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10.1371/journal.pone.0236564

http://scihub22266oqcxt.onion/10.1371/journal.pone.0236564
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32706827!7380591!32706827
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suck abstract from ncbi


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pmid32706827      PLoS+One 2020 ; 15 (7): e0236564
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  • Detection of SARS-CoV-2 RNA by direct RT-qPCR on nasopharyngeal specimens without extraction of viral RNA #MMPMID32706827
  • Hasan MR; Mirza F; Al-Hail H; Sundararaju S; Xaba T; Iqbal M; Alhussain H; Yassine HM; Perez-Lopez A; Tang P
  • PLoS One 2020[]; 15 (7): e0236564 PMID32706827show ga
  • To circumvent the limited availability of RNA extraction reagents, we aimed to develop a protocol for direct RT-qPCR to detect SARS-CoV-2 in nasopharyngeal swabs without RNA extraction. Nasopharyngeal specimens positive for SARS-CoV-2 and other coronaviruses collected in universal viral transport (UVT) medium were pre-processed by several commercial and laboratory-developed methods and tested by RT-qPCR assays without RNA extraction using different RT-qPCR master mixes. The results were compared to that of standard approach that involves RNA extraction. Incubation of specimens at 65 degrees C for 10 minutes along with the use of TaqPath 1-Step RT-qPCR Master Mix provides higher analytical sensitivity for detection of SARS-CoV-2 RNA than many other conditions tested. The optimized direct RT-qPCR approach demonstrated a limit of detection of 6.6x103 copy/ml and high reproducibility (co-efficient of variation = 1.2%). In 132 nasopharyngeal specimens submitted for SARS-CoV-2 testing, the sensitivity, specificity and accuracy of our optimized approach were 95%, 99% and 98.5%, respectively, with reference to the standard approach. Also, the RT-qPCR CT values obtained by the two methods were positively correlated (Pearson correlation coefficient r = 0.6971, p = 0.0013). The rate of PCR inhibition by the direct approach was 8% compared to 9% by the standard approach. Our simple approach to detect SARS-CoV-2 RNA by direct RT-qPCR may help laboratories continue testing for the virus despite reagent shortages or expand their testing capacity in resource limited settings.
  • |*Real-Time Polymerase Chain Reaction[MESH]
  • |Betacoronavirus[MESH]
  • |COVID-19[MESH]
  • |COVID-19 Testing[MESH]
  • |Clinical Laboratory Techniques/*methods[MESH]
  • |Coronavirus Infections/*diagnosis[MESH]
  • |Humans[MESH]
  • |Nasopharynx/virology[MESH]
  • |Pandemics[MESH]
  • |Pneumonia, Viral/*diagnosis[MESH]
  • |RNA, Viral/*isolation & purification[MESH]
  • |Reproducibility of Results[MESH]
  • |SARS-CoV-2[MESH]
  • |Sensitivity and Specificity[MESH]


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