Currently the only method to assess liver preservation injury is based on liver appearance and donor medical history. Previous work has shown that high-frequency ultrasound could detect ischemic cell death due to changes in cell morphology. In this study, we use high-frequency ultrasound integrated backscatter to assess liver damage in experimental models of liver ischemia. Ultimately, our goal is to predict organ suitability for transplantation using high-frequency imaging and spectral analysis techniques. To examine the effects of liver ischemia at different temperatures, livers from Wistar rats were surgically excised, immersed in phosphate buffer saline and stored at 4 and 20 °C for 24 h. To mimic organ preservation, livers were excised, flushed with University of Wisconsin (UW) solution and stored at 4 °C for 24 h. Preservation injury was simulated by either not flushing livers with UW solution or, before scanning, allowing livers to reach room temperature. Ultrasound images and corresponding radiofrequency data were collected over the ischemic period. No significant increase in integrated backscatter (~2.5 dBr) was measured for the livers prepared using standard preservation conditions. For all other ischemia models, the integrated backscatter increased by 4–9 dBr demonstrating kinetics dependent on storage conditions. The results provide a possible framework for using high-frequency imaging to non-invasively assess liver preservation injury.