BACKGROUND: The design of melanoma sentinel lymph node (SLN) histologic protocols is based on the premise that most metastases are found in the central parts of the nodes, but the evidence for this belief has never been thoroughly tested. METHODS: The nodal location of melanoma metastases in 149 prospectively analyzed, completely step sectioned, positive SLNs from 96 patients was examined using 3 theoretical protocols, evaluating respectively: (1) the 3 most central step sections only; (2) the 3 most peripheral step sections only; and (3) 3 step sections evenly distributed throughout the individual SLNs. In addition, the size of the metastases located exclusively outside the 2 regional protocols (ie, 3 central sections, and 3 peripheral sections) were measured and compared with each other. RESULTS: The metastasis detection rates of the central, the peripheral, and the evenly distributed protocols were 77%, 79%, and 78%, respectively. No difference in either the mean volume or the maximum diameter of the metastases located exclusively outside the central and the peripheral protocols was found (volume: 0.036 vs. 0.031 mm and diameter: 0.320 vs. 0.332 mm). CONCLUSIONS: In SLNs, melanoma metastases are located throughout the nodes. Metastases located exclusively outside the peripheral or the central protocol are equally sized. Complete step sectioning of all SLNs will ensure both high metastasis detection rates and detection of all large metastases, and allow for performance of unbiased size estimates.