Introduction: The aim of this study was to assess the biological properties and mineralization potential of the new Theracal PT (Bisco Inc, Schaumburg, IL) compared with its predecessor Theracal LC (Bisco Inc) and the hydraulic silicate-based cement Biodentine (Septodont, Saint-Maur-des-Fosses, France) on human dental pulp stem cells (hDPSCs) in vitro. Methods: Standardized sample discs were obtained for each material (n 5 30) together with 1:1, 1:2, and 1:4 material eluates. Previously characterized hDPSCs were cultured with the different materials in standardized conditions, and the following assays were performed: a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, a wound healing assay, Annexin-V-FITC and 7-AAD staining (BD Biosciences, San Jose, CA), reactive oxygen species production analysis, cell adhesion and morphology evaluation via scanning electron microscopy and immunofluorescence, quantification of the expression of osteo/odontogenic markers via real-time quantitative reverse-transcriptase polymerase chain reaction, and alizarin red S staining. Statistical significance was established at P , .05. Results: All of the tested dilutions of Theracal LC exhibited a significantly higher cytotoxicity and reactive oxygen species production (P , .001) and a lower cell migration rate than the control group (hDPSCs cultured in growth medium without material extracts) at all of the measured time points (P , .001). Both 1:4 Theracal PT and Biodentine-treated hDPSCs exhibited similar levels of cytocompatibility to that of the control group, a significant up-regulation of at least 1 odontogenic marker (Biodentine: dentin sialophosphoprotein (P , .05); Theracal PT: osteonectin and runt-related transcription factor 2 [P , .001]), and a significantly higher mineralized nodule formation (P , .001). Conclusions: The newly introduced TheraCal PT offers an improved in vitro cytocompatibility and mineralization potential on hDPSCs compared with its predecessor, TheraCal LC, and comparable biological properties to Biodentine