Lastly, the question of how to measure epigenetic modification remains a challenge for investigators in this area. There are many laboratory protocols for measuring epigenetic change. In the case of DNA methylation alone, for example, there are several available assays. These include methylated DNA immunoprecipitation (MeDIP);[53] bisulfite reaction based DNA sequencing methods, such as methylation specific PCR and/or bisulfite genomic sequencing PCR;[54,55] Restriction Landmark Genomic Scanning for Methylation (RLGS-M) techniques;[56] and genome-wide screens, such as CpG island microarrays.[57] More recently, the Reduced Representation Bisulfite Sequencing (RRBS) method and the Illumina HumanMethylation450 beadchip (Illumina 450K) have come into more common use. While the sensitivity and specificity of these approaches relative to one another, to the best of our knowledge, remains unmeasured, one recent study pursued a head-to-head comparison of two very similar bisulfite sequence-based assays in human embryonic stem cell replicates and found a nearly 20% difference in methylated CpG islands identified.[58] Despite the more common use of the RRBS and Illumina 450K protocols, there remains no gold standard assay. In this way, differential use of assays may lead to non-differential misclassification bias, ultimately increasing the chances of type II error in this literature.
