Desenvolvimento de sensor bioletroquímico para detecção da hanseníase
Ano de defesa: | 2008 |
---|---|
Autor(a) principal: | |
Orientador(a): | |
Banca de defesa: | , |
Tipo de documento: | Dissertação |
Tipo de acesso: | Acesso aberto |
Idioma: | por |
Instituição de defesa: |
Universidade Federal de Uberlândia
|
Programa de Pós-Graduação: |
Programa de Pós-graduação em Química
|
Departamento: |
Ciências Exatas e da Terra
|
País: |
BR
|
Palavras-chave em Português: | |
Área do conhecimento CNPq: | |
Link de acesso: | https://repositorio.ufu.br/handle/123456789/17293 |
Resumo: | Leprosy is a chronic, granulomatous and dermatological disease, from infection caused by the bacillus Mycobacterium leprae. It is object of intervention of the state, because when untreated cause disability in the economically active population. Brazil remains one of the four endemic countries responsible for 34% of all cases registered at the beginning of 2007. The disease diagnosis can only be done after the onset of disease symptoms. Prophylactic measures are possible when using techniques such as ELISA and PCR, which can be performed only in specialized laboratories, not considering the needs of the endemic countries. Such problems can be circumvented by using tools of low cost, simple and sensitive handling as biosensors. In this work, poly(4-aminophenol) matrix, was used for immobilization and detection of specific PCR product for Hansen bacillus. The surface modification with the film was confirmed observing the electrochemical differences of the electrode without modification front at redox pair Fe2+/Fe3+ and supported electrolyte. The characterization of the polymeric matrix was carried out by FTIR, electrochemical impedance spectroscopy, electrochemical quartz crystal microbalance and studies of thermal stability. The FTIR data indicate polymerization possible by C-N-C bonds. The stability of the film at 98 ° C was measured and even showed 60% of its electroative properties after twenty minutes of heat treatment. being, therefore, adequate for the used methodology, in this work, for biomolecules immobilization. The detection of nitrogenated bases on the new matrix was possible in pH 4.5 and 7.4. The eletrolite, phosphate buffer in pH 7.4, was the most adequate to nitrogenated bases detection and in turn, for PCR products. This electrolyte was used for immobilization of the denaturated PCR product (RLEP3), contained 78pb, and specific for the Hansen bacillus. The DNA fragment immobilization was confirmed monitoring the oxidation peaks of guanosine monophosphate and adenosine monophosphate and also by means of electrochemical impedance spectroscopy studies. Hybridization indicator, ferrocenecarboxyaldehyde and methylene blue, had been used to differentiate complementary or not complementary target. With the hibridization occurrence, a reduction in the current signal values was observed, for both indicators, monitoring the indicator. The small detection limit, 10-10 mol.L-1, of the complementary target, was reached using methylene blue as indicator. When the probe was submitted to not complementary target, it was not observed change in the current values of immobilized probe. At the end of thirty days the biosensor kept 60% of their electrochemical properties These studies demonstrated the viability of constructing of an electrochemical biosensor using eletroactive indicators of low-cost, which can be used in real samples with the DNA of Hansen bacillus. For the first time, in the open literature, it was demonstred the modified electrodes production with poly(4-aminophenol) as matrix to immobilization and detection of specific PCRproduct to leprosy detection. |
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2016-06-22T18:48:21Z2014-02-192016-06-22T18:48:21Z2008-02-22AFONSO, André Santiago. Desenvolvimento de sensor bioletroquímico para detecção da hanseníase. 2008. 