Specificity was assessed using 8 pools each containing at least 80 different synthetic miRNAs. Closely related miRNAs are placed in different pools. This example shows results from the hsa-miR-1 assay. The miRCURY LNA miRNA PCR System performs perfectly with a high signal in pool 1, which contains the hsa-miR-1 synthetic miRNA target, and only a borderline signal in pool 7, which contains the most closely related miRNA (hsa-miR-206). The competitor platform uses ligation-based cDNA synthesis and only one miRNA-specific probe. Competitor T shows a lack of specificity. The highest signal for Competitor T is obtained with pool 1. However, non-specific false-positive signals are obtained from all other pools in the absence of hsa-miR-1 template, within a Cq range that would be interpreted as real signals.
Aliquots of the same heart and liver total RNA were expression profiled using miRCURY LNA miRNA PCR Panels in two different locations (on separate days by different operators) using different Roche LightCycler 480 Real-Time PCR instruments. The correlation between average Cq values (triplicate RT reactions) from all miRNAs with signals below 35 Cq values (total of 297 datapoints) is shown.
Raw Cq values from two separate RT reactions (RT1 and RT2) on total RNA purified from 65 µl serum are shown. A total of 730 miRNAs were profiled. Only miRNAs with Cq values below 35 have been included (133 data points).
Different RT reactions using 40 ng heart and liver total RNA were profiled on the miRCURY LNA miRNome Human PCR Panel I and II on different days. The correlation between raw Cq values from all miRNAs with signals below 35 Cq values is shown (total of 297 data points).