Application of immunocytochemistry and BRAF mutational analysis to direct smears of metastatic melanoma

Cancer Cytopathol. 2012 Feb 25;120(1):52-61. doi: 10.1002/cncy.20180. Epub 2011 Jul 25.


Background: The cytodiagnosis of melanoma in fine-needle aspiration (FNA) specimens can be challenging, often requiring the use of immunocytochemistry. As constitutively activating mutations in the BRAF oncogene are present in at least 40% of melanomas, the use of FNA material to interrogate the BRAF mutational status is likely to increase. Because cell blocks, traditionally used for these studies, can occasionally exhibit insufficient tumor cellularity, the authors investigated the utility of direct smears for immunocytochemistry and BRAF mutational analysis.

Methods: Immunocytochemistry for S-100, HMB-45, and Mart-1 was prospectively performed on direct smears in 17 FNAs of metastatic melanoma. Next, BRAF sequencing was performed using DNA isolated from archived Diff-Quik-stained direct smears for 15 cases. In parallel, sequencing was performed using DNA obtained from corresponding cell blocks.

Results: S-100 positivity in the tumor cells was observed in all 17 cases. HMB-45 and Mart-1 positivity was noted in 81% and 88% of cases, respectively. All 3 markers were positive in 76% of cases. Next, of the 15 archived melanoma FNAs tested, BRAF mutations were observed in 8 (53%); 5 and 3 melanomas harbored the V600E and V600K mutation, respectively. Corresponding cell blocks were also tested for all 15 cases, yielding concordant BRAF results in 14 (93%); 1 cell block yielded a false-negative result.

Conclusions: Cytologic direct smears represent a robust and valuable source of cellular material for immunocytochemistry and molecular studies, especially in instances in which inadequate cell block cellularity is anticipated or encountered.

Publication types

  • Evaluation Study

MeSH terms

  • Cytodiagnosis / methods*
  • DNA Mutational Analysis*
  • Humans
  • Immunohistochemistry*
  • Melanoma / diagnosis*
  • Melanoma / genetics
  • Melanoma / pathology
  • Melanoma / secondary*
  • Mutation*
  • Proto-Oncogene Proteins B-raf / genetics*


  • BRAF protein, human
  • Proto-Oncogene Proteins B-raf