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Southeast Asian J Trop Med Public Health ; 2004 Sep; 35(3): 623-9
Artigo em Inglês | IMSEAR | ID: sea-34043

RESUMO

The aim of this study was to develop a rapid, sensitive and robust procedure for the qualitative detection of SARS coronavirus RNA. Three unique detection formats were developed for real-time RNA amplification assays: a post amplification detection step with a virus-specific internal capture probe based on Taqman (RT-PCR TaqMan assay), hybridization probe (RT-PCR hybridization probe assay) and a real-time assay with virus-specific molecular beacon probes (NASBA-Beacon assay). The analytical sensitivity or reproducibility of the test results among those three assays was compared. All assays yielded results by detecting SARS coronavirus targeting the BNI-1 region in less than 2 hours. RNA detection by all the formats was unaffected by the presence of human sputum. The limits of detection were at least 10 copies of input RNA for both RT-PCR formats (RT-PCR TaqMan and RT-PCR hybridization probe assays), while the NASBA-Beacon assay could detect as little as 1 copy per reaction, with high reproducibility of the coefficient of variation (CV) of <10. These results demonstrate that real-time NASBA provides a rapid and sensitive alternative to RT-PCR for the routine qualitative assay of sputum for SARS corona viral RNA detection.


Assuntos
Sistemas Computacionais , Sondas de DNA/diagnóstico , Humanos , Técnicas de Sonda Molecular , Técnicas de Amplificação de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase , Coronavírus Relacionado à Síndrome Respiratória Aguda Grave/isolamento & purificação , Sensibilidade e Especificidade , Síndrome Respiratória Aguda Grave/diagnóstico , Software , Taq Polimerase/diagnóstico
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