Immunohistochemical dual staining as an adjunct in assessment of mitotic activity in melanoma

Background: The mitotic rate was introduced as a major prognostic criterion for the staging of thin (=1.0 mm) melanoma by the 2009 American Joint Committee on Cancer Staging and Classification (seventh edition). The detection of a single mitotic figure changes the tumor stage in thin melanoma. We sought to address the value of a dual staining to facilitate the determination of the mitotic rate and to assign the mitotic activity to melanocytes. Methods: The mitotic rate of melanoma cells was determined by dual phosphohistone-H3 (PHH3)/Melan-A immunohistochemistry. Results were compared with PHH3 staining alone and conventional hematoxylin and eosin (H&L)-stained slides of 15 melanomas with a tumor thickness <1.0 mm. Results: PHH3 staining clearly labeled cells in the mitotic cell cycle. The mitotic rate in the PHH3/Melan-A dual stain was equal to that derived by H&E staining. Time required for counting mitotic figures was significantly reduced. Conclusions: The evaluation of mitotic rate with an immunohistochemical dual stain is faster (mean 63.0%) and more reliable than evaluation by routine H&E staining alone. Dual staining immunohistochemistry may be a useful additional tool to standardize the determination of mitotic rate and may be helpful in evaluation of challenging cases.