Investigation of Layer-by-Layer Assembly of Polyelectrolytes on Fully Functional Human Red Blood Cells in Suspension for Attenuated Immune Response

The encapsulation of live cells with polymeric coatings is a versatile approach to modulate or control the response cells to their environment. The layer-by-layer (LbL) self-assembly of nonimmunogenic polyelectrolytes is employed here to attenuate or suppress the binding of antibodies to live red blood cells (RBCs) and, consequently, decrease their inherent immunogenicity toward foreign RBCs. The optimized shell was composed of four bilayers of alginate (AL) and chitosan-graft-phosphorylcholine (CH-PC) surrounded by two bilayers of AL and poly-L-lysine-graft-polyethylene glycol (PLL-PEG). Experimental parameters, including the polyelectrolytes and RBCs concentrations and the cell handling and purification protocols, were optimized to achieve effective encapsulation of live and functional RBCs in suspension. The viability and functionality of coated RBCs were confirmed by a hemolysis assay and by their ability to take up oxygen. The successful immunocamouflage of RBCs was confirmed by observing that the recognition of the ABO/D (Rh) blood group antigens present on the surface of RBCs by their respective antibodies was muted in the cue of coated RBCs. The results of this studies mark an important step toward the production of universal RBCs.