A New Centrifugation-Free Approach for Reducing IgA Levels in Units of Stored Red Blood Cells

Introduction: Washed red blood cells (RBCs) are indicated for immunoglobulin A (IgA) deficient recipients when RBCs from IgA-deficient donors are not available. Centrifugation-based cell processors commonly used by hospital blood banks cannot consistently reduce IgA to levels suitable for IgA-deficient recipients in a single washing cycle, hence double washing is frequently required. Additionally, cell processors are designed to wash one RBC unit at a time, making washing several units for a multiple-unit transfusion a laborious, lengthy and logistically difficult process. Importantly, these conventional cell washers are complex, expensive machines that may not be available in smaller medical facilities, thus limiting availability of washed RBCs on demand. To address these limitations, we developed a simple, portable, disposable system capable of washing stored RBC units at practical volumetric throughput without the use of centrifugation, while reducing IgA below 0.05 mg/dL in a single run. This new washing system consists of two core modules: (i) a coil of narrow-bore plastic tubing with three sequential bifurcations for high-throughput separation of washed RBCs from washing solution, and (ii) a highly-engineered membrane-hydrogel composite for concentrating the suspension of washed RBCs to the desired level of hematocrit. Materials and Methods: Several units of stored RBCs (n = 4, leukoreduced, 4-6 weeks storage duration) were purchased from the Gulf Coast Regional Blood Center (Houston, TX). Samples of stored RBCs were withdrawn from the units and diluted with 0.9% saline to a hematocrit (Hct) of 10%, to match the dilution typically used in the conventional centrifugation-based washing procedure (2000 mL of saline per typical 350 mL RBC unit with a 60-65% initial Hct). The diluted RBC samples were then washed either by our new washing system, or via conventional centrifugation. The efficiency of the two washing methods was quantified and compared by measuring several key in vitro quality metrics. Results and Discussion: Stored RBCs were diluted to 10% Hct with normal saline and passed through the new washing system at 5.0 mL/min to produce washed RBCs with output Hct of about 42%, and Hct of washing solution (waste) of about 2%. Overall, washing performed by either of the two methods substantially reduced hemolysis, lowered the concentration of free hemoglobin (by >4-fold) and total free protein (by >10-fold), while keeping the intracellular ATP unchanged (Table 1). Washing increased the fraction of well-preserved cells (discocytes, echinocytes 1) from 62.9 ± 14.3% in the initial RBC units to 74.1 ± 6.9% for our experimental device, which was a more pronounced improvement than for conventional centrifugation (67.8 ± 5.1%), although the difference was not statistically significant. Importantly, our new washing system reduced the level of IgA to well below 0.05 mg/dL required by AABB in a single pass, significantly outperforming the traditional, centrifugation-based washing procedure. Conclusions: This study demonstrated the feasibility of washing stored RBCs in normal saline for transfusion to patients with selective IgA deficiency without the use of centrifugation. Our inexpensive, portable, disposal system was significantly better than centrifugation in reducing the IgA level, and performed on par with the conventional procedure for all other key in vitro metrics of RBC quality. The ability of this new washing system to operate in line with existing infusion equipment at bedside could be particularly attractive in remote or resource-limited settings, when conventional centrifugation-based cell washers are unavailable. Acknowledgements: This work was supported by the 2012 NIH Director9s Transformative Research Award (R01HL117329). Disclosures Shevkoplyas: Halcyon Biomedical Incorporated: Employment, Equity Ownership, Patents & Royalties: U.S. Patent Appl. 61/929,357, Research Funding; New Health Sciences, Inc.: Consultancy, Research Funding.