Quantinova probe rt qpcr kit

Serum, blood and urine samples were obtained from patients 0 to 228 days after first symptoms (Extended Data Table 1a). Viral RNA was isolated from 200 µl Zika-suspected samples using either the NucliSENS easyMag system (BioMerieux, Basingstoke, UK) (Ribeirão Preto samples), the ExiPrep Dx Viral RNA Kit (BIONEER, Republic of Korea) (Rio de Janeiro samples) or the QIAamp Viral RNA Mini kit (QIAGEN, Hilden, Germany) (all other samples) according to the manufacturer’s instructions. Ct values were determined for all samples by probe-based RT-qPCR against the prM target (using 5′ FAM as the probe reporter dye) as previously described^{34 (link)}. RT-qPCR assays were performed using the QuantiNova Probe RT-qPCR Kit (20 ul reaction volume; QIAGEN) with amplification in the Rotor-Gene Q (QIAGEN) following the manufacturer’s protocol. Primers/probe were synthesised by Integrated DNA Technologies (Leuven, Belgium). The following reaction conditions were used: reverse transcription (50°C, 10 min), reverse transcriptase inactivation and DNA polymerase activation (95°C, 20 sec), followed by 40 cycles of DNA denaturation (95°C, 10 secs) and annealing-extension (60°C, 40 sec). Positive and negative controls were included in each batch; however, due to the large number of samples tested in a short time it was possible only to run each sample without replication.