Characterization of mAbs against a putative ILT7 ligand. (A) Culture supernatants from different hybridoma clones were included in the co-cultures of ILT7 reporter cells and T47D cells. The percentages of GFP-positive reporter cells were determined. Two clones able to inhibit GFP induction are indicated (arrows). Anti-ILT7 mAb (clone 17.2) was included as a positive control. (B) Five human breast cancer cells were stained with mAb 26F8 (red line) or an isotype-matched control mAb (shaded area) and analyzed by flow cytometry. A similar result obtained with mAb 28G4 is not shown. Data are representative of three independent experiments. (C) T47D cells were co-cultured with ILT7 reporter cells in the presence of control IgG1, 26F8, or 28G4 mAbs at different concentrations. The percentages of GFP-positive reporter cells were plotted. Data are representative of four independent experiments. (D) Surface biotinylated MDA-MB-231 and T47D were immunoprecipitated with control IgG1, 26F8, or 28G4. The precipitated proteins were analyzed by Western blotting with NeutrAvidin-HRP. Arrows indicate the specific protein bands obtained from T47D cells. Data are representative of two independent experiments.