105 f. Dissertação (Mestrado em Ciências Exatas e da Terra) - Universidade Federal de Uberlândia, Uberlândia, 2008.https://repositorio.ufu.br/handle/123456789/17293Leprosy is a chronic, granulomatous and dermatological disease, from infection caused by the bacillus Mycobacterium leprae. It is object of intervention of the state, because when untreated cause disability in the economically active population. Brazil remains one of the four endemic countries responsible for 34% of all cases registered at the beginning of 2007. The disease diagnosis can only be done after the onset of disease symptoms. Prophylactic measures are possible when using techniques such as ELISA and PCR, which can be performed only in specialized laboratories, not considering the needs of the endemic countries. Such problems can be circumvented by using tools of low cost, simple and sensitive handling as biosensors. In this work, poly(4-aminophenol) matrix, was used for immobilization and detection of specific PCR product for Hansen bacillus. The surface modification with the film was confirmed observing the electrochemical differences of the electrode without modification front at redox pair Fe2+/Fe3+ and supported electrolyte. The characterization of the polymeric matrix was carried out by FTIR, electrochemical impedance spectroscopy, electrochemical quartz crystal microbalance and studies of thermal stability. The FTIR data indicate polymerization possible by C-N-C bonds. The stability of the film at 98 ° C was measured and even showed 60% of its electroative properties after twenty minutes of heat treatment. being, therefore, adequate for the used methodology, in this work, for biomolecules immobilization. The detection of nitrogenated bases on the new matrix was possible in pH 4.5 and 7.4. The eletrolite, phosphate buffer in pH 7.4, was the most adequate to nitrogenated bases detection and in turn, for PCR products. This electrolyte was used for immobilization of the denaturated PCR product (RLEP3), contained 78pb, and specific for the Hansen bacillus. The DNA fragment immobilization was confirmed monitoring the oxidation peaks of guanosine monophosphate and adenosine monophosphate and also by means of electrochemical impedance spectroscopy studies. Hybridization indicator, ferrocenecarboxyaldehyde and methylene blue, had been used to differentiate complementary or not complementary target. With the hibridization occurrence, a reduction in the current signal values was observed, for both indicators, monitoring the indicator. The small detection limit, 10-10 mol.L-1, of the complementary target, was reached using methylene blue as indicator. When the probe was submitted to not complementary target, it was not observed change in the current values of immobilized probe. At the end of thirty days the biosensor kept 60% of their electrochemical properties These studies demonstrated the viability of constructing of an electrochemical biosensor using eletroactive indicators of low-cost, which can be used in real samples with the DNA of Hansen bacillus. For the first time, in the open literature, it was demonstred the modified electrodes production with poly(4-aminophenol) as matrix to immobilization and detection of specific PCRproduct to leprosy detection.A hanseníase é uma doença crônica granulomatosa e dermatológica, proveniente de infecção causada pelo bacilo Mycobacterium leprae. É objeto de intervenção do estado, pois quando não tratada causa incapacidade na população economicamente ativa. O Brasil continua sendo um dos quatro países endêmicos responsáveis por 34% de todos os casos registrados no início de 2007. O diagnóstico da doença só pode ser feito depois do aparecimento dos sintomas da doença. Medidas profiláticas são possíveis quando se utilizam técnicas como ELISA e PCR, que podem ser realizados apenas em laboratórios especializados, não atendendo as necessidades dos países endêmicos. Tais problemas podem ser contornados utilizando ferramentas de baixo custo, simples manuseio e sensíveis como biossensores. Neste trabalho, a matriz de poli(4-aminofenol) foi utilizada para imobilização e detecção de produto de PCR específico para bacilo de Hansen. A modificação da superfície com o filme foi confirmada observando as diferenças eletroquímicas do eletrodo sem modificação frente ao par redox Fe2+/Fe3+ e eletrólito suporte. A caracterização da matriz polimérica foi conduzida por meio de FTIR, espectroscopia de impedância eletroquímica, microbalança eletroquímica de cristal de quartzo e estudos de estabilidade térmica. Dados de FTIR indicam possível polimerização via ligações C-N-C. A estabilidade do filme a 98°C foi avaliada e o mesmo manteve 60% de suas propriedades eletroativas após vinte minutos de tratamento térmico, sendo, portanto, adequado para a metodologia utilizada, neste trabalho, para imobilização de biomoléculas. A detecção de bases nitrogenadas sobre a nova matriz foi possível em valores de pH 4,5 e 7,4. O eletrólito, tampão fosfato em pH 7,4 foi o mais adequado para detecção das bases nitrogenadas e por sua vez, para produtos de PCR. Este eletrólito foi utilizado para imobilização do produto de PCR desnaturado (RLEP3) contendo 78pb e específico para o bacilo de Hansen. A imobilização deste fragmento de DNA