Exogenous internal reference standards

Exogenously added internal reference standards are a pure, unrelated RNA of known concentration and integrity that is added to each sample prior to analysis.  Such RNAs can be bought commercially (for example, Ambion’s RNA250) and can be used to correct the data for the presence of enzyme inhibitors in individual RNA samples (Tentcheva et al., 2006). The amount added per reaction should be low; <1% of the amount of sample RNA, so as not to affect the RT-qPCR reaction efficiencies.