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Background: Sentinel lymph node biopsy (SLNB) is a critical staging tool for melanoma patients. The optimal number of lymph nodes removed in SLNB remains unclear. In this study, we retrospectively analysed and tested different criteria for selecting sentinel lymph nodes (SLNs) by radiotracer uptake and blue dye, and their impact on nodal staging. We also evaluated the association between SLN tumour burden and radiotracer uptake., Methods: The study population consisted of melanoma patients undergoing SLNB. During the operation all radioactive and blue nodes were removed and sent for histopathological analysis. The ex vivo radioactive count and presence of blue dye of each node were recorded, and these were correlated with presence and size of metastasis in each SLN., Results: Altogether 175 patients with clinically occult metastasis presented with one or more positive, i.e. metastatic, SLNs. The mean number of lymph nodes removed was 4.5, and the mean number of positive lymph nodes was 1.5 per patient. The most radioactive or hottest node was negative in 38 patients (22%). By removing the hottest node and all nodes with radioactivity >10% of the hottest node, 97% of patients would have been staged correctly. In five patients, metastasis was found solely in a SLN with radioactivity <10% of the hottest node. Of all 267 positive nodes removed, 125 (47%) contained blue dye. Patients with a negative hottest node were associated with lower SLN tumour burden., Conclusions: By removing the hottest node and all nodes with radioactivity >10% of the hottest node, 97% of patients with SLN metastases are correctly staged with or without using blue dye.