A multisite blinded study for the detection of BRAF mutations in formalin-fixed, paraffin-embedded malignant melanoma Academic Article uri icon

abstract

  • Melanoma patients with BRAF mutations respond to treatment with vemurafenib, thus creating a need for accurate testing of BRAF mutation status. We carried out a blinded study to evaluate various BRAF mutation testing methodologies in the clinical setting. Formalin-fixed, paraffin-embedded melanoma samples were macrodissected before screening for mutations using Sanger sequencing, single-strand conformation analysis (SSCA), high resolution melting analysis (HRM) and competitive allele-specific TaqMan® PCR (CAST-PCR). Concordance of 100% was observed between the Sanger sequencing, SSCA and HRM techniques. CAST-PCR gave rapid and accurate results for the common V600E and V600K mutations, however additional assays are required to detect rarer BRAF mutation types found in 3-4% of melanomas. HRM and SSCA followed by Sanger sequencing are effective two-step strategies for the detection of BRAF mutations in the clinical setting. CAST-PCR was useful for samples with low tumour purity and may also be a cost-effective and robust method for routine diagnostics.

authors

  • Richter, A
  • Grieu, F
  • Carrello, A
  • Amanuel, B
  • Namdarian, K
  • Rynska, A
  • Lucas, A
  • Michael, V
  • Bell, A
  • Fox, SB
  • Hewitt, CA
  • Do, H
  • McArthur, GA
  • Wong, SQ
  • Dobrovic, A
  • Iacopetta, B

publication date

  • 2013