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Nucleic Acids Research 2006 34(3):e19; doi:10.1093/nar/gnj022
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Published online 9 February 2006

© The Author 2006. Published by Oxford University Press. All rights reserved
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Combination of methylated-DNA precipitation and methylation-sensitive restriction enzymes (COMPARE-MS) for the rapid, sensitive and quantitative detection of DNA methylation

Srinivasan Yegnasubramanian1, Xiaohui Lin1, Michael C. Haffner1,2, Angelo M. DeMarzo1 and William G. Nelson1,*

1Sidney Kimmel Comprehensive Cancer Center, The Johns Hopkins University School of Medicine 1650 Orleans Street, CRB 116, Baltimore, MD 21231, USA 2Innsbruck Medical University, Christoph-Probst-Platz 1 Innrain 52, A-6020 Innsbruck, Austria

*To whom correspondence should be addressed. Tel: +1 410 614 1661; Fax: +1 410 502 9817; Email: bnelson{at}jhmi.edu

Received November 7, 2005. Revised January 8, 2006. Accepted January 23, 2006.

Hypermethylation of CpG island (CGI) sequences is a nearly universal somatic genome alteration in cancer. Rapid and sensitive detection of DNA hypermethylation would aid in cancer diagnosis and risk stratification. We present a novel technique, called COMPARE-MS, that can rapidly and quantitatively detect CGI hypermethylation with high sensitivity and specificity in hundreds of samples simultaneously. To quantitate CGI hypermethylation, COMPARE-MS uses real-time PCR of DNA that was first digested by methylation-sensitive restriction enzymes and then precipitated by methyl-binding domain polypeptides immobilized on a magnetic solid matrix. We show that COMPARE-MS could detect five genome equivalents of methylated CGIs in a 1000- to 10 000-fold excess of unmethylated DNA. COMPARE-MS was used to rapidly quantitate hypermethylation at multiple CGIs in >155 prostate tissues, including benign and malignant prostate specimens, and prostate cell lines. This analysis showed that GSTP1, MDR1 and PTGS2 CGI hypermethylation as determined by COMPARE-MS could differentiate between malignant and benign prostate with sensitivities >95% and specificities approaching 100%. This novel technology could significantly improve our ability to detect CGI hypermethylation.


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