Quantification of regional DNA methylation by liquid chromatography/tandem mass spectrometry

Zhongfa Liu, Jiejun Wu, Zhiliang Xie, Shujun Liu, Patty Fan-Havard, Tim H.M. Huang, Christoph Plass, Guido Marcucci, Kenneth K. Chan

Research output: Contribution to journalArticlepeer-review

23 Scopus citations


Promoter hypermethylation-associated tumor suppressor gene (TSG) silencing has been explored as a therapeutic target for hypomethylating agents. Promoter methylation change may serve as a pharmacodynamic endpoint for evaluation of the efficacy of these agents and predict the patient's clinical response. Here a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay has been developed for quantitative regional DNA methylation analysis using the molar ratio of 5-methyl-2′-deoxycytidine (5mdC) to 2′-deoxycytidine (2dC) in the enzymatic hydrolysate of fully methylated bisulfite-converted polymerase chain reaction (PCR) amplicons as the methylation indicator. The assay can differentiate 5% of promoter methylation level with an intraday precision ranging from 3 to 16% using two TSGs: HIN-1 and RASSF1A. This method was applied to characterize decitabine-induced promoter DNA methylation changes of these two TSGs in a breast cancer MCF-7 cell line. Promoter methylation of these TSGs was found to decrease in a dose-dependent manner. Correspondingly, the expression of these TSGs was enhanced. The sensitivity and reproducibility of the method make it a valuable tool for specific gene methylation analysis that could aid characterization of hypomethylating activity on specific genes by hypomethylating agents in a clinical setting.

Original languageEnglish (US)
Pages (from-to)106-113
Number of pages8
JournalAnalytical Biochemistry
Issue number2
StatePublished - Aug 15 2009
Externally publishedYes


  • LC-MS/MS
  • Quantification
  • Regional DNA methylation

ASJC Scopus subject areas

  • Molecular Biology
  • Biophysics
  • Biochemistry
  • Cell Biology


Dive into the research topics of 'Quantification of regional DNA methylation by liquid chromatography/tandem mass spectrometry'. Together they form a unique fingerprint.

Cite this