Binding activity of β₂ integrins on DCs. (A) bmDCs (day 7) were pretreated with 5 mM Mg²⁺ either alone or together with the anti-CD18 antibodies 18.2 or GAME46 (10 μg/mL). Cells (5 × 10⁵) were placed in 96-well plates in triplicate for 45 minutes at 37°C. Wells were either coated with 10 μg/mL recombinant ICAM-1–Fc or human IgG as a control. After washing, plates were analyzed for bound cells by microscopy (original magnification × 10) using the NIH Image 1.55 software. One representative experiment of 4 is shown. (B) bmDCs from CD18^+/+ and CD18^−/− mice were prepared and used to assay ICAM-1 binding as in panel A. One of 3 experiments is shown. (C) T cells were cocultured with either untreated or Mg²⁺ pretreated CD18^+/+ or CD18^−/− bmDCS (day 7) in collagen gels. Contact times of individual T-DC pairs were analyzed as in Figure 1B. One representative experiment of 2 is shown. Horizontal bars represent mean values.