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Comparison of methodologies for the detection of BRAF mutations in bone marrow trephine specimens


Aims BRAF V600E detection assists in the diagnosis of hairy cell leukaemia (HCL); however, testing practices vary. We evaluated the clinical utility of 5 BRAF mutation testing strategies for use on bone marrow trephines (BMT).

Methods 11 HCL, 5 HCL ‘mimic’, 2 treated HCL and 10 normal BMT specimens were tested for mutant BRAF, comparing Sanger sequencing, pyrosequencing, amplicon-based next generation sequencing (NGS), automated (Idylla) PCR and immunohistochemistry (IHC).

Results PCR and IHC were cheaper and identified V600E in 100 % of HCL cases. Pyrosequencing detected the mutation in 91%, NGS in 55% of cases and Sanger sequencing in 27%. All assays gave wild-type BRAF results in HCL mimics and normal BMT samples.

Conclusions PCR and IHC were most sensitive and cost-effective, but these have limited scope for multiplexing and are likely to be replaced by NGS gene panels or whole genome sequencing in the medium to long term.

  • Hairy cell leukaemia
  • bone marrow
  • BRAF
  • molecular pathology
  • immunohistochemistry

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