BACKGROUND: A study was done to assess the quality of RBCs stored at 4 degreesC in AS-1, AS-3, or AS-5 for 42 days before biochemical modification and freezing. STUDY DESIGN AND METHODS: RBCs were stored at 4 degreesC for 42 days in AS-1, AS-3, or AS-5 and then biochemically modified with pyruvate, inosine, phosphate, and adenine solution (Rejuvesol), frozen with 40-percent (wt/vol) glycerol, and stored at -80 degreesC for at least 2 months. The RBCs were deglycerolized by the use of a cell washer (Haemonetics 115), and stored for 24 hours at 4 degreesC in a 0.9-percent sodium chloride and 0.2-percent glucose solution before the autologous transfusion. RESULTS: The mean freeze-thaw-wash recovery process produced RBC recovery values of 85 percent, with the mean 24-hour posttransfusion survival at 75 percent, and the mean index of therapeutic effectiveness at 64 percent for the RBCs stored at 4 degreesC in AS-1, AS-3, or AS-5 for 42 days before biochemical modification and freezing. All the units exhibited normal or slightly higher than normal 2,3 DPG levels after deglycerolization and postwash storage at 4 degreesC for 24 hours. CONCLUSION: RBCs stored in AS-1, AS-3, or AS-5 at 4 degreesC for 42 days and then biochemically modified with pyruvate, inosine, phosphate, and adenine and glycerolized, frozen, washed, and stored at 4 degreesC for 24 hours before autologous transfusion had acceptable in vitro and in vivo measurements.