foi confirmada monitorando os picos de oxidação da guanosina monofosfato e adenosina monofosfato e também por meio de estudos de espectroscopia de impedância eletroquímica. Indicadores de hibridização, ferrocenocarboxaldeído e azul de metileno, foram utilizados para diferenciar alvos complementares ou não complementares. Com a ocorrência da hibridação, foi observada redução nos valores de sinal de corrente, para ambos indicadores, monitorando-se o sinal do indicador. O menor limite de detecção, 10-10 mol.L-1, do alvo complementar, foi atingido utilizando azul de metileno como indicador. Quando a sonda foi submetida ao alvo não complementar, não foi observada mudança nos valores de corrente da sonda imobilizada. Após trinta dias de preparação, o biossensor manteve 60% de suas propriedades eletroquímicas Estes estudos demonstraram a viabilidade de construção de um biossensor eletroquímico utilizando indicadores eletroativos de baixo custo, que pode ser usado em amostras reais com DNA do bacilo de Hansen. Pela primeira vez, foi demonstrada a produção de eletrodos modificados com poli(4-aminofenol) como matriz para imobilização e detecção de produto de PCR específico para detecção da hanseníase.Fundação de Amparo a Pesquisa do Estado de Minas GeraisMestre em Químicaapplication/pdfporUniversidade Federal de UberlândiaPrograma de Pós-graduação em QuímicaUFUBRCiências Exatas e da TerraBiossensoresHanseníaseCNPQ::CIENCIAS EXATAS E DA TERRA::QUIMICADesenvolvimento de sensor bioletroquímico para detecção da hanseníaseinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisMadurro, Ana Graci Britohttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4709037T3Oliveira, Carlos Alberto Oliveira dehttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4721729H9Naal, Zekihttp://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4780103H9http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4735748J6Afonso, André Santiagoinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UFUinstname:Universidade Federal de Uberlândia (UFU)instacron:UFUTHUMBNAILAndre.pdf.jpgAndre.pdf.jpgGenerated Thumbnailimage/jpeg1338https://repositorio.ufu.br/bitstream/123456789/17293/3/Andre.pdf.jpgcf6f3641276c337ec5b6b96bbf2658e7MD53ORIGINALAndre.pdfapplication/pdf3511549https://repositorio.ufu.br/bitstream/123456789/17293/1/Andre.pdfe35b6e7c0095e90a8472126f677ac717MD51TEXTAndre.pdf.txtAndre.pdf.txtExtracted texttext/plain159830https://repositorio.ufu.br/bitstream/123456789/17293/2/Andre.pdf.txt4ea6ad43df18b40de00d4b691f20f17bMD52123456789/172932016-06-23 04:46:42.034oai:repositorio.ufu.br:123456789/17293Repositório InstitucionalONGhttp://repositorio.ufu.br/oai/requestdiinf@dirbi.ufu.bropendoar:2016-06-23T07:46:42Repositório Institucional da UFU - Universidade Federal de Uberlândia (UFU)false |
dc.title.por.fl_str_mv |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
title |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
spellingShingle |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase Afonso, André Santiago Biossensores Hanseníase CNPQ::CIENCIAS EXATAS E DA TERRA::QUIMICA |
title_short |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
title_full |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
title_fullStr |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
title_full_unstemmed |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
title_sort |
Desenvolvimento de sensor bioletroquímico para detecção da hanseníase |
author |
Afonso, André Santiago |
author_facet |
Afonso, André Santiago |
author_role |
author |
dc.contributor.advisor1.fl_str_mv |
Madurro, Ana Graci Brito |
dc.contributor.advisor1Lattes.fl_str_mv |
http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4709037T3 |
dc.contributor.referee1.fl_str_mv |
Oliveira, Carlos Alberto Oliveira de |
dc.contributor.referee1Lattes.fl_str_mv |
http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4721729H9 |
dc.contributor.referee2.fl_str_mv |
Naal, Zeki |
dc.contributor.referee2Lattes.fl_str_mv |
http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4780103H9 |
dc.contributor.authorLattes.fl_str_mv |
http://buscatextual.cnpq.br/buscatextual/visualizacv.do?id=K4735748J6 |
dc.contributor.author.fl_str_mv |
Afonso, André Santiago |
contributor_str_mv |
Madurro, Ana Graci Brito Oliveira, Carlos Alberto Oliveira de Naal, Zeki |
dc.subject.por.fl_str_mv |
Biossensores Hanseníase |
topic |
Biossensores Hanseníase CNPQ::CIENCIAS EXATAS E DA TERRA::QUIMICA |
dc.subject.cnpq.fl_str_mv |
CNPQ::CIENCIAS EXATAS E DA TERRA::QUIMICA |
description |
