Fibroblasts were isolated from human dental pulps of healthy third molars from 49 donors of ages ranging from 17 to 68. Significant variability was noted in the success of obtaining primary cultures from these pulps. Variability between the various cultures was also observed in the reliability of maintaining subcultures of the primary cultures as well as recovery from frozen stocks of established cell lines. Of the original 49 explant cultures studied, only three survived long-term passage and freezing. In addition to difficulties and variability in establishing cell lines, the human pulp fibroblasts also showed great variability in proliferative activity which could not be accounted for by donor age, source, or passage number. These findings highlight significant difficulties in establishing reliable human pulp fibroblast cultures and the need for great care in interpreting any in vitro data.