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Over 500,000 bone graft or bio-implant procedures are performed annually in the United States. It has been reported that osseous autograft procurement may result in donor site complications and bio-implant allografts have been associated with disease transmission. Ceramic scaffolds are only osteoconductive, limiting their clinical use. The objective of this study was to create a bone filler substitute with regenerating properties similar to natural bone. Therefore, melatonin and platelet-rich plasma (PRP) were utilized for their known osteoinductive properties. It was hypothesized that melatonin and/or PRP would enhance the osteoinductive and osteoconductive properties of calcium aluminate (CA) scaffolds to promote bone regeneration in a model of calvarial defects. The biocompatibility of CA and CA-Mel scaffolds was tested in vitro and in vivo. Data show that CA-Mel scaffolds, in comparison with CA scaffolds, enhanced the adhesion, viability, and proliferation of normal human osteoblasts cells but not that of NIH3T3 fibroblasts. Data also showed that human adult mesenchymal stem cells grown on CA or CA-Mel scaffolds showed a time-dependent induction into osteoblasts over 14 days revealed through scanning electron microscopy and by alkaline phosphatase analyses. Implantation of CA-Mel scaffolds into critical size calvarial defects in female, ovariectomized rats showed that the CA-Mel scaffolds were biocompatible, allowed for tissue infiltration, and showed evidence of scaffold biodegradation by 3 and 6 months. Bone regeneration, assessed using fluorochrome labeling at 3 and 6 months, was greatest in animals implanted with the CA-Mel scaffold. Overall, results from this study show that CA-Mel scaffolds were osteoconductive and osteoinductive.