In this study, the composite of Collagen-Hydroxyapitite (COL-HA) with microfibrillated cellulose (MFC) was developed as a new bone substitute material. COL-HA was prepared by in-situ method and modified by dehydrothermal treatment. Microfibrillated cellulose (MFC), a nature polysaccharide with plenty of hydroxyl groups, was incorporated into COL-HA composites to improve the properties. The novel COL-HA-MFC scaffold with different ratios of COL-HA and MFC were fabricated by cold isostatic pressing technique and freeze-drying technology. During the forming process, a three-dimensional bone-like structure was shaped in hybrid scaffolds. The microstructural transitions of COL-HA-MFC composites were examined by Fourier transform infrared spectroscope (FTIR), Ultraviolet-visible spectrophotometer (UV), and X-ray diffraction (XRD), which indicated that HA deposited on collagen molecules and MFC bonded with COL-HA. Hydrophilicity, swelling property, mechanical property, and degradability of COL-HA-MFC composites were investigated. Biological properties, such as cytotoxicity and hemolysis, were also studied. The results showed a good swelling capacity for the scaffolds, keeping their original shapes after swelling. The compression strength and degradability of the scaffold materials could be regulated by the MFC content. The compression strength of COL-HA-MFC composite scaffords increased to 20-40 MPa, closing to that of the nature bone (1-200 MPa). The obtained scaffolds are good in biocompatibility with high level of cell growth rate (>70%) and suitable hemolysis rate (≦5%). The work might provide an efficient and alternative approach for collagen-based biomaterials with necessary properties. The COL-HA-MFC composite scaffold showed a potential application in bone tissue engineering.