tetano
Editor, Senior Moderator
Anal Chem
. 2020 May 22.
doi: 10.1021/acs.analchem.0c01884. Online ahead of print.
A Novel One-Step Single-Tube Nested Quantitative Real-Time PCR Assay for Highly Sensitive Detection of SARS-CoV-2
Ji Wang, Kun Cai, Ruiqing Zhang, Xiaozhou He, Xinxin Shen, Jun Liu, Junqiang Xu, Feng Qiu, Wenwen Lei, Jinrong Wang, Xinna Li, Yuan Gao, Yongzhong Jiang, Wenbo Xu, Xuejun Ma
Abstract
Coronavirus disease 2019 (COVID-19) has become a public health emergency. The reverse transcriptase real-time quantitative PCR (qRT-PCR) test is currently considered as a gold standard in the laboratory for the etiological detection of COVID-19. However, qRT-PCR results could be false-negative due to the inadequate sensitivity of qRT-PCR. In this study, we have developed and evaluated a novel one-step single-tube nested quantitative Real-Time PCR (OSN-qRT-PCR) assay for highly sensitive detection of SARS-CoV-2 targeting the ORF1aband N genes. The sensitivity of the OSN-qRT-PCR assay was 1 copy/reaction and 10-fold higher than that of commercial qRT-PCR kit (10 copies/reaction). The clinical performance of the OSN-qRT-PCR assay was evaluated using 181 clinical samples. Among them, 14 qRT-PCR-negative samples (7 had no repetitive results, and 7 had no Ct values) were detected by OSN-qRT-PCR. Moreover, the 7qRT-PCR-positives in the qRT-PCR grey zone (Ct value of ORF1ab ranged from 37.48 to 39.07, and Ct value of N ranged from 37.34 to 38.75) were out of the grey zone thus were deemed to be positive by OSN-qRT-PCR, indicating the positivity of these samples is confirmative. Compared to the qRT-PCR kit, OSN-qRT-PCR assay revealed higher sensitivity and specificity hence better suited to clinical applications for the detection of SARS-CoV-2 in patients with low viral load.
. 2020 May 22.
doi: 10.1021/acs.analchem.0c01884. Online ahead of print.
A Novel One-Step Single-Tube Nested Quantitative Real-Time PCR Assay for Highly Sensitive Detection of SARS-CoV-2
Ji Wang, Kun Cai, Ruiqing Zhang, Xiaozhou He, Xinxin Shen, Jun Liu, Junqiang Xu, Feng Qiu, Wenwen Lei, Jinrong Wang, Xinna Li, Yuan Gao, Yongzhong Jiang, Wenbo Xu, Xuejun Ma
- PMID: 32438806
- DOI: 10.1021/acs.analchem.0c01884
Abstract
Coronavirus disease 2019 (COVID-19) has become a public health emergency. The reverse transcriptase real-time quantitative PCR (qRT-PCR) test is currently considered as a gold standard in the laboratory for the etiological detection of COVID-19. However, qRT-PCR results could be false-negative due to the inadequate sensitivity of qRT-PCR. In this study, we have developed and evaluated a novel one-step single-tube nested quantitative Real-Time PCR (OSN-qRT-PCR) assay for highly sensitive detection of SARS-CoV-2 targeting the ORF1aband N genes. The sensitivity of the OSN-qRT-PCR assay was 1 copy/reaction and 10-fold higher than that of commercial qRT-PCR kit (10 copies/reaction). The clinical performance of the OSN-qRT-PCR assay was evaluated using 181 clinical samples. Among them, 14 qRT-PCR-negative samples (7 had no repetitive results, and 7 had no Ct values) were detected by OSN-qRT-PCR. Moreover, the 7qRT-PCR-positives in the qRT-PCR grey zone (Ct value of ORF1ab ranged from 37.48 to 39.07, and Ct value of N ranged from 37.34 to 38.75) were out of the grey zone thus were deemed to be positive by OSN-qRT-PCR, indicating the positivity of these samples is confirmative. Compared to the qRT-PCR kit, OSN-qRT-PCR assay revealed higher sensitivity and specificity hence better suited to clinical applications for the detection of SARS-CoV-2 in patients with low viral load.