In this study, preparation and characterization of soft crosslinked gelatin sponge for blood hemostasis application was considered. The effects of gelatin and crosslinker concentrations and altering freeze-drying temperature on sponges' density and structure, water absorption ability and biodegradation, cytotoxicity, mechanical properties and hemostatic effect were investigated. The density measurement indicated that the density of freeze-dried sponges increased when the freezing temperature was lowered. Scanning electron microscope and optical microscope images showed that gelatin sponges had uniform small pores (60 μm) after freezing at liquid nitrogen (-196°C). Biodegradation study demonstrated that the crosslinked sponges containing 1% and 2% gelatin lost respectively nearly 40 to 70% of their weight during 24 h. Prepared sponges showed desired water absorption ability (30-40 times of own dry weight) improved by lowering glutaraldehyde concentration. Cell toxicity was not detected in any of the samples. Compression modulus of sponges decreased four times (160 to 40 kPa) as the gelatin content varied from 2 to 1% w/v. Hemostasis study confirmed that the hemolytic ability of sponges increased through raising gelatin content and porosity of sponge. We suggest using gelatin sponges containing 1% w/v gelatin, 0.5% w/v glutaraldehyde frozen in liquid nitrogen, as a potential substitution for local hemostat absorbable sponge.