Cryopreservation of rare pediatric red blood cells for support following bone marrow transplant.

BACKGROUND: A 2-year-old, 10.8 kg male pediatric patient with X-linked chronic granulomatous disease (CGD) with McLeod syndrome (MLS) was scheduled for a hematopoietic stem cell transplant (HSCT). Identification of allogenic red blood cells (RBC) for post-transplant support was unsuccessful prompting the development of a customized method to collect and freeze rare autologous pediatric cells. STUDY DESIGN AND METHODS: A protocol was developed for the collection of small volume pediatric whole blood (WB) via peripheral venipuncture with collection into 10 ml syringes containing anticoagulants. Additionally, a closed system RBC glycerolization and deglycerolization instrument was adapted to process small volume, non-leukoreduced WB. Both collection and WB processes were validated. In total 4 approximately 100 ml autologous units were collected and frozen. Two units were thawed, deglycerolized, and used for clinical transfusion support. To appreciate processing impacts on RBC rigidity, ektacytometry was performed on pre-processed and post-deglycerolization samples. RESULTS: Free hemoglobin (HGB) of validation units after thawing/deglycerolization was <150 mg/dL with an average red cell recovery of 85%. These units also showed little difference between pre-and post-processing Lorrca deformability curves or membrane rigidity. Two pediatric units were thawed and deglycerolized for transfusion. Free HGB was 70 mg/dL and 50 mg/dL post-thaw, and these RBCs had a slight decrease in deformability and increased membrane rigidity. DISCUSSION: Customized WB collection, glycerolization, freezing, and deglycerolization processes were developed to successfully support a pediatric patient with CGD and MLS after autologous HSCT. Both pediatric units showed increased membrane rigidity post-deglycerolization which may be a consequence of the CGD and MLS genetic background.

A comparison of different methods of red blood cell leukoreduction and additive solutions on the accumulation of neutrophil-priming activity during storage.

BACKGROUND: Three methods of leukoreduction (LR) are used worldwide: filtration, buffy coat removal (BCR), and a combination of the previous two methods. Additionally, there are a number of additive solutions (ASs) used to preserve red blood cell (RBC) function throughout storage. During RBC storage, proinflammatory activity accumulates; thus, we hypothesize that both the method of LR and the AS affect the accumulation of proinflammatory activity. STUDY DESIGN AND METHODS: Ten units of whole blood were drawn from healthy donors, the RBC units were isolated, divided in half by weight, and leukoreduced by: 1) BCR, 2) filtration, or 3) BCR and filtration (combination-LR); stored in bags containing AS-3 per AABB criteria; and sampled weekly. The supernatants were isolated and frozen (-80°C). RBC units drawn from healthy donors into AS-1-, AS-3-, or AS-5-containing bags were also stored and sampled weekly, and the supernatants were isolated and frozen. The supernatants were assayed for neutrophil (PMN)-priming activity and underwent proteomic analyses. RESULTS: Filtration and combination LR decreased priming activity accumulation versus buffy coat LR, although the accumulation of priming activity was not different during storage. Combination LR increased hemolysis versus filtration via proteomic analysis. Priming activity from AS-3 units was significant later in storage versus AS-1- or AS-5-stored units. CONCLUSIONS: Although both filtration and combination LR decrease the accumulation of proinflammatory activity versus buffy coat LR, combination LR is not more advantageous over filtration, has increased costs, and may cause increased hemolysis. In addition, AS-3 decreases the early accumulation of PMN-priming activity during storage versus AS-1 or AS-5.