We still know remarkably little about the extent to which neutral markers can provide a biologically relevant description of population structure. In the present study, we address this question, and quantify microsatellite differentiation among a small, structured island population of great tits (Parus major), and a large mainland population 150 km away. Although only a few kilometres apart, we found small but statistically significant levels of differentiation between the eastern and the western part of the island. On the other hand, there was no differentiation between the western part of the island and the mainland population, whereas the eastern part and the mainland did differ significantly. This initially counterintuitive result provides powerful support for the hypothesis that the large genetic difference in clutch size between both parts of the island found earlier is maintained by different levels of gene flow into both parts of the island, and illustrates the capac! ity of microsatellites to provide a meaningful description of population structure. Importantly, because the level of microsatellite differentiation is very low, we were unable to infer any population structure without grouping individuals a priori. Hence, these low levels of differentiation in neutral markers could easily remain undetected, or incorrectly be dismissed as biologically irrelevant. Thus, although microsatellites can provide a powerful tool to study genetic structure in wild populations, they should be used in conjunction with a range of other sources of information, rather than as a replacement.