Abstract:[Objective] To investigate the biocompatibility and osteogenic properties of acellular bone matrix/calcium phosphate/calcium sulfate hemihydrate (ACBM/CPC/CSH) composite artificial bone. [Methods] ACBM/CPC/CSH composite was synthesized by physical mixing method, and the material extract was prepared for acute and subacute toxicity, pyrogen and epidermal irritation tests. The material was implanted in the muscle and histopathological examination was performed on the 7th, 14th and 21d to observe the inflammatory reaction of the muscle. On the 14th day, flow cytometry was taken to observe the changes of CD4+ and CD8+T lymphocyte content in blood and tissues, and the cytotoxicity was detected by CCK-8 method. Bone defect model was established in the rat femoral condyle, and the bone formation was observed by Xray and CT after implantation of ACBM/CPC/CSH group (material group), calcium phosphate/calcium sulfate hemihydrate (CPC/CSH) group (control group) and no material implantation (blank group). [Results] The extract did not cause acute or subacute toxicity in mice, and there was no significant difference in body weight (P>0.05). In the pyrogen test, the body temperature remained normal. Skin erythema did not appear within 72 h after epidermal injection. In the muscle implantation experiment, there was slight inflammation in the muscle tissue on the 7th day, and the inflammation disappeared on the 21st day. As results of flow cytometry on the 14th day, there were no significant differences among the three groups in terms of the peripheral blood lymphocyte content of CD4+ [(9.6±1.8) vs (10.1±1.2) vs (10.7± 1.4), P=0.470], CD8+ [(9.5±1.1) vs (10.3±1.8) vs (10.5±1.7), P=0.249], as well as the spleen CD4+ [(18.1±1.5) vs (17.2±7.3) vs (17.5±1.0), P= 0.195], and CD8+ [(8.8±7.2) vs (7.7±7.6) vs (7.8±7.2), P=0.359]. The X-ray and Micro-CT images showed that bone defects in all groups were repaired to different degrees, and ACBM/CPC/CSH bone materials had the best repair effect. [Conclusion] The ACBM/CPC/CSH composite artificial bone material has good biocompatibility and in vivo osteogenic properties, which is beneficial to the repair of irregular bone defects