Background: A mesoporous bioactive glass (MBG) with a highly specific surface area and well-ordered pores is a suitable material to carry antibiotics. Fibrin and mesenchymal stem cells are used in promoting tissue repair and osteogenesis, respectively. Purposes: Tobramycin-loaded mesoporous bioactive glass (TMBG), fibrin and mesenchymal stem cells are used in this study to promote osteogenesis and against bacterial contamination in vitro. Methods: MBG was synthesized in a two-step acid-catalyzed self-assembly process with hydrothermal treatment. Tobramycin was loaded with MBG and fibrin to make a TMBG fibrin complex. The cell proliferation, osteogenic differentiation, Live/Dead staining and antibiotics inhibition zone test were evaluated for TMBG fibrin complex. Results: The experimental results showed a well-ordered hexagonally structured bioactive glass was synthesized. The TMBG fibrin complex cultivated with rabbit bone marrow mesenchymal stem cells (RBMSCs) showed good biocompatibility and increased expression of collagen type I, OPN, OCN and RUNX2 in cultivation period. The inhibition zone of TMBG fibrin complex increased while the concentration of tobramycin increased against Staphylococcus aureus and Pseudomonas aeruginosa. In addition, the TMBG fibrin complex (with 0.5% tobramycin) had the same antibacterial effect as 10 mg tobramycin. Conclusions: Based on the results mentioned above, the TMBG fibrin complex showed antibacterial effect and exhibited great biocompatibility a potential biomaterial for bone tissue engineering.