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Haemolysis mediated by anti-D monoclonal antibodies in direct and cold target competition ADCC assays
T Bakács1, B M Kumpel, G Tusnády
1National Institute of Oncology, Budapest, Hungary.
Thirteen IgG anti-D human monoclonal antibodies (mAbs) were compared for their ability to mediate lysis of D-positive erythrocytes by PBMC in direct and cold target competition antibody-dependent cell-mediated cytotoxicity (ADCC) assays. In the latter assay, lysis of fluid-phase anti-D-sensitised O Rh D-positive papainised erythrocytes (E-IgG) was inhibited by A (or B) Rh D-negative papainised erythrocytes sensitised by fluid-phase anti-A (or anti-B) mAbs. The competitive and lytic activities of the anti-D mAbs were characterised by the equilibrium dilution (ED) values, which were the reciprocal of the dilution of anti-A (or anti-B) at which lysis of target E-IgG and competitor E-IgG were identical. There was a poor correlation between the number of erythrocyte-bound anti-D molecules and either the sensitivity of E-IgG anti-D to haemolysis in the direct ADCC assay, or to the ED values of the mAbs obtained in the cold target competition ADCC. The discriminatory power of the cold target competition ADCC was better than than of the direct ADCC to detect differences in the lytic potential of the anti-D mAbs.
Thirteen IgG anti-D human monoclonal antibodies (mAbs) were compared for their ability to mediate lysis of D-positive erythrocytes by PBMC in direct and cold target competition antibody-dependent cell-mediated cytotoxicity (ADCC) assays. In the latter assay, lysis of fluid-phase anti-D-sensitised O Rh D-positive papainised erythrocytes (E-IgG) was inhibited by A (or B) Rh D-negative papainised erythrocytes sensitised by fluid-phase anti-A (or anti-B) mAbs. The competitive and lytic activities of the anti-D mAbs were characterised by the equilibrium dilution (ED) values, which were the reciprocal of the dilution of anti-A (or anti-B) at which lysis of target E-IgG and competitor E-IgG were identical. There was a poor correlation between the number of erythrocyte-bound anti-D molecules and either the sensitivity of E-IgG anti-D to haemolysis in the direct ADCC assay, or to the ED values of the mAbs obtained in the cold target competition ADCC. The discriminatory power of the cold target competition ADCC was better than than of the direct ADCC to detect differences in the lytic potential of the anti-D mAbs.