The FcγRIIB1-mediated inhibition of melanoma development in nude mice requires the receptor’s cytoplasmic tail. HT144IIB1 and HT144IIB(Cyto^–) cell lines were used. (a) Expression of FcγRIIB1 as assessed the day before injection by flow cytometry using AT10 or control IgG1 (dotted histograms). (b) In vitro proliferations of melanoma lines were determined by an MTT assay on days 1–4. (c) Tumor uptake incidence as determined by subcutaneous inoculation of nude mice with 1 × 10⁶ cells. Results are expressed as percentage of mice (n = 20) that remained tumor-free after challenge in two experiments. (d) Tumor growth as measured by volume after inoculation of 2 × 10⁶ cells. Ten mice were used per experiment in each group, and mean tumor volume values of mice bearing tumor with SDs are shown. Data are representative of two independent experiments. *P < 0.01 between HT144IIB1 tumors and HT144 or HT144IIB(Cyto^–) tumors. (e) Immunohistochemical analysis of FcγRII expression using KB61 mAb (right panels), on tumor tissues at day 21 after injection in nude mice of HT144IIB1 (bottom) or HT144IIB(Cyto^–) (top) cells. ×30. (f) Recruitment of SHP2 phosphatase to phosphorylated FcγRIIB1. HT144IIB1 cells (6 × 10⁷) were treated or not treated with 100 μM pervanadate for 10 minutes. Cells were lysed and FcγRIIB was immunoprecipitated using AT10 mAb. Immunoprecipitated materials were electrophoresed and Western blotted with anti-FcγRIIB/IC, anti-phosphotyrosine, anti-SHIP1, anti-SHIP2, anti-SHP1, and anti-SHP2 Ab’s. Whole cell lysates of HT144IIB1 (WCL-H) or ST486 (WCL-S) were used as positive controls.