AIMS: To evaluate immunohistochemistry and reverse transcriptase polymerase chain reaction (RT-PCR) for melanoma associated antigens (MAA) in detecting occult melanoma cells in lymph nodes which were missed on routine pathology. METHODS: Occult melanoma cells were sought in 436 lymph nodes from 32 patients with cutaneous melanoma of the lower extremities by immunohistochemistry using the melanoma specific antibody HMB-45. The detection sensitivity of routine histology, immunohistochemistry, and RT-PCR was also compared in 23 lymph node samples from six patients. RESULTS: Immunohistochemistry showed that 15 of 24 patients (62.5%) who had no detectable metastasis by routine histology had at least one lymph node containing HMB-45 positive cells, mostly seen singly in the medullary sinus. No difference was found in known clinicopathological prognostic factors and recurrence rates between the two groups of patients with and without occult HMB-45 positive cells. RT-PCR analyses showed that the nested PCR for tyrosinase was more sensitive than a combination of single round PCR for five different MAA, including tyrosinase, MART-1/Melan A, Pmel-17, TRP-1, and TRP-2, detecting tyrosinase mRNA in six nodes which were negative by HMB-45 immunohistochemistry. CONCLUSIONS: Immunohistochemistry + RT-PCR is more sensitive than routine histology in detecting occult melanoma cells in lymph nodes. The nested PCR for tyrosinase should be used in future studies investigating the prognostic significance of such lymph node micrometastases.
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