Fig. 3

The antibody-mediated killing activities of wild type and Fc mutant G37 and G119 on CAIX expressing tumor cells. (a) Anti-CAIX mAbs (G37 and G119) and the variants (mG37 and mG119) were tested for ADCC activity with human PBMCs in a dose dependent manner. CAIX⁺ SKRC-52 cells were incubated with human PBMCs at a ratio of 1:25 and the indicated concentrations of antibodies, and cytotoxicity measured by LDH release in the culture supernatant. Data represent three independent experiments, each performed in triplicate. (b) CDC activity was determined by culture of rabbit serum with SKRC-52 cells, and cytotoxicity measured as above after 6 h. Data represent three independent experiments, each performed in triplicate. (c) ADCP activity was determined in vitro by flow cytometry, measuring the uptake of fluorescently labeled SKRC-52 cells (PKH) by monocyte derived human macrophages. Data represent the mean percentage of PKH⁺CD16⁺ macrophages from two experiments, as determined by FACS, each experiment performed in triplicate. All data points represent the mean value ± S.E.M.. *, **, and *** represent p value of two-way ANOVA < 0.05, 0.01, and 0.005, respectively (grey asterisks, compared to wild type IgG1; black asterisks, compared to control IgG1)