QIAcuity OneStep Advanced EG Kit (5mL)

The QIAcuity OneStep Advanced EG Kit from QIAGEN is a one-step RT/dPCR master mix intended for use on QIAcuity digital PCR instruments, combining reverse transcription and PCR in a single-tube workflow for quantitative detection of RNA targets.

According to the manufacturer's Quick-Start Protocol, the kit is configured for use with RT² qPCR Primer Assays in singleplex reactions on QIAcuity instruments. It comprises a 4x EvaGreen Master Mix and a 100x OneStep Advanced Reverse Transcription Mix containing a hot-start RT enzyme that allows reactions to be assembled at room temperature. The kit includes QuantiNova Internal Control RNA (QN IC RNA), which can be used optionally as a reverse transcription and amplification control at a 1:1000 final dilution when paired with the separately purchased QuantiTect Primer Assay. The protocol supports Nanoplate 8.5k (24-well and 96-well) and Nanoplate 26k (8-well and 24-well) formats, with total reaction volumes of 12 µL and 40 µL, respectively. Up to four or eight plates may be run in parallel on the QIAcuity Four or QIAcuity Eight instruments.

Reagents must be stored at −30 to −15 °C in a constant-temperature freezer and protected from light. Imaging is performed in the Green channel using an exposure duration of 200 ms and a gain setting of 3. The cycling program comprises 40 min reverse transcription at 50 °C, 2 min RT enzyme inactivation at 95 °C, 40 cycles of denaturation at 95 °C for 10 s followed by combined annealing/extension at 60 °C for 30 s, and a 5 min cooling step at 40 °C.