DNA methylation plays a critical role in the regulation of gene expression and is known to be an essential mechanism for guiding normal cellular development and maintaining tissue identities. Numerous studies have implicated aberrant methylation in the etiology of many human diseases including cancer. Illumina’s Infinium Methylation Assay provides quantitative methylation measurement at the single-CpG-site level, offering the highest resolution for understanding epigenetic changes.
The HumanMethylation450 BeadChip array offers a unique combination of comprehensive, expert-selected coverage, including 99% of RefSeq genes, 96% of CpG islands, and other content categories selected by methylation experts, making it ideal for epigenome-wide association studies (EWAS).
The Infinium Methylation Assay detects cytosine methylation at CpG islands based on highly multiplexed genotyping of bisulfite-converted genomic DNA (gDNA). Upon treatment with bisulfite, unmethylated cytosine bases are converted to uracil, while methylated cytosine bases remain unchanged.
The assay interrogates these chemically differentiated loci using two site-specific probes, one designed for the methylated locus (M bead type) and another for the unmethylated locus (U bead type).
Single-base extension of the probes incorporates a labeled ddNTP, which is subsequently stained with a fluorescence reagent. The level of methylation for the interrogated locus can be determined by calculating the ratio of the fluorescent signals from the methylated vs unmethylated sites.
The Infinium Methylation Assay uses two different bead types to detect CpG methylation. The U bead type matches the unmethylated CpG site; the M bead type matches the methylated site. In the top figure, the unmethylated CpG target site matches with the U probe, enabling single-base extension and detection. It has a single-base mismatch to the M probe, which inhibits extension. If the CpG locus of interest is methylated (bottom figure), the reverse occurs.