MethylLight is a high-throughput assay for measuring DNA methylation using fluorescence-based real-time PCR (TaqMan®). This method allows for the detection of methylated alleles in the presence of a 10,000-fold excess of unmethylated alleles and provides highly quantitative results. The assay is designed to be sensitive, specific, and reproducible, and it can be performed with very small amounts of DNA, making it suitable for use with microdissected paraffin-embedded tissue samples. MethylLight distinguishes between mono- and bi-allelic methylation of the MLH1 gene in human colorectal tumor specimens. The technique is compatible with multiple gene loci and allows for rapid analysis of many samples. The study validates the MethylLight technology by comparing it with conventional methods such as COBRA and shows that it can accurately detect methylation status. The assay is also shown to be highly sensitive, capable of detecting methylation in the presence of a large amount of unmethylated DNA. The MethylLight technology is also shown to be reproducible on heterogeneous samples and can provide significant biological information, such as promoter CpG island hypermethylation in human tumors, which is associated with the transcriptional silencing of genes relevant to the cancer process. The study concludes that MethylLight is a powerful tool for rapidly and accurately generating epigenetic profiles of tumor samples, which can be used to complement ongoing efforts to determine molecular profiles of tumor samples using high-throughput genomic and RNA-based technologies.MethylLight is a high-throughput assay for measuring DNA methylation using fluorescence-based real-time PCR (TaqMan®). This method allows for the detection of methylated alleles in the presence of a 10,000-fold excess of unmethylated alleles and provides highly quantitative results. The assay is designed to be sensitive, specific, and reproducible, and it can be performed with very small amounts of DNA, making it suitable for use with microdissected paraffin-embedded tissue samples. MethylLight distinguishes between mono- and bi-allelic methylation of the MLH1 gene in human colorectal tumor specimens. The technique is compatible with multiple gene loci and allows for rapid analysis of many samples. The study validates the MethylLight technology by comparing it with conventional methods such as COBRA and shows that it can accurately detect methylation status. The assay is also shown to be highly sensitive, capable of detecting methylation in the presence of a large amount of unmethylated DNA. The MethylLight technology is also shown to be reproducible on heterogeneous samples and can provide significant biological information, such as promoter CpG island hypermethylation in human tumors, which is associated with the transcriptional silencing of genes relevant to the cancer process. The study concludes that MethylLight is a powerful tool for rapidly and accurately generating epigenetic profiles of tumor samples, which can be used to complement ongoing efforts to determine molecular profiles of tumor samples using high-throughput genomic and RNA-based technologies.