Validation experiments using MSP-PCR. MSP-PCR was performed for the PRF1, ADORA3, BCL10, and IGSF4 genes, using bisulfite-treated DNA from IGHV mutated samples (M1-3 to M8-12) and IGHV unmutated samples (UM1-4 and UM8-12). (A) Data for samples analyzed in the methylation array; (B) results from additional CLL samples are included. Analysis of each gene in panels A and B was performed during the same experiment and on the same gel; vertical line(s) have been inserted to indicate a repositioned gel lane. N indicates normal PBMC sample; M, methylation-specific primers; and U, unmethylation-specific primers.