 1:100 dilution for immunoblotting. Sensitivity, specificity, diagnostic efficiency and Youden Index were calculated for ELISA. ELISA sensitivity and specificity were 92.5% and 84.8% (extract S); 92.5% and 92.4% (DS); 67.5% and 70.6% (AS); 82.5% and 85.9% (VF); 77.5% and 72.8 (DVF); 80.0 and 80.4% (AVF); 85.0% and 84.8% (jacalin-unbound fraction); 92.5% and 93.5% (Detergent jacalin-unbound fraction); 82.5% and 82.6 % (Aqueous jacalin-unbound fraction), respectively. ELISA diagnostic efficiency and Youden Index were: 87.1% and 0.77 (extract S); 92.4% and 0.85 (DS); 69.7% and 0.38 (AS); 84.8% and 0.68 (VF); 74.2% and 0.50 (DVF); 80.3% and 0.60 (AVF); 84.8% and 0.70 (jacalin-unbound fraction); 93.2% and 0.86 (Detergent jacalinunbound fraction); 82.6% and 0.65 (Aqueous jacalin-unbound fraction), respectively. By immunoblotting, all serum samples from NC patients recognized a 70-50 kDa band in the detergent jacalin-unbound fraction whereas no reactivity for this antigenic fraction was observed in serum samples from patients with other parasitoses or healthy individuals. In conclusion, the detergent jacalin-unbound fraction showed to be more sensitive and specific than other extracts, since it was able to discriminate NC patients from other parasitoses, particularly Echinococcus granulosus-infected patients. |
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Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humanaTaenia soliumTriton X-114Cromatografia de afinidadeNeurocisticercoseTaeniaTaenia soliumTriton X-114Affinity chromatographyDiagnosisNeurocysticercosisCNPQ::CIENCIAS BIOLOGICAS::IMUNOLOGIA::IMUNOLOGIA APLICADANeurocysticercosis (NC) caused by Taenia solium metacestodes usually affects the central nervous system and can be confused with other brain pathologies. The serological diagnosis of NC is of great importance in routine clinical management of patients because it supports the diagnosis in patients that have clinical and radiological findings compatible with this disease. The use of purified antigens has allowed the development of more sensitive and specific tests, resulting in reliable seroepidemiological and screening studies. Purification procedures using Triton X-114 (TX-114) and affinity chromatography are simple and low cost methods. The aim of this study was to characterize and to compare different antigenic fractions from crude saline extract of T. solium metacestodes obtained by phase partitioning using TX-114 and affinity chromatography for the serodiagnosis of human NC. A total of 132 serum samples were analyzed, from which 40 were of patients with NC, 62 were of patients with other parasitic diseases and 30 were from apparently healthy individuals. The crude saline extract (S) and the cyst vesicular fluid (VF) were submitted to phase partitioning using TX-114, resulting in following fractions: detergent (DS and DVF, respectively) and aqueous (AS and AVF, respectively) phases. The extract S was purified in jacalin or N-acetyl-glucosamine columns, but only the jacalin-unbound fraction was then submitted to Concanavalin A column or TX-114 partitioning. All antigens were analyzed in SDS-PAGE 12% and antigen bands were visualized by silver staining. Serum samples were evaluated for IgG detection at 1:200 dilution for ELISA and 1:100 dilution for immunoblotting. Sensitivity, specificity, diagnostic efficiency and Youden Index were calculated for ELISA. ELISA sensitivity and specificity were 92.5% and 84.8% (extract S); 92.5% and 92.4% (DS); 67.5% and 70.6% (AS); 82.5% and 85.9% (VF); 77.5% and 72.8 (DVF); 80.0 and 80.4% (AVF); 85.0% and 84.8% (jacalin-unbound fraction); 92.5% and 93.5% (Detergent jacalin-unbound fraction); 82.5% and 82.6 % (Aqueous jacalin-unbound fraction), respectively. ELISA diagnostic efficiency and Youden Index were: 87.1% and 0.77 (extract S); 92.4% and 0.85 (DS); 69.7% and 0.38 (AS); 84.8% and 0.68 (VF); 74.2% and 0.50 (DVF); 80.3% and 0.60 (AVF); 84.8% and 0.70 (jacalin-unbound fraction); 93.2% and 0.86 (Detergent jacalinunbound fraction); 82.6% and 0.65 (Aqueous jacalin-unbound fraction), respectively. By immunoblotting, all serum samples from NC patients recognized a 70-50 kDa band in the detergent jacalin-unbound fraction whereas no reactivity for this antigenic fraction was observed in serum samples from patients with other parasitoses or healthy individuals. In conclusion, the detergent jacalin-unbound fraction showed to be more sensitive and specific than other extracts, since it was able to discriminate NC patients from other parasitoses, particularly Echinococcus granulosus-infected patients.Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorDoutor em Imunologia e Parasitologia AplicadasA neurocisticercose (NC) causada pela forma metacestódea de Taenia solium freqüentemente acomete o sistema nervoso central podendo ser confundida com outras síndromes neurológicas. O diagnóstico sorológico da NC é de grande importância na rotina laboratorial porque auxilia no diagnóstico de pacientes que apresentam sinais clínicos e radiológicos compatíveis com esta doença. A utilização de antígenos purificados tem permitido o desenvolvimento de testes mais sensíveis e específicos, resultando em estudos soroepidemiológicos e de triagem mais confiáveis. Processos de purificação antigênica utilizando Triton X-114 (TX-114) e cromatografia de afinidade consistem em técnicas simples e de baixo custo. O objetivo deste estudo foi caracterizar e comparar diferentes frações antigênicas da forma metacestódea de T. solium obtidos por separação de fases usando TX-114 e cromatografia de afinidade para o sorodiagnóstico da NC humana. Foram analisadas 132 amostras de soro, sendo 40 de pacientes com NC, 62 de pacientes com outras parasitoses e 30 de indivíduos aparentemente saudáveis. O extrato salino total (S) e o líquido de vesícula (LV) foram submetidos ao processo de fracionamento utilizando TX-114 para a obtenção das frações antigênicas: detergente (DS e DLV, respectivamente) e aquosa (AS e ALV, respectivamente). O extrato S foi purificado em colunas de jacalina e N-acetilglicosamina, sendo apenas a fração não ligante de jacalina (FNL-J) submetida à coluna de concanavalina A (ConA) e fracionamento com TX-114. Todos antígenos foram analisados em SDS-PAGE 12% e corados pela prata para visualização das bandas antigênicas. As amostras de soro foram avaliadas para detecção de IgG pelo teste ELISA na diluição de 1:200 e pelo immunoblotting na diluição de 1:100. Sensibilidade, especificidade, eficiência do diagnóstico e Índice de Youden foram calculados para o teste ELISA. As sensibilidades e especificidades dos testes ELISA foram: 92,5% e 84,8% (S); 92,5% e 92,4% (DS); 67,5% e 70,6% (AS); 82,5% e 85,9% (LV); 77,5% e 72,8% (DLV); 80,0% e 80,4% (ALV); 85,0% e 84,8% (FNL-J); 92,5% e 93,5% (DFNL-J); 82,5% e 82,6% (AFNL-J), respectivamente. A eficiência do diagnóstico e o Índice de Youden dos testes ELISA foram: 87,1% e 0,77 (S); 92,4% e 0,85 (DS); 69,7% e 0,38 (AS); 84,8% e 0,68 (LV); 74,2% e 0,50 (DLV); 80,3% e 0,60 (ALV); 84,8% e 0,70 (FNL-J); 93,2% e 0,86 (DFNL-J); 82,6% e 0,65% (AFNL-J), respectivamente. No immunoblotting a banda de 70-50 kDa foi detectada em todas as amostras de soro de pacientes com NC para a fração DFNL-J, enquanto nenhuma reatividade foi observada para esta fração antigênica nas amostras de pacientes com outras parasitoses ou indivíduos saudáveis. Concluindo, a fração DFNL-J foi mais sensível e específica que os outros extratos, sendo capaz de diferenciar pacientes com NC daqueles com outras parasitoses, particularmente nas infecções por Echinococcus granulosus.Universidade Federal de UberlândiaBRPrograma de Pós-graduação em Imunologia e Parasitologia AplicadasCiências BiológicasUFUMineo, Jose Robertohttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4780327P9Costa-Cruz, Julia Mariahttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4797155D9Taketomi, Ernesto Akiohttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4788625D6Espindola, Foued Salmenhttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4727087Y6Takayanagui, Osvaldo Massaitihttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4781698D6Vergara, Mário Leon Silvahttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4799608Y0Machado, Gleyce Alves2016-06-22T18:46:26Z2008-04-112016-06-22T18:46:26Z2008-03-25info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesisapplication/pdfapplication/pdfMACHADO, Gleyce Alves. Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana . 2008. 111 f. Tese (Doutorado em Ciências Biológicas) - Universidade Federal de Uberlândia, Uberlândia, 2008.https://repositorio.ufu.br/handle/123456789/16620porinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UFUinstname:Universidade Federal de Uberlândia (UFU)instacron:UFU2021-09-16T17:54:47Zoai:repositorio.ufu.br:123456789/16620Repositório InstitucionalONGhttp://repositorio.ufu.br/oai/requestdiinf@dirbi.ufu.bropendoar:2021-09-16T17:54:47Repositório Institucional da UFU - Universidade Federal de Uberlândia (UFU)false |
dc.title.none.fl_str_mv |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
title |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
spellingShingle |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana Machado, Gleyce Alves Taenia solium Triton X-114 Cromatografia de afinidade Neurocisticercose Taenia Taenia solium Triton X-114 Affinity chromatography Diagnosis Neurocysticercosis CNPQ::CIENCIAS BIOLOGICAS::IMUNOLOGIA::IMUNOLOGIA APLICADA |
title_short |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
title_full |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
title_fullStr |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
title_full_unstemmed |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
title_sort |
Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana |
author |
Machado, Gleyce Alves |
author_facet |
Machado, Gleyce Alves |
author_role |
author |
dc.contributor.none.fl_str_mv |
Mineo, Jose Roberto http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4780327P9 Costa-Cruz, Julia Maria http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4797155D9 Taketomi, Ernesto Akio http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4788625D6 Espindola, Foued Salmen http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4727087Y6 Takayanagui, Osvaldo Massaiti http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4781698D6 Vergara, Mário Leon Silva http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4799608Y0 |
dc.contributor.author.fl_str_mv |
Machado, Gleyce Alves |
dc.subject.por.fl_str_mv |
Taenia solium Triton X-114 Cromatografia de afinidade Neurocisticercose Taenia Taenia solium Triton X-114 Affinity chromatography Diagnosis Neurocysticercosis CNPQ::CIENCIAS BIOLOGICAS::IMUNOLOGIA::IMUNOLOGIA APLICADA |
topic |
Taenia solium Triton X-114 Cromatografia de afinidade Neurocisticercose Taenia Taenia solium Triton X-114 Affinity chromatography Diagnosis Neurocysticercosis CNPQ::CIENCIAS BIOLOGICAS::IMUNOLOGIA::IMUNOLOGIA APLICADA |
description |
Neurocysticercosis (NC) caused by Taenia solium metacestodes usually affects the central nervous system and can be confused with other brain pathologies. The serological diagnosis of NC is of great importance in routine clinical management of patients because it supports the diagnosis in patients that have clinical and radiological findings compatible with this disease. The use of purified antigens has allowed the development of more sensitive and specific tests, resulting in reliable seroepidemiological and screening studies. Purification procedures using Triton X-114 (TX-114) and affinity chromatography are simple and low cost methods. The aim of this study was to characterize and to compare different antigenic fractions from crude saline extract of T. solium metacestodes obtained by phase partitioning using TX-114 and affinity chromatography for the serodiagnosis of human NC. A total of 132 serum samples were analyzed, from which 40 were of patients with NC, 62 were of patients with other parasitic diseases and 30 were from apparently healthy individuals. The crude saline extract (S) and the cyst vesicular fluid (VF) were