In this chapter we outline a RNA extraction method for very low immune cell populations isolated from the central nervous system of mice undergoing experimental autoimmune encephalomyelitis. We compare various normalization and quantification techniques to examine miRNA expression. Our data highlight that employing a mean normalization procedure using a number of well-selected housekeeping miRNA genes, followed by absolute quantification with a standard curve generated from a commercial miRNA oligo, gave the most robust and reproducible miRNA expression results.
Keywords: Absolute quantification; Experimental autoimmune encephalomyelitis; Immune cells; Mean normalization; RT-PCR; Relative quantification; Standard curve; microRNA.