The amount of bone generated using current tissue engineering approaches is insufficient for many clinical applications. Previousin vitrostudies suggest that culturing cells as 3D aggregates can enhance their osteogenic potential, but the effect on bone formationin vivois unknown. Here, we use agarose wells to generate uniformly sized mesenchymal stromal cell (MSC) aggregates. When combined with calcium phosphate ceramic particles and a gel prepared from human platelet-rich plasma, we generated a tissue engineered construct which significantly improvedin vivobone forming capacity as compared to the conventional system of using single cells seeded directly on the ceramic surface. Histology demonstrated the reproducibility of this system, which was tested using cells from four different donors.In vitrostudies established that MSC aggregation results in an up-regulation of osteogenic transcripts. And finally, thein vivoperformance of the constructs was significantly diminished when unaggregated cells were used, indicating that cell aggregation is a potent trigger ofin vivobone formation by MSCs. Cell aggregation could thus be used to improve bone tissue engineering strategies.