Erythrocyte Physiology and the Storage Lesion

Changes during refrigerated storage

A red-cell unit late in storage differs measurably from the same unit on the day it was collected. The red-cell storage lesion is the progressive, partly reversible set of metabolic, structural, and functional changes that develops during storage at 1 to 6 °C. Cold temperature slows metabolism and membrane transport. Glycolysis continues in the closed container, consuming glucose and producing lactate and hydrogen ions. The timing and size of each change depend on the storage system and duration. ATP may rise during the first week and then decline. 2,3-BPG generally falls during conventional storage. Falling pH and substrate depletion impair metabolic enzymes and eventually contribute to loss of ATP and 2,3-BPG.^{1-31 Bloch EM, et al. Technical Manual. 2026.3 Flatt JF, et al. The involvement of cation leaks in the storage lesion of red… 2014.}

Physiology behind storage changes

Membrane and cation control

The biconcave red cell has enough membrane surface to deform through small vessels and splenic slits. Integral proteins such as band 3 and the glycophorins carry blood-group antigens and transport solutes. Spectrin, actin, ankyrin, and protein 4.1R form or anchor the flexible membrane skeleton. Loss of membrane surface, oxidation, calcium accumulation, and ATP depletion reduce deformability and promote splenic clearance.^{1,21 Bloch EM, et al. Technical Manual. 2026.2 Yoshida T, et al. Red blood cell storage lesion. 2019.}

ATP-dependent pumps normally maintain low intracellular sodium and calcium and high intracellular potassium. Refrigeration slows these pumps, so potassium leaves the red cells and accumulates in the component supernatant while sodium enters. Falling ATP, calcium entry, oxidation, and membrane vesiculation contribute to rigidity and hemolysis. The supernatant potassium increase is most important when a large extracellular volume is transfused rapidly or to a small recipient.^{1,31 Bloch EM, et al. Technical Manual. 2026.3 Flatt JF, et al. The involvement of cation leaks in the storage lesion of red… 2014.}

Metabolic support

Mature erythrocytes lack nuclei and mitochondria, so cytosolic glucose metabolism must supply ATP, reducing power, and 2,3-bisphosphoglycerate. These products connect metabolic decline to membrane failure, oxidation, methemoglobin accumulation, and altered oxygen affinity.

Low ATP impairs cation control and deformability. Low NADPH leaves hemoglobin and membrane proteins vulnerable to oxidation. Failure to reduce methemoglobin lowers functional oxygen-carrying capacity. The terms 2,3-bisphosphoglycerate (2,3-BPG) and 2,3-diphosphoglycerate (2,3-DPG) identify the same molecule.^{1,21 Bloch EM, et al. Technical Manual. 2026.2 Yoshida T, et al. Red blood cell storage lesion. 2019.}

Hemoglobin oxygen affinity

2,3-BPG binds deoxygenated hemoglobin and promotes oxygen release. Cooperative oxygen binding produces the sigmoid oxygen dissociation curve. P50 is the PO₂ at which hemoglobin is 50% saturated; adult hemoglobin has a P50 near 26 to 27 mm Hg under standard conditions. A higher P50 means lower affinity and easier oxygen release, while a lower P50 means higher affinity.^{1,51 Bloch EM, et al. Technical Manual. 2026.5 Jaafar LS, et al. 2,3-Diphosphoglycerate. 2025.}

Refrigerated red cells lose 2,3-BPG, which shifts the curve left. After transfusion, 2,3-BPG synthesis resumes and oxygen affinity moves toward the recipient's physiologic position. Recovery varies with the recipient's acid-base state, phosphate availability, anemia, and illness.^{1,2,51 Bloch EM, et al. Technical Manual. 2026.2 Yoshida T, et al. Red blood cell storage lesion. 2019.5 Jaafar LS, et al. 2,3-Diphosphoglycerate. 2025.}

Product quality and posttransfusion recovery

FDA evaluation of a red-cell storage system includes end-of-storage hemolysis and 24-hour in vivo recovery, the percentage of labeled cells remaining in circulation 24 hours after transfusion. In the FDA decision summary for one 42-day system, the hemolysis plan evaluated whether at least 95% of units remained below 1% hemolysis, and mean 24-hour recovery had to reach at least 75%. The full acceptance plan and labeled conditions are specific to that system.^{66 US Food and Drug Administration. De Novo Classification Request for the HEMANEXT ONE System. 2023.}

Percent hemolysis is the end-of-storage number a quality-control record shows. In vitro hemolysis estimates the proportion of the component's hemoglobin released into the cell-free fluid. The extracellular volume fraction is 1 minus the hematocrit expressed in L/L. Correct a diluted supernatant result for dilution and use the same concentration units for both hemoglobin measurements before calculating:^{77 Canadian Blood Services. Red blood cells.}

hemolysis (%) = 100 × (1 − hematocrit) × supernatant hemoglobin ÷ total component hemoglobin

For example, suppose a fivefold-diluted supernatant measures 36 mg/dL. The undiluted concentration is 180 mg/dL, or 0.180 g/dL. With hematocrit 0.65 L/L and total component hemoglobin 19.5 g/dL, hemolysis is 100 × 0.35 × 0.180 ÷ 19.5 = 0.32%, rounded to two decimal places. Compare the result with the specification for that product and study. This calculation measures hemoglobin outside intact cells only. Posttransfusion survival needs the in vivo recovery study described above.^{77 Canadian Blood Services. Red blood cells.}

After transfusion, membrane-pump activity resumes, and ATP and 2,3-BPG begin to recover. 2,3-BPG synthesis begins within hours, and both ATP and 2,3-BPG have been reported to recover over roughly 7 to 72 hours of circulation in the recipient.^{1,21 Bloch EM, et al. Technical Manual. 2026.2 Yoshida T, et al. Red blood cell storage lesion. 2019.} Cells with irreversible membrane or oxidative injury are cleared, which lowers the recovered circulating fraction. Unit age gives only part of the picture, because donor biology and processing alter the rate of change. Release and selection therefore rest on that approved system, the labeled expiration date, component inspection, quality control, and any recipient-specific procedure.^{44 AABB, et al. Circular of Information for the Use of Human Blood and Blood… 2024.}

The clinical importance of temporary 2,3-BPG depletion and the benefit of restoring 2,3-BPG during cardiac surgery or shock remain uncertain.^{22 Yoshida T, et al. Red blood cell storage lesion. 2019.}