Evaluating plasma RNA yield using RT-qPCR for miRNAs.

As biofluid RNA is below the detection limit of standard quantification tools, all RT-qPCRs were performed using 1.5 μL RNA extracted from 200 μL plasma. Lines indicate paired samples i.e. aliquots of plasma from the same blood draw. (A) RT-qPCR for the spike-in cel-miR-39-3p using the four kits with and without glycogen. (B-D) RT-qPCR for three endogenous miRNAs isolated using the four kits with and without glycogen. (E) The miRNeasy serum/plasma protocol recommends elution in 14 μL, compared with 100 μL for the mirVana protocol. To control for effects of this, RNA extractions were performed with both protocols, eluting in 30 μL or 50 μL. RT-qPCR for cel-miR-39-3p, hsa-miR-16-5p and hsa-miR-21-5p were performed to evaluate the different elution volumes.