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.
Identifieur interne : 002908 ( PubMed/Curation ); précédent : 002907; suivant : 002909Development 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.
Auteurs : Wasun Chantratita [Thaïlande] ; Wiroj Pongtanapisit ; Wantanich Piroj ; Chutatip Srichunrasmi ; Somying SeesuaiSource :
- The Southeast Asian journal of tropical medicine and public health [ 0125-1562 ] ; 2004.
Descripteurs français
- KwdFr :
- Expectoration (virologie), Humains, Logiciel, Réaction de polymérisation en chaîne, Sensibilité et spécificité, Sondes d'ADN, Syndrome respiratoire aigu sévère (diagnostic), Syndrome respiratoire aigu sévère (virologie), Systèmes informatiques, TAQ polymerase, Techniques d'amplification d'acides nucléiques (), Techniques de sonde moléculaire, Virus du SRAS (isolement et purification).
- MESH :
- diagnostic : Syndrome respiratoire aigu sévère.
- isolement et purification : Virus du SRAS.
- virologie : Expectoration, Syndrome respiratoire aigu sévère.
- Humains, Logiciel, Réaction de polymérisation en chaîne, Sensibilité et spécificité, Sondes d'ADN, Systèmes informatiques, TAQ polymerase, Techniques d'amplification d'acides nucléiques, Techniques de sonde moléculaire.
English descriptors
- KwdEn :
- Computer Systems, DNA Probes, Humans, Molecular Probe Techniques, Nucleic Acid Amplification Techniques (methods), Polymerase Chain Reaction, SARS Virus (isolation & purification), Sensitivity and Specificity, Severe Acute Respiratory Syndrome (diagnosis), Severe Acute Respiratory Syndrome (virology), Software, Sputum (virology), Taq Polymerase.
- MESH :
- chemical : DNA Probes, Taq Polymerase.
- diagnosis : Severe Acute Respiratory Syndrome.
- isolation & purification : SARS Virus.
- methods : Nucleic Acid Amplification Techniques.
- virology : Severe Acute Respiratory Syndrome, Sputum.
- Computer Systems, Humans, Molecular Probe Techniques, Polymerase Chain Reaction, Sensitivity and Specificity, Software.
Abstract
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.
PubMed: 15689078
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uniqKey="Piroj W" first="Wantanich" last="Piroj">Wantanich Piroj</name></author><author><name sortKey="Srichunrasmi, Chutatip" sort="Srichunrasmi, Chutatip" uniqKey="Srichunrasmi C" first="Chutatip" last="Srichunrasmi">Chutatip Srichunrasmi</name></author><author><name sortKey="Seesuai, Somying" sort="Seesuai, Somying" uniqKey="Seesuai S" first="Somying" last="Seesuai">Somying Seesuai</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2004">2004</date><idno type="RBID">pubmed:15689078</idno><idno type="pmid">15689078</idno><idno type="wicri:Area/PubMed/Corpus">002908</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">002908</idno><idno type="wicri:Area/PubMed/Curation">002908</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">002908</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">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.</title><author><name sortKey="Chantratita, Wasun" sort="Chantratita, Wasun" uniqKey="Chantratita W" first="Wasun" last="Chantratita">Wasun Chantratita</name><affiliation wicri:level="1"><nlm:affiliation>Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Bangkok, Thailand. rawct@mahidol.ac.th</nlm:affiliation><country xml:lang="fr">Thaïlande</country><wicri:regionArea>Department of Pathology, Faculty of Medicine, Ramathibodi Hospital, Bangkok</wicri:regionArea></affiliation></author><author><name sortKey="Pongtanapisit, Wiroj" sort="Pongtanapisit, Wiroj" uniqKey="Pongtanapisit W" first="Wiroj" last="Pongtanapisit">Wiroj Pongtanapisit</name></author><author><name sortKey="Piroj, Wantanich" sort="Piroj, Wantanich" uniqKey="Piroj W" first="Wantanich" last="Piroj">Wantanich Piroj</name></author><author><name sortKey="Srichunrasmi, Chutatip" sort="Srichunrasmi, Chutatip" uniqKey="Srichunrasmi C" first="Chutatip" last="Srichunrasmi">Chutatip Srichunrasmi</name></author><author><name sortKey="Seesuai, Somying" sort="Seesuai, Somying" uniqKey="Seesuai S" first="Somying" last="Seesuai">Somying Seesuai</name></author></analytic><series><title level="j">The Southeast Asian journal of tropical medicine and public health</title><idno type="ISSN">0125-1562</idno><imprint><date when="2004" type="published">2004</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Computer Systems</term><term>DNA Probes</term><term>Humans</term><term>Molecular Probe Techniques</term><term>Nucleic Acid Amplification Techniques (methods)</term><term>Polymerase Chain Reaction</term><term>SARS Virus (isolation & purification)</term><term>Sensitivity and Specificity</term><term>Severe Acute Respiratory Syndrome (diagnosis)</term><term>Severe Acute Respiratory Syndrome (virology)</term><term>Software</term><term>Sputum (virology)</term><term>Taq Polymerase</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Expectoration (virologie)</term><term>Humains</term><term>Logiciel</term><term>Réaction de polymérisation en chaîne</term><term>Sensibilité et spécificité</term><term>Sondes d'ADN</term><term>Syndrome respiratoire aigu sévère (diagnostic)</term><term>Syndrome respiratoire aigu sévère (virologie)</term><term>Systèmes informatiques</term><term>TAQ polymerase</term><term>Techniques d'amplification d'acides nucléiques ()</term><term>Techniques de sonde moléculaire</term><term>Virus du SRAS (isolement et purification)</term></keywords><keywords scheme="MESH" type="chemical" xml:lang="en"><term>DNA Probes</term><term>Taq Polymerase</term></keywords><keywords scheme="MESH" qualifier="diagnosis" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" qualifier="diagnostic" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="isolation & purification" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Nucleic Acid Amplification Techniques</term></keywords><keywords scheme="MESH" qualifier="virologie" xml:lang="fr"><term>Expectoration</term><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term><term>Sputum</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Computer Systems</term><term>Humans</term><term>Molecular Probe Techniques</term><term>Polymerase Chain Reaction</term><term>Sensitivity and Specificity</term><term>Software</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Humains</term><term>Logiciel</term><term>Réaction de polymérisation en chaîne</term><term>Sensibilité et spécificité</term><term>Sondes d'ADN</term><term>Systèmes informatiques</term><term>TAQ polymerase</term><term>Techniques d'amplification d'acides nucléiques</term><term>Techniques de sonde moléculaire</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">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.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">15689078</PMID><DateCompleted><Year>2005</Year><Month>05</Month><Day>31</Day></DateCompleted><DateRevised><Year>2019</Year><Month>12</Month><Day>10</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0125-1562</ISSN><JournalIssue CitedMedium="Print"><Volume>35</Volume><Issue>3</Issue><PubDate><Year>2004</Year><Month>Sep</Month></PubDate></JournalIssue><Title>The Southeast Asian journal of tropical medicine and public health</Title><ISOAbbreviation>Southeast Asian J. Trop. Med. Public Health</ISOAbbreviation></Journal><ArticleTitle>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.</ArticleTitle><Pagination><MedlinePgn>623-9</MedlinePgn></Pagination><Abstract><AbstractText>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. 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