BACKGROUND: The elucidation of the storage lesion is important for the improvement of red blood cell (RBC) storage. Ex vivo storage is also a model system for studying cell-signaling events in the senescence and programmed cell death of RBCs. The membrane hosts critical steps in these mechanisms and undergoes widespread remodeling over the storage period. STUDY DESIGN AND METHODS: Fresh and CPDA-stored RBCs from 21 blood donors were evaluated as whole cells, membrane ghosts, and cytoskeletons by sodium dodecyl sulfate–polyacrylamide gel electrophoresis, immunoblotting, immunofluorescence microscopy, and in situ assays. Band 3 content, immunoglobulin G (IgG) content, specific protein movement to and from the membrane, and caspase system activation were measured. RESULTS: During storage, Band 3 protein was aggregated and its content decreased as did the content of several lipid raft–related proteins. IgG binding to the membrane increased. Sorcin and synexin moved from the cytosol to the membrane, stomatin and flotillins left the membrane, the Fas protein was oligomerized, and caspase was activated. CONCLUSION: The remodeling of the RBC membrane during storage includes loss and oxidative cross-linking of Band 3 as well as IgG binding. This process occurs with lipid raft development and loss and is probably driven by caspase activation. Oxidative injury appears to be an important driver of RBC aging during storage.