Immunohistochemistry cannot replace DNA analysis for evaluation ofBRAFV600E mutations in papillary thyroid carcinoma

Abstract

IntroductionThe BRAF V600E mutation is the most common genetic event occurring in papillary thyroid cancer (PTC). Recently, the possibility of using immunohistochemistry (IHC) to detect the BRAF V600E mutation has been reported.Materials and MethodsIn 140 patients with classical PTC, the status of the BRAF V600E mutation was determined by IHC (using two alternative staining protocols, IHC-1 and IHC-2) and molecular biology methods: Sanger sequencing (SEQ) and real-time PCR (qPCR).ResultsThe BRAF V600E mutation was detected in 57.1% (80/140) patients by IHC-1 and 62.9% (88/140) patients by IHC-2. The highest correlation in detecting the BRAF V600E mutation was found between IHC-2 and qPCR (94.2%), and between IHC-1 and qPCR (83.9%). Correlations between IHC-1 and SEQ and between IHC-2 and SEQ were 71.5% and 76.2%, respectively. The IHC-2 protocol had higher sensitivity, PPV, and NPV, and Cohen’s kappa than IHC- 1. The presence of BRAF V600E mutation in IHC-2 statistically correlated with age at diagnosis, histopathological stage, and extrathyroidal extension.ConclusionsThe results obtained in this study indicate a lack of concordance between BRAF V600E detection by IHC and molecular methods. The IHC method cannot replace molecular methods for the detection of the BRAF V600E mutation.

VE1 Immunohistochemistry Improves the Limit of Genotyping for Detecting BRAFV600E Mutation in Papillary Thyroid Cancer

“…Although Sanger sequencing has been widely acknowledged as the standard technique for detection of point mutation, from the literature review, we found out that the discordant rates are much higher when using Sanger sequencing (7-23%) to validate the performance of VE1 IHC [20,23,25,31,35,[38][39][40], than using a more sensitive molecular method, such as real-time PCR (0.1-8%) [19,22,23,[41][42][43][44][45]. Several groups achieved a discordance rate of < 2% when VE1 immunostaining was compared to real-time PCR [22,43].…”

“…Sanger sequencing has been widely utilized as the gold standard method; however, it has a reported detection sensitivity of only 10-20% of mutant allele frequency, and therefore, this method alone might not be sufficient to detect the mutation in cases with low tumor cellularity [18][19][20]. Pyrosequencing and real-time polymerase chain reaction (PCR) are known to be more sensitive than Sanger sequencing, with a reported detection sensitivity of 5% and 1% mutant alleles, respectively [21][22][23]. Since different techniques have different performance rates, the methods employed to detect the mutation might have a significant impact on the prevalence rate of the mutation.…”

Utility of the BRAF p.V600E immunoperoxidase stain in FNA direct smears and cell block preparations from patients with thyroid carcinoma

“…In patients with advanced thyroid cancer, targeted therapy using selective BRAF inhibitors is considered based on the presence of the BRAF p.V600E mutation. Although several studies have described BRAF p.V600E immunohistochemistry in surgical specimens of PTC as a rapid and cheaper alternative with a high sensitivity and specificity, molecular testing by mutational analysis remains the gold standard because IPOX rarely may fail to detect the BRAF p.V600E mutation . In the current study cohort, the majority of the surgical cases had BRAF p.V600E mutation status determined by molecular testing using next‐generation sequencing, with 9 cases that were evaluated by BRAF p.V600E (VE1) IPOX.…”

BRAF V600E expression in ameloblastomas—A 36‐patient cohort from Helsinki University Hospital

“…It has been discussed that immunohistochemistry for identifying BRAFV600E‐mutated protein from formalin‐fixed, paraffin‐embedded samples is reliable compared to molecular techniques, thus making it a beneficial tool to detect these mutations as part of the normal diagnostic (Brown et al, ; Capper et al, ). Recently also opposite results have emerged (Szymonek et al, ). Decalcification with formic acid, however, affects immunoreactivity.…”