Phosphate-buffered sucrose (PBSc) solution is effective for short-term hypothermic preservation of tissue during feline kidney transplantation. A high-molecular-weight polyethylene glycol (35,000 Da, PEG35) reportedly enhanced the protective effects against cold-induced tubular injuries in animal kidney transplantation models. We investigated the ability of PBSc solution containing PEG35 to preserve cultured feline kidney cells using in vitro WST-8 cell proliferation assays. PEG35 significantly improved cell viability during 24 hr of cold preservation. PBSc containing 20 g/l PEG35 achieved an effect almost equal to that of University of Wisconsin (UW) solution, the gold standard preservation solution used in human clinical kidney transplantation, for up to 24 hr of preservation. Our results suggest that PBSc containing PEG35 provides an excellent medium for graft cold storage during feline kidney transplantation.