Bone defect repair, particularly in the alveolar region, remains a significant hurdle in periodontics. In recent years, the spotlight in regenerative medicine has fallen on 3D-printed bone scaffolds, especially those constructed of polylactic acid (PLA) infused with hydroxyapatite. This research introduced a novel approach by developing a 3D-printed PLA scaffold enriched with hydroxyapatite derived from fish skin waste (FSHA). Mechanical compression tests revealed that the 3D-printed PLA-FSHA scaffolds had a compressive strength (13.4 ± 5.53 MPa) in the same ballpark as their reference PLA counterparts (20.3 ± 1.08 MPa). Scanning electron micrographs highlighted an average pore size in the scaffold (572 ± 33 μm) conducive to angiogenesis and facilitating cell migration and proliferation. In vitro, cytotoxicity was ascertained using the MTT assay on L929 fibroblast cells. Further in vitro cytocompatibility assessments through actin-DAPI staining and measurements of bone regeneration markers - alkaline phosphatase, osteocalcin and osteopontin-demonstrated that the PLA-FSHA scaffolds not only were biocompatible but also showcased performance on par with the commercial graft, osseograft. This lays the foundation for future in vivo evaluations of bone regenerative capabilities.