BACKGROUND: Membrane-associated carbohydrate changes act as signals for removal of senescent and damaged red blood cells (RBCs) from the circulation and could play a role in the RBC storage lesion and RBC survival after transfusion. In this study, a panel of lectins was used to investigate the expression of carbohydrates on RBCs that had been separated before storage into young and old RBCs. STUDY DESIGN AND METHODS: Leukodepleted RBCs were separated before storage into young and old RBCs (n = 9 paired units) by centrifugation and sampled at nominated time points during 42 days of storage. Changes to carbohydrate expression at the RBC membrane during storage were determined by flow cytometry with a panel of fluorescein-labeled lectins. RESULTS: Old RBCs showed lower fluorescence intensity throughout storage, suggesting reduced binding of lectins compared to young RBCs. Progressively increased binding of lectins specific for galactose and N-acetylglucosamine residues was observed during storage of young and old RBCs. CONCLUSION: Changes to lectin binding during storage of RBCs suggest that significant changes occur to the carbohydrate structures at the RBC membrane. These findings provide further insight into the mechanisms of the RBC storage lesion and potential influence on RBC survival after transfusion.