Objectives: Acute kidney injury is experienced by up to 50% of the patients undergoing partial nephrectomy, whereas current approaches are limited to surgical management. To tackle this issue, we developed an implantable biomaterial that can target rapid hemostasis and wound healing at the site of morbidity. Specifically, we developed a porcine kidney-based biomaterial (dkEMS; decellularized kidney extracellular matrix-based sponge) and tested for its clinical efficacy in vitro and in vivo. Methods: First, kidney decellularized extracellular matrix (kdECM) was prepared by obtaining fresh, healthy porcine kidneys provided by a veterinarian from the National Institute of Animal Science. Prepared kidneys were then frozen and sliced into 0.6mm~0.8mm thickness, later to be washed. After washing, the tissues were freeze-dried and dissolved into a solution. To prepare dkEMS, the kdECM solution was mixed into different concentrations of 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) mixture and frozen at -20°C to promote cross-linking of the collagen fibers and formation of ice particles. The ice crystals were removed during freeze-drying process, upon which became empty pores in its dry state dkEMS. The prepared dkEMS was characterized for its material properties and biocompatibility. Finally, it was implanted in acute kidney injury rat model, and histological analyses were made until day 56. Results: The degree of cross-linking of the kdEMS at varying EDC/NHS concentrations were measured to be around 38%, 58%, and 81% for 12mM/3mM, 24mM/6mM, 48mM/12mM EDC/NHS respectively. In addition, 12mM/3mM kdEMS showed highest biocompatibility rate. When compared with gelatin sponge, kdEMS showed similar or higher hemocompatibility, and its 56-day follow-up showed a histological result that resembles closely to the native kidney. Conclusions: kdEMS was prepared and analyzed for its physical/chemical properties, biocompatibility, and hemocompatibility. Material analysis showed the successful preparation of kdEMS, and histological results in rat kidney showed its favorable ability in tissue regeneration and recovery after acute kidney injury model.