A Novel System for the Quantification of the ADCC Activity of Therapeutic Antibodies

Figure 9

Quantification of the ADCC activity of rituximab using iLite effector cells and CD20⁺ Raji target cells in the presence of normal human serum. Frozen RTU iLite effector cells (1.2 × 10⁵ cells/well), CD20⁻, and CD20⁺ target cells were thawed rapidly, and the effector cells were mixed with the CD20⁻ or CD20⁺ target cells at an E:T ratio of 3 : 1 and incubated for 4 hours at 37°C in the presence of RPMI 1640 medium +10% FBS either alone or together with a 1/20 final dilution of samples of human serum from normal donors in the presence of increasing concentrations of rituximab prior to the addition of Nano-Glo Dual-luciferase reagent (Promega, Madison, WI) and the sequential determination of FL and NL activity. Results are expressed as fold induction relative to the control sample without rituximab. (a) Fold induction and (b) fold induction following subtraction of the values obtained with the iLite effector cells and the CD20⁻ target cells as described in Materials and Methods.

(a)

(b)