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Volume 27, Number 7—July 2021

Multiplex Real-Time Reverse Transcription PCR for Influenza A Virus, Influenza B Virus, and Severe Acute Respiratory Syndrome Coronavirus 2

Bo Shu1, Marie K. Kirby1, William G. Davis, Christine Warnes, Jimma Liddell, Ji Liu, Kai-Hui Wu, Norman Hassell, Alvaro J. Benitez, Malania M. Wilson, Matthew W. Keller, Benjamin L. Rambo-Martin, Yamundow Camara, Jörn Winter, Rebecca J. Kondor, Bin Zhou, Stacey Spies, Laura E. Rose, Jonas M. Winchell, Brandi M. Limbago, David E. Wentworth2, and John R. Barnes2Comments to Author 
Author affiliations: Centers for Disease Control and Prevention, Atlanta, Georgia, USA (B. Shu, M.K. Kirby, W.G. Davis, C. Warnes, J. Liddell, J. Liu, K.-H. Wu, N. Hassell, A.J. Benitez, M.M. Wilson, M.W. Keller, B.L. Rambo-Martin, Y. Camara, J. Winter, R.J. Kondor, B. Zhou, S. Spies, L.E. Rose, J.M. Winchell, B.M. Limbago, D.E. Wentworth, J.R. Barnes); Battelle Memorial Institute, Atlanta (W.G. Davis, J. Liddell); Leidos Inc, Atlanta (Y. Camara)

Main Article

Table 4

Evaluation of the Influenza SARS-CoV-2 Multiplex Assay sensitivity using quantified synthetic RNAs*

RNA copies/reaction Cycle threshold values
Influenza A
Influenza B
RNase P (human)
50,000 23.72 24.02 23.86 22.02 21.59 21.88 20.05 20.26 20.08 22.02 22.13 21.69
5,000 27.01 27.38 28.01 25.17 25.03 25.04 24.03 24.12 24.15 25.12 24.68 25.09
500 31.74 31.73 32.48 28.52 28.02 28.88 27.25 28.01 27.60 28.19 28.02 28.11
50 35.60 35.34 36.65 32.58 31.58 31.17 31.89 31.33 32.93 31.47 31.54 30.59
5 36.50 38.40 0 33.76 34.65 36.15 34.24 34.03 34.25 34.33 35.42 39.02

*Multiplex real-time reverse transcription PCR for influenza A virus, influenza B virus, and SARS-CoV-2. Influenza A, Influenza B, and RP targets were quantified using Armored RNA Quant CDC-9 (Asuragen, Inc.,, which includes the M gene of influenza A virus strain A/Brisbane/02/2018_H1N1, the NS gene of influenza B virus strain B/Colorado/06/2017, and RP; the SARS-CoV-2 target was quantified using Twist Synthetic SARS-CoV-2 RNA (Twist Bioscience, SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

Main Article

1These first authors contributed equally to this article.

2These senior authors contributed equally to this article.

Page created: May 13, 2021
Page updated: June 16, 2021
Page reviewed: June 16, 2021
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