Comprehensive long-term studies of isolated populations provide valuable comparative data that may be used to evaluate different methods for quantifying the relationship between genetic diversity and fitness. Here, we report on data collected from large and well-characterized cohorts of the two numerically dominant species of Darwin’s finches on Isla Daphne Major, Gala´pagos, Ecuador – Geospiza fortis and G. scandens. Multilocus microsatellite (SSR) genetic diversity estimates (heterozygosity and d2) and pedigreebased estimates of the inbreeding coefficient (f) were compared to each other and to two fitness components: lifespan and recruitment. In the larger sample of G. fortis, heterozygosity (H)was correlated with both fitness components, but no relationship was detected in the smaller sample of G. scandens. Analyses of the inbreeding coefficient detected highly significant relationships between f and recruitment, but no relationship between f and overall lifespan. The d2 statistic showed no relationship to either fitness component. When the two SSR-based estimators were compared to f, d2 was correlated with f in G. fortis in the predicted direction, while in G. scandens the relationship was positive. Multilocus heterozygosity was correlated with f in G. fortis but not in the G. scandens sample. A pedigree simulation demonstrated that the variation in true autozygosity can be large among individuals with the same level of inbreeding. This observation may supplement the interpretation of patterns relevant to the local (locus-specific) and general (genome-wide) effects hypotheses, which have been proposed to explain the mechanism responsible for associations between genetic diversity and fitness.