500 ng of DNA was used for bisulfite conversion using the Zymo Research EZ-96 DNA Methylation Kit. The bisulfite converted DNA extracts were hybridized to Infinium 450K BeadChips (Illumina). Due to the differences in sample plates between the completed Zymo assay and the Illumina assay, pre-amplification was performed manually by following the Illumina MSA4 SOP. Once complete, sample and reagent barcodes were simmed through the Illumina LIMS tracking software. Four microlitres (200 ng) of sample is required (Illumina guidelines) for the pre-amplification reaction using the Tecan Freedom Evo. No further quantification step was performed after the completion of the Zymo assay. Labelling was performed automatically during the post-amplification xStain process with Biotin and DNP labelled antibodies. The HumanMethylation450_15017482_v.1.1 arrays were scanned with iScan (Illumina) in accordance with the manufacturer’s protocol.