Leprosy is a chronic, granulomatous and dermatological disease, from infection caused by the bacillus Mycobacterium leprae. It is object of intervention of the state, because when untreated cause disability in the economically active population. Brazil remains one of the four endemic countries responsible for 34% of all cases registered at the beginning of 2007. The disease diagnosis can only be done after the onset of disease symptoms. Prophylactic measures are possible when using techniques such as ELISA and PCR, which can be performed only in specialized laboratories, not considering the needs of the endemic countries. Such problems can be circumvented by using tools of low cost, simple and sensitive handling as biosensors. In this work, poly(4-aminophenol) matrix, was used for immobilization and detection of specific PCR product for Hansen bacillus. The surface modification with the film was confirmed observing the electrochemical differences of the electrode without modification front at redox pair Fe2+/Fe3+ and supported electrolyte. The characterization of the polymeric matrix was carried out by FTIR, electrochemical impedance spectroscopy, electrochemical quartz crystal microbalance and studies of thermal stability. The FTIR data indicate polymerization possible by C-N-C bonds. The stability of the film at 98 ° C was measured and even showed 60% of its electroative properties after twenty minutes of heat treatment. being, therefore, adequate for the used methodology, in this work, for biomolecules immobilization. The detection of nitrogenated bases on the new matrix was possible in pH 4.5 and 7.4. The eletrolite, phosphate buffer in pH 7.4, was the most adequate to nitrogenated bases detection and in turn, for PCR products. This electrolyte was used for immobilization of the denaturated PCR product (RLEP3), contained 78pb, and specific for the Hansen bacillus. The DNA fragment immobilization was confirmed monitoring the oxidation peaks of guanosine monophosphate and adenosine monophosphate and also by means of electrochemical impedance spectroscopy studies. Hybridization indicator, ferrocenecarboxyaldehyde and methylene blue, had been used to differentiate complementary or not complementary target. With the hibridization occurrence, a reduction in the current signal values was observed, for both indicators, monitoring the indicator. The small detection limit, 10-10 mol.L-1, of the complementary target, was reached using methylene blue as indicator. When the probe was submitted to not complementary target, it was not observed change in the current values of immobilized probe. At the end of thirty days the biosensor kept 60% of their electrochemical properties These studies demonstrated the viability of constructing of an electrochemical biosensor using eletroactive indicators of low-cost, which can be used in real samples with the DNA of Hansen bacillus. For the first time, in the open literature, it was demonstred the modified electrodes production with poly(4-aminophenol) as matrix to immobilization and detection of specific PCRproduct to leprosy detection. |
publishDate |
2008 |
dc.date.issued.fl_str_mv |
2008-02-22 |
dc.date.available.fl_str_mv |
2014-02-19 2016-06-22T18:48:21Z |
dc.date.accessioned.fl_str_mv |
2016-06-22T18:48:21Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
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info:eu-repo/semantics/masterThesis |
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masterThesis |
status_str |
publishedVersion |
dc.identifier.citation.fl_str_mv |
AFONSO, André Santiago. Desenvolvimento de sensor bioletroquímico para detecção da hanseníase. 2008. 105 f. Dissertação (Mestrado em Ciências Exatas e da Terra) - Universidade Federal de Uberlândia, Uberlândia, 2008. |
dc.identifier.uri.fl_str_mv |
https://repositorio.ufu.br/handle/123456789/17293 |
identifier_str_mv |
AFONSO, André Santiago. Desenvolvimento de sensor bioletroquímico para detecção da hanseníase. 2008. 105 f. Dissertação (Mestrado em Ciências Exatas e da Terra) - Universidade Federal de Uberlândia, Uberlândia, 2008. |
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Universidade Federal de Uberlândia |
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Programa de Pós-graduação em Química |
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UFU |
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BR |
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Ciências Exatas e da Terra |
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Universidade Federal de Uberlândia |
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