This unit describes a PCR-based assay for distinguishing between the two X chromosomes in female cells and assessing the percentage of cells having each parental X chromosome active. Methylation of CpG residues in gene promoters is a major mechanism of transcriptional silencing. In mammalian female cells, hypermethylation is the way in which one X chromosome is inactivated. The X-inactivation assay described in the Basic Protocol relies on methylation sensitivity. In this unit, the highly polymorphic and therefore typically heterozygous (CAG)n region of the 5 end of the coding region of the human androgen receptor gene (HUMARA), at Xq11.2, is used to distinguish and compare the methylation activity of the X chromosomes.