Different degrees of clinical success have been reported for synthetic, allograft, and xenograft bone substitutes in human trials. Although these substitutes have been clinically investigated, their in vitro effects on cell differentiation remain unclear. Proliferation, differentiation, and attachment of dental pulp stem cells (DPSCs) to β-tricalcium phosphate (β-TCP), freeze-dried bone allograft (FDBA), and deproteinized bovine bone mineral (DBBM) were compared in this study. MTT assay, measurement of total DNA, and reverse transcriptase polymerase chain reaction were performed. β-TCP had the highest potential for DPSC attachment and proliferation, while FDBA induced osteoblastic differentiation of DPSCs. Further in vivo investigations are necessary to select a clinically appropriate scaffold.