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Limited availability and use of whole blood (WB) following trauma is driven by perceptions that hemostatic function is limited by platelet dysfunction within 5 days storage. We sought to define the hemostatic function of WB stored at 4°C for up to 25 days, elucidate changes in metabolic parameters and mitochondrial dysfunction in platelets in WB, and the effect of supplementation using resveratrol (Res) or cytochrome c (Cyt c).Whole blood was collected, aliquoted, and stored at 4°C without agitation. Resveratrol or Cyt c was supplemented before storage, or 10 days post-storage. Serial samples were collected and analyzed for hemostatic function by platelet mapping thromboelastography. Platelets isolated from WB were counted and mitochondrial function assessed by oxygen consumption, mitochondrial membrane potential, and biochemical parameters.Platelet function of WB was maintained up to 15 days at 4°C before a significant decrease was observed at 25 days. Resveratrol or Cyt c improved WB aggregation potential when supplemented 10 days post-storage. Platelet oxygen consumption was maintained until 10-day storage but significantly decreased thereafter in the absence of change in platelet count. Cytochrome c increased oxygen consumption on Day 15 and platelet mitochondrial membrane potential steadily decreased over time, an effect attenuated by Res or Cyt c supplementation 10 days post-storage. Potassium and lactate levels increased during storage, while pH levels decreased, with no observed effect following Res or Cyt c supplementation.Storing cold WB with Res or Cyt c supplementation enhances ex vivo aggregation by improving platelet function, thereby extending overall storage life. These findings have potential significance for improving WB availability in immediate trauma situations, including treatment in a battlefield trauma setting.Translational study, diagnostic test or criteria, level II.