STUDY OF REHYDRATION INJURY IN LYOPHILIZED RED BLOOD CELLS

Injuries caused during rehydration, including membrane phase transition and volume response, can lead to low recovery rates of freeze-dried red blood cells (RBCs) following rehydration. Thus, rehydration temperature and procedures must be optimized to prevent leakage of intracellular substances from cells and the occurrence of extreme volume response. To examine the phase transition of lyophilized RBC membranes, RBC ghosts were prepared and freeze-dried and their phase transition temperature was determined as 37 degrees C using differential scanning calorimetry. In addition, the membrane phase transitions of freeze-dried RBCs were analyzed at different temperatures. The highest cell recovery rate was obtained at 37 degrees C, which was consistent with the phase transition injury analysis. The volume response injury mechanism was examined using Environmental Scanning Electrical Microscopy (ESEM) with lyophilized cells. Rehydration velocity was controlled by adjusting the relative humidity under ESEM. The results indicate that during the primary stage of rehydration, cell thickness and diameter first increase and then decrease. The response magnitude and potential of cell thickness was greater than those of cell diameter. These results indicate that the control of rehydration conditions is crucial to avoid hemolysis caused by membrane phase transitions and extreme volume response in freeze-dried RBCs. This study suggests that the injury mechanism underlying the rehydration of freeze-dried RBCs is caused by phase transitions and cell volume response. These findings will be instrumental in the preservation of RBCs and other cell types by freeze-drying.