Small and isolated island populations provide ideal systems to study the effects of limited population size, genetic drift and gene flow on genetic diversity. We assessed genetic diversity within and differentiation among 19 mockingbird populations on 15 Galapagos islands, covering all four endemic species, using 16 microsatellite loci. We tested for signs of drift and gene flow, and used historic specimens to assess genetic change over the last century and to estimate effective population sizes. Within-population genetic diversity and effective population sizes varied substantially among island populations and correlated strongly with island size, suggesting that island size serves as a good predictor for effective population size. Genetic differentiation among populations was pronounced and increased with geographical distance. A century of genetic drift did not change genetic diversity on an archipelago-wide scale, but genetic drift led to loss of genetic diversity in small populations, especially in one of the two remaining populations of the endangered Floreana mockingbird. Unlike in other Galapagos bird species such as the Darwin's finches, gene flow among mockingbird populations was low. The clear pattern of genetically distinct populations reflects the effects of genetic drift and suggests that Galapagos mockingbirds are evolving in relative isolation.