FIG. 3.
Effect of Salmonella infection on the stability of MHC-II complexes. BM-DC from BALB/c mice were infected with GFP-expressing Salmonella WT or sseC-deficient (SPI2) strains as indicated at an MOI of 25. As controls, BM-DC were uninfected or were stimulated with LPS. If indicated, BM-DC were also stimulated with 50 μg of OVA/ml. To enrich Salmonella-infected BM-DC, 24 h after infection the cells were subjected to FACS for CD11c-positive and GFP-positive cells. BM-DC from the uninfected control assays were purified by MACS for CD11c-positive cells. About 6 × 105 cells were collected for each assay and subjected to the SDS stability assay as described in Materials and Methods. (A) Western blot analysis of SDS-stable MHC-II complexes (MHC-II) were performed with monoclonal antibody to mouse I-Ad/I-Ed. As loading controls, Western blots were stripped and reprobed with an antibody to p38. (B) For quantification, intensities for the MHC-II bands on Western blots were determined by densitometry and expressed as the percent transmission. Means and standard deviations of two blots are shown. The data shown are representative for three independent experiments with similar outcomes.