Suppression of the initiation and persistence of IgE-CAI by Ba103 administration in vivo. (A) BALB/c mice were passively sensitized with an intravenous injection of TNP-specific IgE 1 day before being challenged with an intradermal injection of TNP-OVA into the left ear and control OVA into the right ear. The mice were treated with a single intravenous injection of 30 μg Ba103 or control IgG at the indicated time point before or after the antigen challenge. In addition, daily from days −1 to 5 except on the day of the Ba103 injection, the mice were given 30 μg control IgG in place of Ba103. Thus, the control mice (■) received 7 injections of control IgG from days −1 to 5 while the Ba103-treated mice (○) received 1 injection of Ba103 on the indicated day and 6 injections of control IgG on the others. The thicknesses of the left and right ears were measured at the indicated time points. The values of ΔEar thickness, the differences in ear thickness (left-right) at each time point, are plotted. Data are expressed as the mean (± SEM) of 3 mice in each group and are representative of 3 repeated experiments. In these experiments, as well as those shown in panel D, we focused on the incidence of IgE-CAI (the delayed-onset ear swelling); therefore, the early changes in ear thickness, including those of the immediate-phase and late-phase ear swelling, are not plotted. (B) In parallel experiments performed as described in panel A, ear specimens were prepared on day 3 after challenge from mice treated with Ba103 or control IgG on day −1, and stained with Giemsa. Images were acquired using an Olympus IX71 microscope equipped with a 10×/0.30 numerical aperture objective lens (Olympus, Tokyo, Japan). Images were taken using an Olympus DP70 digital camera with the software of DP controller version 2.1.1.183. The images were subsequently processed with Adobe Photoshop 8.0.1 software (Adobe Systems, San Jose, CA). (C) In parallel experiments performed as described in panel A, on day 3 after the TNP-OVA challenge, cells were isolated by treatment with collagenase from the ears of 3 mice injected with Ba103 (■) or control IgG (▩) on day 2 after challenge. After incubation with IgE at 4°C for 30 minutes to saturate FcϵRI, cells from each group of mice were stained for IgE, Gr-1, c-kit, and Siglec-F. Basophils, eosinophils, and neutrophils were defined as IgE⁺c-kit⁻ cells, Siglec-F⁺Gr-1^low cells, and Siglec-F⁻Gr-1^high cells, respectively. The number of each cell type per ear is shown, and data are representative of 3 repeated experiments. (D) WBB6F1-W/W^v mice were passively sensitized with IgE and then challenged with antigens as described in panel A. The mice were treated with an intravenous injection of 30 μg Ba103 on day 1 and 30 μg control IgG on day 3 (○), control IgG on day 1 and Ba103 on day 3 (◇), or control IgG on both days (■). Data are expressed as the mean (± SEM) of 5 mice in each group and are representative of 3 repeated experiments.