Analytical sensitivity and stability of DNA methylation testing in stool samples for colorectal cancer detection

Linda J. W. Bosch, Sandra Mongera, Jochim S. Terhaar Sive Droste, Frank A. Oort, Sietze T. van Turenhout, Maarten T. Penning, Joost Louwagie, Chris J. J. Mulder, Manon van Engeland, Beatriz Carvalho, Gerrit A. Meijer*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review


Stool-based molecular tests hold large potential for improving colorectal cancer screening. Here, we investigated the analytical sensitivity of a DNA methylation assay on partial stool samples, and estimated the DNA degradation in stool over time. In addition, we explored the detection of DNA methylation in fecal immunochemical test (FIT) fluid. Partial stool samples of colonoscopy-negative individuals were homogenized with stool homogenization buffer, spiked with different numbers of HCT116 colon cancer cells and kept at room temperature for 0, 24, 48, 72 and 144 h before DNA isolation. Analytical sensitivity was determined by the lowest number of cells that yielded positive test results by DNA methylation or mutation analysis. DNA methylation in FIT fluid was measured in 11 CRC patients and 20 control subjects. The analytical sensitivity for detecting DNA methylation was 3000 cells per gram stool, compared to 60000 cells per gram stool for detection of DNA mutations in the same stool samples. No degradation up to 72 h was noted when a conservation buffer was used. DNA methylation was detected in 4/11 CRC FIT samples and in none of the 20 control FIT samples. Methylation based stool DNA testing showed a high analytical sensitivity for tumor DNA in partial stool samples, which was hardly influenced by DNA degradation over time, provided an adequate buffer was used. The feasibility of detecting DNA methylation in FIT fluid demonstrates the opportunity to combine testing for occult blood with DNA methylation in the same collection device.
Original languageEnglish
Pages (from-to)309-315
JournalCellular oncology
Issue number4
Publication statusPublished - Aug 2012


  • DNA methylation
  • CRC
  • Screening
  • Stool
  • Analytical sensitivity
  • Stability
  • FIT
  • iFOBT


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