Rapidly freeze‐dried human red blood cells for pre‐transfusion alloantibody testing reagents

Abstract

Prior to transfusion of red blood cells (RBCs), recipients must be tested for the presence of alloantibodies to avoid immune complications. Liquid‐preserved reagent RBCs with known blood group antigen phenotypes are used for testing. However, these reagents have practical constraints, including limited shelf‐life and require constant refrigeration. To address these issues, we explore the effects of rapid freeze‐drying conditions with trehalose cryoprotectant (0.1–1 M concentrations) on human RBCs and storage of freeze‐dried RBCs (FDRBCs) at room temperature (RT) for up to 12 months. We report that rapid freeze‐drying of RBCs for 2.5 hr with 0.5 M trehalose achieves recoverable cells with near‐normal morphological shape, although size‐reduced. The FDRBCs are metabolically active and functional in antibody‐agglutination tests by the column agglutination test (CAT) for ABO and Rhesus‐D blood group antigens. Expression of the Duffy blood group protein (CD234) decreases by 50% after freeze‐drying RBCs. The initial recovery rate is ≤25%; however, 43% of these FDRBCs are still recoverable after RT storage for 12 months. In this proof‐of‐principle study, we show that rapid freeze‐drying can stabilize RBCs. Further refinements to improve the recovery rate and preservation of antigenic epitopes will make FDRBCs a practical alternative source of reagent RBCs for pre‐transfusion alloantibody identification.

The future of blood services amid a tight balance between the supply and demand of blood products: Perspectives from the ISBT Young Professional Council

“…Lyophilized platelets are, however, not well characterized, with only two Phase 1 dose-escalation studies conducted to date [42,43]. Some studies have also explored the feasibility of producing lyophilized RBCs [44][45][46][47][48], but this product is still in its infancy.…”

Artificial cells for in vivo biomedical applications through red blood cell biomimicry

“…freeze-dried platelets retain some clotting properties) 223 , 224 . For RBCs, the recovery rate has been shown to be less than 25% for pre-transfusion alloantibody screening 225 . These results might be improved by addition of lyoprotectants inside the cells, but this is rather difficult and the loading process might damage the cells.…”

Characterization of freeze-dried oxidized human red blood cells for pre-transfusion testing by synchrotron FTIR microspectroscopy live-cell analysis

“…2 Efforts to develop effective freeze-drying protocols to preserve RBCs together with cryoprotectants have been hindered by poor RBC recovery rates following cell rehydration. 3 In the process of addressing these limitations, we made an interesting discovery that a pre-freeze oxidation step, without the need of a cryoprotectant, appears to stabilize the RBC membrane and maintains intact RBCs throughout the subsequent freeze-drying and rehydration steps (Alves, manuscript in preparation). Although it is known that oxidative stress modifies the chemical composition of the RBC membrane and cytoskeleton leading to rigidification and alteration in cell mechanical properties, 4,5 the effect of oxidation in combination with freeze-drying of RBCs has not been investigated.…”