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A novel hybrid biomaterial composed of calcium sulfate (CS) and gelatin (GEL) was prepared with the potential of being used as bone filler or scaffold owing to its osteoconduction. Such composite biomaterial, cross-linked or un-cross-linked, could provide a suitable absorbing rate and prevent the CS crystals migrating from the implant for tissue engineering. The structure of the composite was analyzed with infrared (IR) spectroscopy, X-ray diffraction (XRD) and scanning electron microscopy (SEM). The results indicated that the crystal pattern of CS was affected by the addition of GEL. The GEL part affected the development of the CS dihydrate (CSD) crystal by slowing the conversion from CS hemihydrate (CSH) to CSD; thus, the composite actually contained CSD, CSH and GEL. The compressive strength of the CS/CLGEL composite was also investigated. The compressive strength was correlated to the weight proportions of CS in the CS/cross-linked GEL (CS/CLGEL) composite, and the highest compressive strength of 82 MPa was obtained for the composite containing 40 wt% CS. The in vitro absorption test and the SEM results showed that a porous scaffold was formed in situ with the absorption of CS in the CS/CLGEL composite in a certain time. Therefore, the CS/CLGEL composite material can be used as an in situ porous scaffold with a high initial mechanical strength, and the remaining porous GEL scaffold will enable further in-growth of cells. Human osteoblasts were cultured in contact with the CS/CLGEL composite and the primary results suggested that human osteoblasts could attach and spread on the surface of CS/CLGEL films. The preliminary animal model experiment was operated for assessing the potential of the CS/CLGEL composite as a biodegradable bone substitute. The primary results showed that the CS/CLGEL composite filler could promote new bone in-growth, which will stimulate further study. [ABSTRACT FROM AUTHOR]