A comparison of three methods for detecting KRAS mutations in formalin-fixed colorectal cancer specimens.

D. Gonzalez de Castro, B. Angulo, B. Gomez, D. Mair, R. Martinez, A. Suarez-Gauthier, F. Shieh, M. Velez, V.H. Brophy, H.J. Lawrence, F. Lopez-Rios

Research output: Contribution to journalArticlepeer-review

70 Citations (Scopus)


BACKGROUND: KRAS mutation testing is required to select patients with metastatic colorectal cancer (CRC) to receive anti-epidermal growth factor receptor antibodies, but the optimal KRAS mutation test method is uncertain. METHODS: We conducted a two-site comparison of two commercial KRAS mutation kits - the cobas KRAS Mutation Test and the Qiagen therascreen KRAS Kit - and Sanger sequencing. A panel of 120 CRC specimens was tested with all three methods. The agreement between the cobas test and each of the other methods was assessed. Specimens with discordant results were subjected to quantitative massively parallel pyrosequencing (MPP). DNA blends were tested to determine detection rates at 5% mutant alleles. RESULTS: Reproducibility of the cobas test between sites was 98%. Six mutations were detected by cobas that were not detected by Sanger, and five were confirmed by MPP. The cobas test detected eight mutations which were not detected by the therascreen test, and seven were confirmed by MPP. Detection rates with 5% mutant DNA blends were 100% for the cobas and therascreen tests and 19% for Sanger. CONCLUSION: The cobas test was reproducible between sites, and detected several mutations that were not detected by the therascreen test or Sanger. Sanger sequencing had poor sensitivity for low levels of mutation.
Original languageEnglish
Pages (from-to)345-351
Number of pages7
JournalBritish Journal of Cancer
Issue number2
Publication statusPublished - 10 Jul 2012


  • Adult
  • Aged
  • Aged, 80 and over
  • Colorectal Neoplasms
  • Female
  • Formaldehyde
  • Humans
  • Male
  • Middle Aged
  • Mutation
  • Proto-Oncogene Proteins
  • Reproducibility of Results
  • Sequence Analysis, DNA
  • Tissue Fixation
  • ras Proteins


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