We have used qPCR to do this, and also found good correlation with microarray results. qPCR results are of course far more accurate, and therefore preferable as a measure of extent of knockdown. Our assay of choice has been the miRNA (FAM) kits from Applied Biosystems.

We have used this for in vivo transfections as well as with cultured cells.

The question raised is most relevant. We obtained increasing knockdown on injection of up to 50 pmol inhibitor, but observe lower knockdown with higher dosages, e.g. 75 or 100 pmol. I would love to know why!

If you are using the Exiqon LNA you can't confirm the miRNAs knockdown thorough the Real Time PCR, because LNA-miRNA binding not leads to degradation of the miRNA, so with the assay you are detecting also the miRNA that is binding. You should analyze some confirmed target gene.

The inhibitors are synthetic oligonucleotides that contain the sequences of the corresponding miRNAs and they could interfere with the qRT-PCR primers and probes. The levels of transfected miRNA frequently seen using RT-PCR do not represent the functional levels. Therefore, such reactions produce rather misleading results and reflect neither levels nor activity of the modulated miRNAs (example of reference

Thomson et al., 2013 ).