Earlier studies on human fibroblast cultures have demonstrated an inverse relationship between the total number of population doublings (PDs) and donor age. As more recent studies were unable to replicate these findings, we set out to analyze growth characteristics of fibroblast cultures from nonagenarians who represent the extreme of human lifespan. Therefore, we obtained skin biopsies from 68 participants of the Leiden 85-plus Study, all aged 90 years. None of the 68 strains failed to proliferate and all were easily cultured under highly standardized conditions. Within a time window of 30 months, all strains displayed a high and reproducible replicative capacity that was maintained for at least 50 PDs. A decline in mitotic activity was observed between 26 and 81 PDs. Out of the 68 cell strains, 58 strains reached the post-mitotic phase with an onset between 51 and 108 PDs. The growth pattern of each senescent strain was fitted by a piecewise linear model, which allowed calculation of the transition towards the phase of decreased growth speed, as well as by a nonlinear continuous model; goodness-of-fit was high and not different between the models (both > 0.99). Growth characteristics were not associated with morbidity or mortality of the donors. We conclude that fibroblasts from nonagenarians maintain a high-replicative capacity despite a huge variability in the onset of senescence. These results cast further doubt on the association between in vitro growth characteristics of fibroblast cultures and the length of human lifespan.