TY - JOUR
T1 - Performance characteristics of a high-throughput automated transcription-mediated amplification test for SARS-CoV-2 detection
AU - Pham, Jimmykim
AU - Meyer, Sarah
AU - Nguyen, Catherine
AU - Williams, Analee
AU - Hunsicker, Melissa
AU - McHardy, Ian
AU - Gendlina, Inessa
AU - Yitzchak Goldstein, D.
AU - Fox, Amy S.
AU - Hudson, Angela
AU - Darby, Paul
AU - Hovey, Paul
AU - Morales, Jose
AU - Mitchell, James
AU - Harrington, Karen
AU - Majlessi, Mehrdad
AU - Moberly, Joshua
AU - Shah, Ankur
AU - Worlock, Andrew
AU - Walcher, Marion
AU - Eaton, Barbara
AU - Getman, Damon
AU - Clark, Craig
N1 - Publisher Copyright:
Copyright © 2020 Pham et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.
PY - 2020/10
Y1 - 2020/10
N2 - The COVID-19 pandemic caused by the new SARS-CoV-2 coronavirus has imposed severe challenges on laboratories in their effort to achieve sufficient diagnostic testing capability for identifying infected individuals. In this study, we report the analytical and clinical performance characteristics of a new, high-throughput, fully automated nucleic acid amplification test system for the detection of SARS-CoV-2. The assay utilizes target capture, transcription-mediated amplification, and acridinium ester-labeled probe chemistry on the automated Panther system to directly amplify and detect two separate target sequences in the open reading frame 1ab (ORF1ab) region of the SARS-CoV-2 RNA genome. The probit 95% limit of detection of the assay was determined to be 0.004 50% tissue culture infective dose (TCID50)/ml using inactivated virus and 25 copies/ml (c/ml) using synthetic in vitro transcript RNA targets. Analytical sensitivity (100% detection) was confirmed to be 83 to 194 c/ml using three commercially available SARS-CoV-2 nucleic acid controls. No cross-reactivity or interference was observed with testing of six related human coronaviruses, as well as 24 other viral, fungal, and bacterial pathogens, at high titers. Clinical nasopharyngeal swab specimen testing (n = 140) showed 100%, 98.7%, and 99.3% positive, negative, and overall agreement, respectively, with a validated reverse transcription-PCR nucleic acid amplification test (NAAT) for SARS-CoV-2 RNA. These results provide validation evidence for a sensitive and specific method for pandemic-scale automated molecular diagnostic testing for SARS-CoV-2.
AB - The COVID-19 pandemic caused by the new SARS-CoV-2 coronavirus has imposed severe challenges on laboratories in their effort to achieve sufficient diagnostic testing capability for identifying infected individuals. In this study, we report the analytical and clinical performance characteristics of a new, high-throughput, fully automated nucleic acid amplification test system for the detection of SARS-CoV-2. The assay utilizes target capture, transcription-mediated amplification, and acridinium ester-labeled probe chemistry on the automated Panther system to directly amplify and detect two separate target sequences in the open reading frame 1ab (ORF1ab) region of the SARS-CoV-2 RNA genome. The probit 95% limit of detection of the assay was determined to be 0.004 50% tissue culture infective dose (TCID50)/ml using inactivated virus and 25 copies/ml (c/ml) using synthetic in vitro transcript RNA targets. Analytical sensitivity (100% detection) was confirmed to be 83 to 194 c/ml using three commercially available SARS-CoV-2 nucleic acid controls. No cross-reactivity or interference was observed with testing of six related human coronaviruses, as well as 24 other viral, fungal, and bacterial pathogens, at high titers. Clinical nasopharyngeal swab specimen testing (n = 140) showed 100%, 98.7%, and 99.3% positive, negative, and overall agreement, respectively, with a validated reverse transcription-PCR nucleic acid amplification test (NAAT) for SARS-CoV-2 RNA. These results provide validation evidence for a sensitive and specific method for pandemic-scale automated molecular diagnostic testing for SARS-CoV-2.
KW - Aptima
KW - Automation
KW - High throughput
KW - SARS-CoV-2
KW - TMA
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U2 - 10.1128/JCM.01669-20
DO - 10.1128/JCM.01669-20
M3 - Article
C2 - 32727828
AN - SCOPUS:85091576873
SN - 0095-1137
VL - 58
JO - Journal of Clinical Microbiology
JF - Journal of Clinical Microbiology
IS - 10
M1 - e0166920
ER -