Figure 7

Cleavage-deficient point-mutant rIde_Ssuis_homologue_C195S binds more strongly to IgM⁺ B cells than does rIde_Ssuis_homologue. A Flow cytometry analysis of FITC⁺ cells. Porcine PBMC (n = 6) were incubated with FITC-labelled proteins: (i) rIde_Ssuis_homologue (rIde_Ssuis_h), (ii) rIde_Ssuis_homologue_C195S (rIde_Ssuis_h_C195S), (iii) rIde_Ssuis_C_domain or (iv) rMRP (all 4 µg/10⁶ cells, 45 min). Upper row: Cells were analyzed directly after fixation, with a total of 4 washing steps before analysis. Lower row: Cells were stained for IgM⁺ B cells for a total of 14 washing steps before analysis. The percentage of FITC⁺ cells (left hand side) and the respective median fluorescence intensity (MFI, right hand side) are shown. Statistical analysis was conducted with ordinary one-way ANOVA with Tukey’s multiple comparisons test (p < 0.0001 ****, p < 0.001 ***, p < 0.01 **, p < 0.05 *, p > 0.05 ns). Only comparisons with p < 0.05 are shown. The bars and error bars represent the means and standard deviations, respectively. B FITC-labelled recombinant proteins do not bind to CD3⁺ T cells. Porcine PBMC (n = 6) were incubated in cell culture medium alone or with the indicated recombinant proteins as described above. The CD3 + T cells were stained and analysed by flow cytometry (for a total of 14 washing steps before analysis).