DEVELOPMENT OF INTEGRATED BEAD AND FIBER ANTIBODY FILTERS FOR SPECIFIC REMOVAL OF ANTI-A ANTIBODIES FROM BLOOD

摘要

Removal of anti-A/B antibodies from blood in the peri-transplantation period eliminates the risk of hyperacute rejection in ABO-incompatible transplantation. We are developing anti-A/B immunoadsorption devices, compatible with whole blood perfusion. In this study we developed a new antibody filtering device based on integrated microfiltration fibers with antibody capturing beads distributed within the interstitial fiber space (BSAF device). As blood flows through the fibers in BSAF devices, Starling flow carries plasma from the inner fiber lumen to the beads in the shell compartment where antibodies diffuse and bind to covalently attached antigens within the shell-side porous beads. We first investigated the possibility of using synthetic blood group A/B-trisaccharide epitopes, conjugated with poly-N hydroxyethylacrylamide spacers, as the immunoadsorbent material in our antibody filtering devices. The glycopolymers were equipped with biotin tags and deposited on streptavidin-coated sensor chips. Antibody removal capacity per unit surface area and kinetics of antibody binding to immobilized glycoconjugates were quantified using surface plasmon resonance. We then developed a simple mathematical model to guide the choice of key design and operational parameters for a clinical BSAF device. The model demonstrated that for a given flow rate and reservoir volume, antibody removal rate in a BSAF was dependent on the magnitude of a lumped dimensionless parameter which characterized the ratio of antibody uptake rate by the beads to the Starling flow rate in the device. The highest antibody removal rate was predicted for a perfusion limited regime. Once this maximum limit was obtained, any further increase in the antibody removal rate was only possible by increasing the flow rate in the device. Key model predictions were validated in a series of in vitro monoclonal anti-A antibody capture studies in BSAF devices packed with anti-A specific beads. Once validated, we used the model to design a BSAF device that would generate a clinically relevant rate of anti-A removal. We fabricated and tested scaled down prototypes of the "clinical" BSAF device and showed significant reduction in IgM and IgG anti-A antibody titers within two hours of whole blood perfusion through our fabricated BSAF devices.