Determination of the CpG Island Methylator Phenotype (CIMP) in colorectal cancer using MethyLight

Abstract

MethyLight is a sensitive, quantitative, TaqMan-based, real-time PCR assay for measuring DNA methylation profiles. The MethyLight-based data is presented as a percentage relative to an M.SssI-treated methylated DNA reference sample (PMR). We have extended the applicability of MethyLight technology to assay for the presence of the CpG island methylator phenotype (CIMP) in human colorectal cancer. We provide here a detailed, step-by-step protocol of the MethyLight assay for detection of CIMP with high sensitivity and specificity in colorectal cancer using a five marker panel composed of CACNA1G, IGF2, NEUROG1, RUNX3 and SOCS1. Our criteria state that a sample with ≥3 of five markers posititve for methylation (PMR>10) is CIMP positive (CIMP+), while a sample with ≤2 of the five markers positive for methylation is considered CIMP negative (CIMP-).