Study design and hemoglobin levels. (A) On day 1, each human volunteer donated 2 autologous RBC units by apheresis. One unit was transfused into the same participant after 3 to 7 days of storage (ie, “fresh”); the other unit was transfused after 40 to 42 days of storage (ie, “older”). One whole blood phlebotomy was performed 3 to 7 days before the older RBC transfusion to prevent post-transfusion erythrocytosis and to control for effects of recent blood loss on laboratory parameters. Blood samples were collected 90 minutes before transfusion and at 0, 1, 2, 4, 24, and 72 hours after transfusion. (B) Data are mean ± SEM for hemoglobin levels from before transfusion to 72 hours after transfusion of either fresh or older RBCs. The P value is as specified in the figure comparing the paired area under the curve of the mean hemoglobin levels for the N = 14 volunteers from 0 to 24 hours after the fresh and older RBC transfusions. (C) The individual hemoglobin levels for each subject up to 24 hours after transfusion. Vertical arrows indicate pretransfusion time points; and horizontal dashed lines, reference range values for men (blue) and women (pink).