submitted to phase partitioning using TX-114, resulting in following fractions: detergent (DS and DVF, respectively) and aqueous (AS and AVF, respectively) phases. The extract S was purified in jacalin or N-acetyl-glucosamine columns, but only the jacalin-unbound fraction was then submitted to Concanavalin A column or TX-114 partitioning. All antigens were analyzed in SDS-PAGE 12% and antigen bands were visualized by silver staining. Serum samples were evaluated for IgG detection at 1:200 dilution for ELISA and 1:100 dilution for immunoblotting. Sensitivity, specificity, diagnostic efficiency and Youden Index were calculated for ELISA. ELISA sensitivity and specificity were 92.5% and 84.8% (extract S); 92.5% and 92.4% (DS); 67.5% and 70.6% (AS); 82.5% and 85.9% (VF); 77.5% and 72.8 (DVF); 80.0 and 80.4% (AVF); 85.0% and 84.8% (jacalin-unbound fraction); 92.5% and 93.5% (Detergent jacalin-unbound fraction); 82.5% and 82.6 % (Aqueous jacalin-unbound fraction), respectively. ELISA diagnostic efficiency and Youden Index were: 87.1% and 0.77 (extract S); 92.4% and 0.85 (DS); 69.7% and 0.38 (AS); 84.8% and 0.68 (VF); 74.2% and 0.50 (DVF); 80.3% and 0.60 (AVF); 84.8% and 0.70 (jacalin-unbound fraction); 93.2% and 0.86 (Detergent jacalinunbound fraction); 82.6% and 0.65 (Aqueous jacalin-unbound fraction), respectively. By immunoblotting, all serum samples from NC patients recognized a 70-50 kDa band in the detergent jacalin-unbound fraction whereas no reactivity for this antigenic fraction was observed in serum samples from patients with other parasitoses or healthy individuals. In conclusion, the detergent jacalin-unbound fraction showed to be more sensitive and specific than other extracts, since it was able to discriminate NC patients from other parasitoses, particularly Echinococcus granulosus-infected patients. |
publishDate |
2008 |
dc.date.none.fl_str_mv |
2008-04-11 2008-03-25 2016-06-22T18:46:26Z 2016-06-22T18:46:26Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/doctoralThesis |
format |
doctoralThesis |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
MACHADO, Gleyce Alves. Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana . 2008. 111 f. Tese (Doutorado em Ciências Biológicas) - Universidade Federal de Uberlândia, Uberlândia, 2008. https://repositorio.ufu.br/handle/123456789/16620 |
identifier_str_mv |
MACHADO, Gleyce Alves. Análise de frações antigênicas da forma metacestódea de Taenia solium, obtidas pelo Triton X-114 e por cromatografia de afinidade, no diagnóstico sorológico da neurocisticercose humana . 2008. 111 f. Tese (Doutorado em Ciências Biológicas) - Universidade Federal de Uberlândia, Uberlândia, 2008. |
url |
https://repositorio.ufu.br/handle/123456789/16620 |
dc.language.iso.fl_str_mv |
por |
language |
por |
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info:eu-repo/semantics/openAccess |
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openAccess |
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application/pdf application/pdf |
dc.publisher.none.fl_str_mv |
Universidade Federal de Uberlândia BR Programa de Pós-graduação em Imunologia e Parasitologia Aplicadas Ciências Biológicas UFU |
publisher.none.fl_str_mv |
Universidade Federal de Uberlândia BR Programa de Pós-graduação em Imunologia e Parasitologia Aplicadas Ciências Biológicas UFU |
dc.source.none.fl_str_mv |
reponame:Repositório Institucional da UFU instname:Universidade Federal de Uberlândia (UFU) instacron:UFU |
instname_str |
Universidade Federal de Uberlândia (UFU) |
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UFU |
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UFU |
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Repositório Institucional da UFU |
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Repositório Institucional da UFU |
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Repositório Institucional da UFU - Universidade Federal de Uberlândia (UFU) |
repository.mail.fl_str_mv |
diinf@dirbi.ufu.br |
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1805569606513852416 |