Figure 1.

Illumina Infinium HumanMethylation450 assay. (a) Infinium I assay. Each individual CpG is interrogated using two bead types: methylated (M) and unmethylated (U). The probe design assumes that all CpGs underlying the probe body have the same methylation status as the target CpG. Both bead types will incorporate the same labeled nucleotide for the same target CpG, thereby producing the same color fluorescence. The nucleotide that is added is determined by the base downstream of the 'C' of the target CpG. The proportion of methylation, β, can be calculated by comparing the intensities from the two different probes in the same color: β= M/(U + M). (b) Infinium II assay. Each target CpG is interrogated using a single bead type. A probe may have up to three underlying CpG sites, with a degenerate R base corresponding to the 'C' of each CpG. Methylation state is detected by single base extension at the position of the 'C' of the target CpG, which always results in the addition of a labeled 'G' or 'A' nucleotide, complementary to either the 'methylated' C or 'unmethylated' T, respectively. Each locus is detected in two colors, and methylation status is determined by comparing the two colors from the one position: β = Green (M)/(Red (U) + Green (M)). (c) The number of CpG dinucleotides in the body of the probe according to Infinium probe type. Infinium I probes have significantly more CpGs in the probe body.