We evaluated the usefulness of immunohistochemical examination for E-cadherin, p16, and cyclin D1 in discriminating melanoma from Spitz tumors. Immunoperoxidase staining was performed on formalin-fixed tissue specimens from 46 Spitz tumors and 42 concurrent melanoma specimens. The percentages of immunoreactive melanocytes in the epidermis and dermis were estimated semiquantitatively. Qualitatively abnormal immunoreactivity patterns were also tabulated. Dermal p16 immunoreactivity was the best quantitative discriminator: decreased nuclear immunoreactivity (<25% of dermal melanocytes) was 3-fold more likely in melanoma than in Spitz tumors (P = .004). Loss of both nuclear and cytoplasmic dermal p16 immunoreactivity was 8-fold more likely in melanoma (P = .01). Qualitative irregularities in the zonal distribution of E-cadherin immunoreactivity were 2-fold higher in melanoma (P = .01), but these were often focal or subtle. There was no statistically significant difference in cyclin D1 immunoreactivity. In atypical Spitz tumors, the dermal p16 immunoreactivity and frequency of qualitative E-cadherin abnormalities were intermediate between those of ordinary Spitz nevi and melanoma. Also, contrasting immunoreactivity patterns were helpful in determining Breslow thickness in specimens containing melanoma and contiguous dermal nevi.