FIGURE 4.
BAG-6686–936 binds to NKp30 in vitro and on NK cells. Co-immunoprecipitation (IP) of NKp30-IgG1-Fc and BAG-6686–936. A and B, E. coli BAG-6686–936 immobilized on beads via polyhistidine tag-specific antibody (A) and High Five insect cell BAG-6686–936 immobilized on StrepTactin-coupled beads (B) were tested for IgG1-Fc fusion protein binding and analyzed by SDS-PAGE and Western blot with anti-human Fc (α-hFc) antibodies. C and D, graded amounts of IgG1-Fc fusion proteins were probed for binding to E. coli BAG-6686–936 (C) and to High Five insect cell BAG-6686–936 (D), and KD values were calculated after fitting of the curves to a 1:1 Langmuir binding model. E and F, binding of E. coli (E) and High Five (F) BAG-6686–936 to NK-92 cells was analyzed via flow cytometry using mouse monoclonal polyhistidine tag-specific or Strep-tag II-specific antibodies prior to detection with APC-conjugated anti-mouse secondary antibodies. Data are representative of at least three independent experiments. Error bars represent mean values ± S.E.