Identification of the most stably expressed reference genes using GeNorm analysis. RNA samples from whole-blood from 20 MS patients and 20 matched controls were analyzed using qRT-PCR. A. Average expression stability values (M) of the reference genes during step-wise exclusion of the least stable control gene. The x- axis shows the gene with the highest M value (the lowest stability) for this set of genes. The least stable gene was excluded after each iteration. B. Determination ofthe optimal number of reference genes for normalization. V is the pair-wise variation of two sequential normalization factors. The least number of genes for each V<0.15 was selected as the optimal set of genes for normalization . In this study, the four most stably expressed genes (TRAP1, FPGS, DECR1 and PPIB) represent the optimal set of genes for normalization.