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Silk Fibroin (SF) is a protein polymer with great biocompatibility, which can promote cell proliferation and differentiation, and enhance bone repair. In this paper, the effects of distinctive concentrations of SF solutions on the physicochemical and biological properties of the SF-HA-SA scaffolds were investigated. The SF-HA-SA porous scaffolds were prepared utilizing the pneumatic extrusion 3D printing technique, composed of hydroxyapatite (HA) and different concentrations of SF solution, and sodium alginate (SA) as a binder. The results shown the SF-HA-SA scaffolds can promote cell proliferation with the increase of SF concentration in scaffolds, and the strength meets the necessities of trabecular bone defects of bone and cartilage repair. It provides an important reference for the application of SF in bone tissue engineering.