Bone marrow aspirates provide a rich source of cells for use in tissue engineering of bone and other clinical indications. However, progenitor cells such as mesenchymal stem cells (MSCs) account for a small fraction of nucleated cells in bone marrow aspirate (BMA), requiring extensive culture expansion. Accessory cell populations such as endothelial or hematopoietic cells can potentiate the bone-forming potential of MSCs, and cell-secreted extracellular matrix (ECM) can increase cell seeding efficiency and osteogenic differentiation of heterogeneous cell populations. In this study, we hypothesized that cell-secreted ECM could be used to sequester MSCs and accessory cells from BMA for bone regeneration. To generate 3D implantable constructs, BMA was resuspended in media with or without type I collagen or ECM and injected into a perfusion bioreactor system. The addition of protein coatings increased cell seeding efficiency compared to uncoated scaffolds. Compared to fresh BMA, the culture of BMA on all scaffolds reduced the proportion of CD45+ myeloid cells and increased CD31+CD45- endothelial cells. Compared to uncoated scaffolds, we observed a 143- and 30-fold increase in MSCs when fresh BMA was cultured on ECM- or collagen-coated scaffolds, respectively. Upon subcutaneous implantation, ECM-coated scaffolds promoted cell survival and early vascularization. However, bone formation was comparable across all implant groups, suggesting additional osteogenic cues are necessary to increase the bone forming potential of fresh BMA. These results motivate further investigation into strategies which elicit more robust bone regeneration by tissue aspirates.