Factorial design optimization of red blood cell PEGylation with a low molecular weight polymer
A proposed method to overcome the donor-recipient blood group incompatibility is to mask the blood group antigens by the covalent attachment of poly(ethylene glycol) (PEG) to the red blood cell (RBC) membrane. Despite much work in the development of PEG-coating of RBCs, there is a paucity of data on the optimization of the PEG-coating technique by using low molecular weight polymers. It is the aim of this study to determine the optimum conditions for PEG-coating using a cyanuric chloride reactive methoxy-PEG derivative as a model polymer. Activated PEG of 2 kDa molecular mass was covalently attached to human RBCs under various reaction conditions. Inhibition of binding of a blood-type specific antiserum (anti-D) was employed to evaluate the effect of the PEG-coating, quantified by flow cytometry. RBC morphology was examined by scanning electron microscopy (SEM). Statistical analysis of experimental design together with SEM results showed that the optimum PEGylation conditions are: temperature 14 degrees C, reaction time 20 min, and concentration of reactive PEG 10 mg/mL.