Immunoperoxidase technique modified by counterstain with azure B as a diagnostic aid in evaluating heavily pigmented...

Heavily-pigmented melanocytic neoplasms are difficult to evaluate on routine hematoxylin and eosin stained slides because pigmented melanocytes are difficult to distinguish from the numerous melanophages that are usually seen in the background of these lesions. Immunoperoxidase staining for S100 protein or HMB-45 antibody using diaminobenzidine (DAB) as chromogen, which forms a brown product, does not adequately distinguish melanocytes from melanophages. We modified this technique by replacing hematoxylin as the counterstain with azure B, which stains melanin green-blue. Thus, positive melanocytes appear brown while melanin granules in their cytoplasm are green-blue. However, negative melanophages only stain green-blue. This technique is useful in evaluating heavily pigmented melanocytic lesions such as malignant melanomas, melanosis of regressing malignant melanoma, residual malignant melanoma in areas of granulation tissue with melanophages, blue nevi, pigmented spindle cell variant of Spitz's nevi and combined nevi.