Figure 1 Melting curve analysis of BRAF mutations in thyroid surgical tissue samples. Fluorescently labelled oligonucleotide probes were used with extracted DNA to detect mutations in exon 15 of BRAF. Multiple probes complementary to the wild‐type (WT) sequences were placed within the same reaction, and the different sites were identified by their specific probe/target duplex melting temperatures. The position of each probe/target melting temperature and the relative ratio of the melting peak areas determined WT profiles. After amplification in a LightCycler, the instrument begins a melting programme where the reactions are cooled to anneal the probes and then slowly heated (0.1°C/s) while fluorescence is continuously monitored. Somatic mutations are identified by changes from a characteristic WT melting curve profile. When melting curves from non‐mutated and mutated samples are compared, additional melting peaks or changes in peak–area ratios indicate a sequence alteration (nucleotide mismatch) under the probe. Melting curve analysis revealed that the WT BRAF sequence Tm was 64.92°C±0.35°C, the mutation at nucleotide 1799, resulted in a shift to 60.11°C±0.46°C.