CD44 and breast cancer: To test whether the TGF-β1 causes the epigenetic modification on GSN expression in breast cancer cells, the method of methylation-specific PCR (MSPCR) for assessing the methylation and unmethylation on the CpG island at the promoter region of GSN (intron 1) was used in MDA-MB231 cells without FACS sorting (control), in CD44+/CD22- subpopulation sorted cells without TGF-β1 pretreatment, and in CD44+/CD22- subpopulation sorted cells with 2 ng/ml TGF-β1 pretreatment for 3 days (Fig. 6a).