Immobilized DC-HIL-Fc inhibits T-cell activation triggered by anti-CD3 Ab. CD4⁺ (A-B) or CD8⁺ (C-D) T cells (2 × 10⁵ each) purified from BALB/c spleens were cultured for 48 hours in microculture wells precoated with increasing doses of anti-CD3 Ab and with a constant dose (10 μg/mL) of DC-HIL-Fc (•), control hIgG (○), or neither (none; ▵). After pulsing with ³H-thymidine, cells and culture supernatant were harvested. ³H-thymidine incorporation into cells (A,C) and IL-2 production (B,D) were determined, and values (cpm or ng/mL) plotted at a logarithmic scale, respectively. (E) Titration of inhibitory function of DC-HIL-Fc. CD4⁺ T cells were cultured for 48 hours in microculture wells precoated with a constant dose (0.3 μg/mL) of anti-CD3 Ab and increasing doses of DC-HIL-Fc or control hIgG. (F) CD28 costimulation rescues DC-HIL-Fc–induced inhibition of CD4⁺ T-cell activation. Purified CD4⁺ T cells were cultured in wells precoated with anti-CD3 Ab (0.3 μg/mL), hIgG or DC-HIL-Fc (5 μg/mL), and anti-CD28 mAb (increasing doses). (G-H) Previously activated T cells were prepared from Tac-TgN(DO11.10)-Rag2^tm1mice and reactivated by immobilized anti-CD3 Ab (varying doses) and DC-HIL-Fc or hIgG (constant dose). (I-J) Inhibitory function of DC-HIL-Fc was titrated against reactivation of previously activated T cells by anti-CD3 Ab (1 μg/mL). Proliferation (G,I) and IL-2 production (H,J) were measured. Results are expressed as mean values ± SDs. Data shown are representative of 6 (A-B), 3 (G-J), and 2 (C-F) experiments, respectively.