Current clinical lung preservation techniques have not eliminated ischaemia-reperfusion (I/R) injury, despite many improvements. The optimal combination of flush and storage temperatures remain unclear in lung preservation. This is the first study to investigate a range of temperatures with 24-h inflated storage using consistent state-of-the-art preservation techniques. A rat lung transplant model was used to investigate the optimal combination of flush and storage temperatures. In six groups, rat lungs were flushed at 4 °C, 10 °C or room temperature (F4/F10/FRt) with Perfadex and stored inflated for 24 h in Perfadex on melting ice or at 10 °C (Sice/S10). Left donor lungs were transplanted for analysis. During 2-h reperfusion, the lung graft function was measured (blood gases, maximum ventilation pressure and static compliance) and lung graft injury was also assessed (W/D ratio, total lung protein, Tryptase, Myeloperoxidase). Right donor lungs were assessed for W/D ratio only after flush and storage. For baseline measurements, left lungs without intervention were used. The combination of FRt–Sice showed a significantly higher pO2, lower Pmax, low W/D ratios and total protein levels of left lungs after reperfusion when compared with F4–Sice and baseline. Storage at 10 °C did not improve preservation. We conclude that FRt–Sice creates the best lung graft preservation.