Two-Layer Method (TLM) preservation consists of a storage chamber containing a layer of highly oxygenated, water-immiscible liquid perfluorocarbon (perfluorodecalin, C10F18) surmounted by a layer of conventional UW (or similar) organ preservation solution. The perfluorocarbon is sufficiently dense (~2 g/ml) that the pancreas floats on top of it, in contact with both layers. The perfluorocarbon combines low toxicity with an oxygen content 75 times greater than the UW solution used in standard pancreas storage. When preserved under these conditions, the pancreas absorbs oxygen by diffusion and steadily consumes it, supporting sufficient aerobic metabolism to maintain tissue ATP concentrations at near-physiologic levels and prevent, or even reverse, pancreas anoxic injury. In animal models of pancreas ischemic and storage injury, TLM preservation has been strikingly successful at improving the outcome of both islet isolation and pancreas transplantation.