Abstract 21 High-Throughput Blood Separation System for CD34+ Cell Isolation

Abstract

IntroductionResearch into the therapeutic roles and clinical potential of CD34+ cells is growing [Weissman and Shizuru, Blood. 2008;112(9):3543-3553]. Accordingly, advances in cell isolation methods have risen to meet the research demand. Cord blood banks have a unique opportunity to source CD34+ cells because of their relatively high prevalence in cord blood [Dauber et al. Cytotherapy. 2011;13(4):449-458]. However, current isolation systems are not optimized for processing cord blood, and even blood-compatible systems typically require the blood to be processed before use. FerroBio (www.42bio.com) has developed a magnetic separation system that facilitates scalable, high-volume cell isolation from complex biological fluids, such as blood, without pre-processing. This technology is now being adapted for CD34+ cell isolation from cord blood.ObjectiveIn this study, the conditions in which magnetic beads efficiently bind target cells were explored. The objective of this work was to evaluate optimal blood mixing conditions to bring beads into contact with cells in suspension in the blood. This will allow unmodified cord blood units (CBUs) to flow directly into our apparatus.MethodsThe FerroBio separation process is comprised of three simple steps (Figure 1A). Here, conditions of step 1 were explored to maximize target cell recovery and viability. Variables included type of agitation equipment (Figure 1B), temperature, time, and relative air volume in the blood bag. Recovery and viability were measured via flow cytometry and calculated relative to unprocessed blood from the same CBU.ResultsTwo mixing conditions were identified for maximum recovery of CD34+ cells (Figure 1C, blue bars). While not statistically different from each other, the Drum Roller method only requires one piece of standard equipment, is gentler, and can accommodate multiple CBUs simultaneously. No statistical difference in viability was observed in any conditions (90.7% ± 16.3%, n = 9). Increasing incubation time and the relative amount of air in the bag did not increase recovery (not shown).Figure 1. (A): Overview of the FerroBio Magnetic Separation Process, highlighting step (1) in which magnetic beads are mixed within the CBU to bind the target CD34+ cells. (B): Illustrations of the agitation equipment used to mix the CBU during step (1). (C): The resulting CD34+ recovery (1-way ANOVA, P = .0245). Image credit for panel (A): BioRender.DiscussionThe FerroBio separator can streamline CD34+ cell isolation by processing high blood volumes in a fast, closed system with high recovery rates. Additional studies are under way to evaluate the functionality of the isolated CD34+ cells and efficiency of bead removal.