Related Experiment Video
Updated: May 21, 2026
Employing Digital Droplet PCR to Detect BRAF V600E Mutations in Formalin-fixed Paraffin-embedded Reference Standard Cell Lines
Published on: October 8, 2015
Detecting BRAF Mutations in Formalin-Fixed Melanoma: Experiences with Two State-of-the-Art Techniques
Nicola L Schoenewolf1, Reinhard Dummer, Daniela Mihic-Probst
1Department of Dermatology, University Hospital Zürich, Zürich, Switzerland.
Background:
Melanoma is characterized by a high frequency of BRAF mutations. It is unknown if the BRAF mutation status has any predictive value for therapeutic approaches such as angiogenesis inhibition.
Patients And Methods:
We used 2 methods to analyze the BRAF mutation status in 52 of 62 melanoma patients. Method 1 (mutation-specific real-time PCR) specifically detects the most frequent BRAF mutations, V600E and V600K. Method 2 (denaturing gel gradient electrophoresis and direct sequencing) identifies any mutations affecting exons 11 and 15.
Results:
Eighteen BRAF mutations and 15 wild-type mutations were identified with both methods. One tumor had a double mutation (GAA) in codon 600. Results of 3 samples were discrepant. Additional mutations (V600M, K601E) were detected using method 2. Sixteen DNA samples were analyzable with either method 1 or method 2. There was a significant association between BRAF V600E mutation and survival.
Conclusion:
Standardized tissue fixation protocols are needed to optimize BRAF mutation analysis in melanoma. For melanoma treatment decisions, the availability of a fast and reliable BRAF V600E screening method may be sufficient. If other BRAF mutations in exons 11 and 15 are found to be of predictive value, a combination of the 2 methods would be useful.
