A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes
Autor(a) principal: | |
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Data de Publicação: | 2010 |
Outros Autores: | , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Revista do Instituto de Medicina Tropical de São Paulo |
Texto Completo: | https://www.revistas.usp.br/rimtsp/article/view/31317 |
Resumo: | Hepatitis B virus (HBV) is a major cause of chronic liver disease worldwide. Besides genotype, quantitative analysis of HBV infection is extensively used for monitoring disease progression and treatment. Affordable viral load monitoring is desirable in resource-limited settings and it has been already shown to be useful in developing countries for other viruses such as Hepatitis C virus (HCV) and HIV. In this paper, we describe the validation of a real-time PCR assay for HBV DNA quantification with TaqMan chemistry and MGB probes. Primers and probes were designed using an alignment of sequences from all HBV genotypes in order to equally amplify all of them. The assay is internally controlled and was standardized with an international HBV panel. Its efficacy was evaluated comparing the results with two other methods: Versant HBV DNA Assay 3.0 (bDNA, Siemens, NY, USA) and another real-time PCR from a reference laboratory. Intra-assay and inter-assay reproducibilities were determined and the mean of CV values obtained were 0.12 and 0.09, respectively. The assay was validated with a broad dynamic range and is efficient for amplifying all HBV genotypes, providing a good option to quantify HBV DNA as a routine procedure, with a cheap and reliable protocol. |
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Revista do Instituto de Medicina Tropical de São Paulo |
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A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes Ensaio quantitativo em tempo real para o DNA do vírus da hepatite B (HBV) desenvolvido para detectar todos os genótipos do HBV HBV DNA quantificationReal Time PCRHepatitis B virusViral load Hepatitis B virus (HBV) is a major cause of chronic liver disease worldwide. Besides genotype, quantitative analysis of HBV infection is extensively used for monitoring disease progression and treatment. Affordable viral load monitoring is desirable in resource-limited settings and it has been already shown to be useful in developing countries for other viruses such as Hepatitis C virus (HCV) and HIV. In this paper, we describe the validation of a real-time PCR assay for HBV DNA quantification with TaqMan chemistry and MGB probes. Primers and probes were designed using an alignment of sequences from all HBV genotypes in order to equally amplify all of them. The assay is internally controlled and was standardized with an international HBV panel. Its efficacy was evaluated comparing the results with two other methods: Versant HBV DNA Assay 3.0 (bDNA, Siemens, NY, USA) and another real-time PCR from a reference laboratory. Intra-assay and inter-assay reproducibilities were determined and the mean of CV values obtained were 0.12 and 0.09, respectively. The assay was validated with a broad dynamic range and is efficient for amplifying all HBV genotypes, providing a good option to quantify HBV DNA as a routine procedure, with a cheap and reliable protocol. O vírus da Hepatite B (HBV) é uma das principais causas de doença crônica do fígado no mundo. Além do genótipo, a análise quantitativa do HBV é amplamente utilizada para monitorar a progressão da doença e o tratamento. Em locais com recursos escassos, métodos baratos para o monitoramento da carga viral são desejáveis e, em países em desenvolvimento, sua utilidade já foi demonstrada para outros vírus, como o da Hepatite C e HIV. Neste trabalho, descrevemos a validação de um teste de PCR em Tempo Real para a quantificação do DNA do HBV utilizando sondas Taqman/MGB. Os oligos e sondas foram escolhidos usando um alinhamento contendo seqüências de todos os genótipos do HBV para garantir uma amplificação igual de todos eles. O teste possui um controle interno e foi padronizado com um painel internacional de HBV. Sua eficácia foi testada comparando-se os resultados com outros dois métodos: Versant HBV DNA Assay 3.0 (bDNA, Siemens, NY, USA) e outro PCR em tempo real realizado em um laboratório de referência. As reprodutibilidades intra e inter-ensaio foram determinadas e a média dos valores de CV obtidos foram de 0,12 e 0,09, respectivamente. O teste foi validado com uma ampla faixa dinâmica e amplificou com eficiência os diferentes genótipos de HBV, fornecendo uma boa opção para a quantificação de rotina do DNA do HBV, com um protocolo barato e confiável. Universidade de São Paulo. Instituto de Medicina Tropical de São Paulo2010-06-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionapplication/pdfhttps://www.revistas.usp.br/rimtsp/article/view/31317Revista do Instituto de Medicina Tropical de São Paulo; Vol. 52 No. 3 (2010); 119-124 Revista do Instituto de Medicina Tropical de São Paulo; Vol. 52 Núm. 3 (2010); 119-124 Revista do Instituto de Medicina Tropical de São Paulo; v. 52 n. 3 (2010); 119-124 1678-99460036-4665reponame:Revista do Instituto de Medicina Tropical de São Pauloinstname:Instituto de Medicina Tropical (IMT)instacron:IMTenghttps://www.revistas.usp.br/rimtsp/article/view/31317/33202Copyright (c) 2018 Revista do Instituto de Medicina Tropical de São Pauloinfo:eu-repo/semantics/openAccessSitnik, RobertaPaes, ÂngelaMangueira, Cristovão PitangueiraPinho, João Renato Rebello2012-07-07T19:30:54Zoai:revistas.usp.br:article/31317Revistahttp://www.revistas.usp.br/rimtsp/indexPUBhttps://www.revistas.usp.br/rimtsp/oai||revimtsp@usp.br1678-99460036-4665opendoar:2022-12-13T16:51:59.623478Revista do Instituto de Medicina Tropical de São Paulo - Instituto de Medicina Tropical (IMT)true |
