The formation of new cementum is an important issue in clinical periodontology, as cementum is required to provide attachment for newly formed periodontal tissues to the root surface. In this study a model of cementogenesis in vitro was used in order to test the effects of root surface demineralization on the migration, attachment and formation of a cementum-like tissue by osteoblastic cells cultured on cementum and to test the specificity of cementum matrix in modulating those effects by comparison of root co-cultures with bone co-cultures. It was demonstrated that root surface demineralization did not significantly alter the orientation, number and attachment of cells to the root co-cultures. The results also demonstrated that cementum and bone matrix appear to behave differently in culture, as seen by their distinct action on the morphological profile of the attached cells and the extracellular matrix deposited by these cells. These results demonstrate that although cementum matrix appears to stimulate the production of cementum-like tissue, this action is not confined to cementum matrix alone, since a similar material was also deposited on dentine and bone surfaces. Thus, these results do not support a specific action of cementum matrix on the modulation of the cementoblast phenotype. The use of co-cultures of neonatal rat calvaria cells with root slices represents a promising model of cementogenesis in vitro; however, studies should be undertaken towards the identification of markers to distinguish between cementoblast and osteoblast phenotypes in order to further validate this model.