Prevention of the immune agglutination of methyl acetimidate-reacted sickle erythrocytes by prior reaction with pyridoxal 5'-phosphate.

The appearance of an immune response in some sickle cell anemia patients to reinfused autologous erythrocytes which had been treated with methyl acetimidate (Gabuzda, T. G., Chao, T. L., Berenfeld, M. R., and Gelbart, T. (1980) Blood 56, 1041--1047) was a serious impediment to the clinical application of this reagent as an extracorporeal antisickling agent. The immune reaction was doubtlessly due to acetamidination of membrane protein amino groups. Protection of these amino groups with a reversible reagent prior to treatment of the cells with methyl acetimidate should prevent the antigenic reaction. This result was realized by preequilibration of erythrocytes with pyridoxal 5'-phosphate prior to addition of methyl acetimidate. Subsequent washing of the cells to remove excess reagents and to hydrolyze the pyridoxal 5'-phosphate/membrane protein Schiff base adduct regenerates native protein amino groups of the erythrocyte membrane.