Comprehensive evaluation of an one pot quadruplex RT-qPCR assay for the simultaneously subtyping influenza A virus into H1, H3, N1 and N2

Abstract

In the current study, an one pot quadruplex RT-qPCR assay was established and comprehensively evaluated. The assay's limit of detection could reach as low as 10¹ copies/reaction, with good repeatability profile and no cross-reaction with other respiratory pathogens. During clinical evaluation both by blinded samples and real clinical samples, the assay exhibited a 100% coincidence rate with individual commercial RT-qPCR assays. To the best of our knowledge, this is the first report on the simultaneous subtyping influenza A virus into H1, H3, N1, and N2 by one pot quadruplex RT-qPCR assay, which could improve the preparedness for future influenza outbreaks.