Rationale: Acute lung injury (ALI) that develops six hours after transfusion (TRALI) is the leading cause of transfusion-related mortality. Several transfusion characteristics have been postulated as risk factors for TRALI, but the evidence is limited to retrospective studies. Objective: To compare patient and transfusion risk factors between the patients who do and do not develop ALI. Methods: In this prospective cohort study, consecutive transfused critically ill patients were closely observed for development of ALI. Donor samples were collected from the transfusion bags. Risk factors were compared between patients who did develop ALI following transfusion and transfused controls, matched by age, gender and admission diagnosis. Measurements and main results: Seventy-four out of 901 transfused patients developed ALI within 6 hours after transfusion (8%). Compared to transfused controls, ALI patients were more likely to have sepsis (37% vs 22%, p=0.016), and a history of chronic alcohol abuse (37% vs 18%, p=0.006). When adjusted for patient characteristics, transfusion of plasma from female (odds ratio-OR 5.09, 95%CI 1.37-18.85), rather than male donors (OR 1.60, 95%CI 0.76 to 3.37), number of pregnancies among the donors (OR 1.19, 95%CI 1.05 to 1.34), number of donor units positive for antigranulocyte (OR 4.85, 95%CI, 1.32-17.86) and HLA class II antibodies (OR 3.08, 95%CI 1.15-8.25), and the concentration of lysophosphatidylcholine in the donor product (OR 1.69, 95%CI 1.10 to 2.59) were associated with development of ALI. Conclusions: Both patient and transfusion risk factors determine the probability of ALI after transfusion. Transfusion factors represent attractive targets for prevention of ALI