dc.title.none.fl_str_mv |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes Ensaio quantitativo em tempo real para o DNA do vírus da hepatite B (HBV) desenvolvido para detectar todos os genótipos do HBV |
title |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes |
spellingShingle |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes Sitnik, Roberta HBV DNA quantification Real Time PCR Hepatitis B virus Viral load |
title_short |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes |
title_full |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes |
title_fullStr |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes |
title_full_unstemmed |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes |
title_sort |
A real-time quantitative assay for hepatitis B DNA virus (HBV) developed to detect all HBV genotypes |
author |
Sitnik, Roberta |
author_facet |
Sitnik, Roberta Paes, Ângela Mangueira, Cristovão Pitangueira Pinho, João Renato Rebello |
author_role |
author |
author2 |
Paes, Ângela Mangueira, Cristovão Pitangueira Pinho, João Renato Rebello |
author2_role |
author author author |
dc.contributor.author.fl_str_mv |
Sitnik, Roberta Paes, Ângela Mangueira, Cristovão Pitangueira Pinho, João Renato Rebello |
dc.subject.por.fl_str_mv |
HBV DNA quantification Real Time PCR Hepatitis B virus Viral load |
topic |
HBV DNA quantification Real Time PCR Hepatitis B virus Viral load |
description |
Hepatitis B virus (HBV) is a major cause of chronic liver disease worldwide. Besides genotype, quantitative analysis of HBV infection is extensively used for monitoring disease progression and treatment. Affordable viral load monitoring is desirable in resource-limited settings and it has been already shown to be useful in developing countries for other viruses such as Hepatitis C virus (HCV) and HIV. In this paper, we describe the validation of a real-time PCR assay for HBV DNA quantification with TaqMan chemistry and MGB probes. Primers and probes were designed using an alignment of sequences from all HBV genotypes in order to equally amplify all of them. The assay is internally controlled and was standardized with an international HBV panel. Its efficacy was evaluated comparing the results with two other methods: Versant HBV DNA Assay 3.0 (bDNA, Siemens, NY, USA) and another real-time PCR from a reference laboratory. Intra-assay and inter-assay reproducibilities were determined and the mean of CV values obtained were 0.12 and 0.09, respectively. The assay was validated with a broad dynamic range and is efficient for amplifying all HBV genotypes, providing a good option to quantify HBV DNA as a routine procedure, with a cheap and reliable protocol. |
publishDate |
2010 |
dc.date.none.fl_str_mv |
2010-06-01 |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
https://www.revistas.usp.br/rimtsp/article/view/31317 |
url |
https://www.revistas.usp.br/rimtsp/article/view/31317 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
https://www.revistas.usp.br/rimtsp/article/view/31317/33202 |
dc.rights.driver.fl_str_mv |
Copyright (c) 2018 Revista do Instituto de Medicina Tropical de São Paulo info:eu-repo/semantics/openAccess |
rights_invalid_str_mv |
Copyright (c) 2018 Revista do Instituto de Medicina Tropical de São Paulo |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Universidade de São Paulo. Instituto de Medicina Tropical de São Paulo |
publisher.none.fl_str_mv |
Universidade de São Paulo. Instituto de Medicina Tropical de São Paulo |
dc.source.none.fl_str_mv |
Revista do Instituto de Medicina Tropical de São Paulo; Vol. 52 No. 3 (2010); 119-124 Revista do Instituto de Medicina Tropical de São Paulo; Vol. 52 Núm. 3 (2010); 119-124 Revista do Instituto de Medicina Tropical de São Paulo; v. 52 n. 3 (2010); 119-124 1678-9946 0036-4665 reponame:Revista do Instituto de Medicina Tropical de São Paulo instname:Instituto de Medicina Tropical (IMT) instacron:IMT |
instname_str |
Instituto de Medicina Tropical (IMT) |
instacron_str |
IMT |
institution |
IMT |
reponame_str |
Revista do Instituto de Medicina Tropical de São Paulo |
collection |
Revista do Instituto de Medicina Tropical de São Paulo |
repository.name.fl_str_mv |
Revista do Instituto de Medicina Tropical de São Paulo - Instituto de Medicina Tropical (IMT) |
repository.mail.fl_str_mv |
||revimtsp@usp.br |
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1798951647429263360 |