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  • Title: Ultra-deep sequencing confirms immunohistochemistry as a highly sensitive and specific method for detecting BRAF V600E mutations in colorectal carcinoma.
    Author: Rössle M, Sigg M, Rüschoff JH, Wild PJ, Moch H, Weber A, Rechsteiner MP.
    Journal: Virchows Arch; 2013 Nov; 463(5):623-31. PubMed ID: 24085553.
    Abstract:
    The activating BRAF (V600) mutation is a well-established negative prognostic biomarker in metastatic colorectal carcinoma (CRC). A recently developed monoclonal mouse antibody (clone VE1) has been shown to detect reliably BRAF (V600E) mutated protein by immunohistochemistry (IHC). In this study, we aimed to compare the detection of BRAF (V600E) mutations by IHC, Sanger sequencing (SaS), and ultra-deep sequencing (UDS) in CRC. VE1-IHC was established in a cohort of 68 KRAS wild-type CRCs. The VE1-IHC was only positive in the three patients with a known BRAF (V600E) mutation as assessed by SaS and UDS. The test cohort consisted of 265 non-selected, consecutive CRC samples. Thirty-nine out of 265 cases (14.7%) were positive by VE1-IHC. SaS of 20 randomly selected IHC negative tumors showed BRAF wild-type (20/20). Twenty-four IHC-positive cases were confirmed by SaS (24/39; 61.5%) and 15 IHC-positive cases (15/39; 38.5%) showed a BRAF wild-type by SaS. UDS detected a BRAF (V600E) mutation in 13 of these 15 discordant cases. In one tumor, the mutation frequency was below our threshold for UDS positivity, while in another case, UDS could not be performed due to low DNA amount. Statistical analysis showed sensitivities of 100% and 63% and specificities of 95 and 100% for VE1-IHC and SaS, respectively, compared to combined results of SaS and UDS. Our data suggests that there is high concordance between UDS and IHC using the anti-BRAF(V600E) (VE1) antibody. Thus, VE1 immunohistochemistry is a highly sensitive and specific method in detecting BRAF (V600E) mutations in colorectal carcinoma.
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