Knowledge Bank

Red blood cell (RBC) units that are stored ex vivo hypothermically will degrade over time. Also known as RBC storage lesion, the RBC unit accumulates cell-free hemoglobin (Hb) over its 42-day storage limit set by the US Food and Drug Administration (FDA). Once the RBC unit containing cell-free Hb is transfused into a patient, the Hb will scavenge nitric oxide, causing vasoconstriction and systemic hypertension. The iron in the cell-free Hb will also cause oxidative tissue damage. We examined optimizing the removal of cellular waste using tangential flow filtration (TFF). The TFF RBC washing system was run under room temperature conditions at 25 oC in a biosafety cabinet. Constant system volume was maintained with a wash solution of 0.9 wt% saline and 0.2 wt% dextrose, and the blood was pumped throughout the system with either a centrifugal or peristaltic pump. 13 separate and unexpired (≤42 days old) RBC units were used in the RBC washing process (10 with peristaltic, 3 with centrifugal). UV-visible absorbance spectroscopy and Winterbourn equations were used to quantify cell-free Hb. Hemox Analyzer, Coulter counter, and Brookfield rheometer were used to analyze pre- and post-wash RBC samples. Standard hematocrit (HCT) analysis was used to determine RBC volume fraction in solution throughout the wash process. There was a significant difference in HCT, but not for specific pairs of diacycles during the wash process when the peristaltic pump was used. However, there was no significant change in red blood cell count after the wash process with the peristaltic pump. In terms of cell-free Hb in the washed blood, there was a significant difference, but not between specific pairs of diacycles when using the peristaltic pump throughout the wash process. Between the initial and final (6x) diacycle, the oxygen equilibrium curve P¬50 had a significant decrease, and the related cooperativity had a significant increase when the peristaltic pump was used. As for the centrifugal pump, there was no significant change in HCT throughout the wash process. Similarly, the cell count did not significantly change for the process with the centrifugal pump. There was no significant difference for P¬50 and cooperativity values for the centrifugal pump blood wash process between the initial and final diacycles. Overall, this study attempted to optimize the use of TFF for washing RBC units. With a goal of removing cell-free Hb, we introduced the use of dextrose in the wash solution, a peristaltic pump, and a change in system temperature when compared to previous TFF facilitated RBC washing experiments. We determined that the use of a peristaltic pump is a poor choice to drive the blood wash process.