Genomic CpG methylation profiles of peripheral blood mononuclear cells in childhood AD patients and control subjects

Peripheral blood samples were taken from a total of 48 individuals (24 AD patients and 24 controls) and subjected to access DNA CpG methylation state using Infinium HumanMethylation450 BeadChips. This array consists of probes to determine the site-specific methylation status of more than 450K CpG loci across the human genome. Approximately 500 ng of bisulfate-converted DNA (BCD) was used per BeadChip for hybridization in accordance with the protocol and instructions of Infinium HD methylation. After completing the whole-genome amplification step and then controlled enzymatic end-point fragmentation, BCD samples were precipitated and re-suspended for hybridization for 16 h at 48C. During this process, the DNA molecules are annealed to two bead types with methylated loci (C)-specific and unmethylated loci (T)-specific oligomers. Afterward, the non-hybridized or mis-hybridized DNA was washed away, followed by single-base extension using 2,4-dinitrophrnol- and biotin-labeled ddNTPs and the annealed BCD as templates. Bisulfite converted DNA from 48 PBMC samples (with 24 control subjects and 24 AD patients) was hybridized to the Illumina Infinium HumanMethylation450 BeadChip array.