*4.2.4. Two-layer method*

Great focus has remained on improving the quality of pancreas transport to the islet transplant centers, including novel ways to provide oxygen rich media to the graft whilst in cold storage during shipping. In late 1988 Kuroda et al. was the first to report the use of the Two-Layer Method (TLM) for shipping of the pancreas prior to islet cell isolation [86]. The TLM uses a perfluorochemical (PFC) and the organ perfusion fluid; initially Euro-Collins' solution was used but was replaced by UW solution. The benefits of the use of the PFC are due to it being a biologically inert liquid that acts as an oxygen-supplying media. A pancreas preserved using the TLM is theoretically oxygenated through the PFC and substrates are supplied by the UW solution. This allows the pancreas preserved using the TLM to generate adenosine triphosphate during storage, prolonging the preservation time [87]. Strong debate still remains over its benefits, if any, when compared to the use of UW solution during CS [88, 89] and a recent publication of guidelines recommended against the use of the TLM for preservation of the pancreas preceding islet isolation [85].
