Surface Treatment of Red Blood Cells with Monomethoxypoly(ethylene glycol) Activated by Succinimidyl Carbonate

In the present study, the optimum conditions were determined for PEGylation of human red blood cells by methoxypoly(ethylene glycol) (mPEG) with molecular mass of 5 kDa, activated by succinimidyl carbonate. Factorial and Taguchi design methods for experiments were used to study the effects of process variables on coating of the cells and determine the optimum conditions for PEGylation. The inhibition of agglutination by a blood-type specific antiserum (anti-D) was employed to evaluate the effect of the polymer coating. The remaining single cells after incubation with anti-D sera were counted using a simple hemocytometer (Improved Neubauer Ruling). The extent of surface coating was evaluated by attachment of cells to FITC labeled-anti-D serum and recording as the fluorescence intensity ratio of FITC-anti-D bound cells of the PEG-RBCs versus control (uncoated) RBCs. The morphology of red blood cells was determined by scanning electron microscopy (SEM) technique. Statistical analysis of experimental design together with SEM results showed that the optimum conditions were pH = 8.7, T=14°C, and t = 60 min while the suitable polymer concentration was found to be 12 mg/mL.