- Background: In contrast to the relative scarcity of donor kidneys and hearts, the potential supply of deceased donor pancreata is exceeding the demand. However, this organ surplus is not being fully realized because, in current transplantation practice, the duration of pancreas storage before transplantation is limited to 8–10 hours due to the extreme vulnerability of pancreatic tissue to anaerobic damage caused by preservation. Objectives: To reduce cold ischemic injury in order to increase the utilization of donor pancreases in Israel for whole-organ and cell transplantation. Methods: We evaluated a novel two-layer preservation oxygenated cold storage method that uses perfluorocarbon to continuously supply oxygen to the pancreas during preservation in conventional University of Wisconsin solution. Results: Pancreatic tissue morphology, viability and adenosine-triphosphate content were serially examined during preservation of the pig pancreas for 24 hours either by a two-layer or by conventional simple cold storage. Already after 12 hours of storage, the superiority of the two-layer method over the University of Wisconsin method was apparent. Starting at this time point and continuing throughout the 24 hours of preservation, the tissue architecture, mitochondrial integrity, cellular viability and ATP tissue concentration were improved in samples preserved in oxygenated UW/PFC as compared to controls stored in conventional UW solution alone. Conclusions: The UW/PFC two-layer preservation method allowed tissue ATP synthesis and amelioration of cold ischemic tissue damage during extended 24 hour pancreas preservation. This method could be implemented in clinical practice to maximize utilization of pancreata for whole-organ and islet transplantation as well as for pancreas sharing with remote centers.