Fig. 6
H3v-47 mAb exhibits ADCC activity. a Cross-linking of FcγRIIIa. Binding curves were obtained by performing ELISA with serial dilutions of each antibody (H3v-47, H3v-12, or control mAb VRC01 [to HIV]) onto HA-coated plates and assessing the ability of HA-bound mAbs to engage both Fc-binding sites on the soluble FcγRIIIa dimer. b Primary NK cell activation. Antibody (H3v-47, H3v-12, or VRC01) at 0.1, 1, or 10 µg/mL each were added independently on 96-well plates coated with purified A/Sydney/5/1997 H3 HA and incubated for 2 h. The plates were washed, and 5 × 105 purified NK cells were added to each well. The cells were stained with anti-human CD107a allophycocyanin-H7 Ab, anti-human CD3 PerCP, anti-human CD56 allophycocyanin, and anti-IFNγ AF700. The data for 20,000–50,000 events were acquired using an LSRFortessa flow cytometer. The percentage of NK cell activation was calculated as the percentage of NK cells that expressed CD107a and/or IFNγ. The dotted lines in both a and b indicate the limit of detection. The results are representative of three independent experiments. Error bars indicate standard error of the mean (SEM)
