An Immunohistochemical Comparison Study of SOX10, Pan Melanoma Cocktail and S100 in Malignant Melanoma

Design Formalin-fixed, paraffin-embedded tissue microarrays (TMAs) consisting of malignant and metastatic melanoma, including SCM and DM (n = 72), were stained by immunohistochemistry and compared in a direct study using a mouse monoclonal SOX10 [BC34] (nuclear), a Pan Melanoma antibody cocktail (cytoplasmic) (Biocare Medical) and a polyclonal S100 antibody (cytoplasmic/ nuclear) (Dako). TMA tissue sections and normal lymph node tissues were deparaffinized and hydrated down to water. Slides were placed in a modified citrate buffer antigen retrieval solution in a pressure cooker and heated to 110°C for 15 minutes. SOX10, Pan Melanoma Cocktail and S100 were optimized for immunohistochemistry and incubated for 30 minutes, followed by an alkaline phosphatase (AP) polymer detection system and visualized with Fast Red chromogen. A melanoma triple cocktail of SOX10, MART-1 and Tyrosinase was applied simultaneously to tissues sections, detected with an AP polymer detection system and visualized with Fast Red. A sequential two-color double stain for a single section was accomplished by incubating SOX10 for 30 minutes, followed by a horseradish peroxidase polymer detection system and visualization with DAB; the Pan Melanoma Cocktail was incubated for 30 minutes followed by an AP polymer detection system and visualization with Fast Red. Results In malignant melanoma cases, SOX10, Pan Melanoma and S100 stained 88%, 90% and 88%, respectively (Figure 1, Table 1). In metastatic melanoma, SOX10, Pan Melanoma and S100 stained 81%, 89% and 81%, respectively (Figures 2A-C, Table 1). The combination of SOX10 and Pan Melanoma stained 92% (24/26) of metastatic melanomas (Figure 2D, data not shown). In addition, SOX10 and S100 both stained 100% of DM and SCM, whereas Pan Melanoma stained 50% (Figure 3, Table 1). Normal lymph node was negative for SOX10 and Pan Melanoma but was strongly positive for S100 (Figure 4). The melanoma triple cocktail (Fast Red only) and the sequential double stain of SOX10 (DAB) and Pan Melanoma (Fast Red) were successfully achieved and could be visualized in a single section (Figure 5). Introduction Traditionally, melanoma markers such as S100, HMB45, and MART-1 have been used as a panel of antibodies to identify melanoma.1-5 The S100 antibody has been primarily used as a first-line screener for melanoma for the last three decades. However, S100 has been shown to lack specificity as it stains interdigitating reticulum cells in the paracortex of lymph nodes, a common site of metastatic melanoma and its mimics.1, 6 A cocktail of MART-1 and Tyrosinase (known as Pan Melanoma) has been shown to be a very sensitive and specific marker in metastatic melanomas and studies have shown its sensitivity was comparable to that of S100.7 However, MART-1 and Tyrosinase have demonstrated a lack of sensitivity in desmoplastic melanoma (DM) and spindle cell melanoma (SCM).1, 5 SOX10 is a nuclear transcription factor that plays an important role in melanocytic cell differentiation. It has been shown to be a sensitive marker for melanoma including spindle and desmoplastic subtypes.8-10 A mouse monoclonal SOX10 antibody has recently been developed that was shown to stain the majority of melanomas and most importantly, stained 98% (47/48) of DM and SCM.10