Novel multiplex MethyLight protocol for detection of DNA methylation in patient tissues and bodily fluids.

Sci Rep

1] Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON, Canada [2] Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada [3] Department of Pathology and Laboratory Medicine, University Health Network, Toronto, Ontario, Canada.

Published: March 2014

Aberrant DNA methylation is a hallmark of cancer and is an important potential biomarker. Particularly, combined analysis of a panel of hypermethylated genes shows the most promising clinical performance. Herein, we developed, optimized and standardized a multiplex MethyLight assay to simultaneously detect hypermethylation of APC, HOXD3 and TGFB2 in DNA extracted from prostate cancer (PCa) cell lines, archival tissue specimens, and urine samples. We established that the assay is capable of discriminating between fully methylated and unmethylated alleles with 100% specificity and demonstrated the assay as highly accurate and reproducible as the singleplex approach. For proof of principle, we analyzed the methylation status of these genes in tissue and urine samples of PCa patients as well as PCa-free controls. These data show that the multiplex MethyLight assay offers a significant advantage when working with limited quantities of DNA and has potential applications in research and clinical settings.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3961737PMC
http://dx.doi.org/10.1038/srep04432DOI Listing

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