Application Guide - QIAxcel Advanced

QIAxcel Advanced Application Guide 10/2016 67 Test A was set up for simultaneous detection of nine respiratory viruses: The QIAGEN® OneStep RT-PCR Kit was used for the amplification and PCR was performed as previously described (3). All PCR tubes (or 96-well reaction plate) can be loaded directly onto the QIAxcel Advanced electrophoresis system without post-PCR operation. For interpretation during analysis, the QIAxcel DNA High Resolution Kit in combination with the QX DNA Size Marker 25-500 bp and the QX Alignment Marker 15 bp/600 bp were used. In Test B, seven respiratory viruses were detected: • Parainfluenza virus type 2 (PIV2) • Parainfluenza virus type 3 (PIV3) • Respiratory syncytial virus A (RSVA) • Respiratory syncytial virus B (RSVB) • Coronavirus subtypes NL63 (CoV NL63) • Human metapneumovirus (HMPV) • Human bocavirus (HBoV) • Influenza A virus (FluA) • Influenza B virus (FluB) • Seasonal influenza A virus subtypes H1N1 (sH1N1) • Parainfluenza virus type 1 (PIV1) • Human rhinovirus (HRV) • Coronavirus subtypes OC43 (CoV OC43) • Coronavirus subtypes 229E (CoV 229E) • Coronavirus subtypes HKU1 (CoV HKU1) • Adenovirus (Adv) Results and Discussion Test A allowed identification of nine respiratory viral targets (Figure 1A), while test B enabled identification of another seven respiratory viral targets (Figure 1B). The results show discrete band patterns for each of the virus type/subtype-specific amplicons. The QIAxcel DNA High Resolution Kit is capable of resolving amplicons that differ by as few as 5 bp. It is estimated that, with this assay, 100 samples can be tested within 1 working day. Of that, a maximum of 1 hour is hands-on time. This estimate is based on 30 minutes to prepare the PCR mixture, 3 hours to complete the RT-PCR process, and time for detection using the QIAxcel Advanced System (15 minutes for 12 tests). The fast and high resolving capacity provided by this experimental setup increases the potential to assess these 16 respiratory virus types/subtypes in future routine surveillance analyses. Conclusions • The two-tube assay in combination with the QIAxcel Advanced System and the QIAxcel DNA High Resolution Kit provided a reliable method for respiratory virus identification. • The assay had sensitivity and efficiency suitable for consideration for routine surveillance.

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