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Chantratita, W. et al.

Chantratita, Pongtanapisit, Piroj, Srichunrasmi, Seesuai,

Development and comparison of the real-time amplification based methods--NASBA-Beacon, RT-PCR taqman and RT-PCR hybridization probe assays--for the qualitative detection of sars coronavirus.

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.[1]

References

Development and comparison of the real-time amplification based methods--NASBA-Beacon, RT-PCR taqman and RT-PCR hybridization probe assays--for the qualitative detection of sars coronavirus. Chantratita, W., Pongtanapisit, W., Piroj, W., Srichunrasmi, C., Seesuai, S. Southeast Asian J. Trop. Med. Public Health (2004) [Pubmed]

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