FIG. 1.

Immunoblot analysis of secreted ExoT. Bacterial strains were cultured under the same conditions used for phenotypic assays (LB broth with 200 μg of carbenicillin per ml for plasmid-containing strains, standing for 18 h at 37°C). Proteins secreted into the culture medium were precipitated with ammonium sulfate, electrophoresed on SDS-polyacrylamide gels, transferred to nitrocellulose, and hybridized with rabbit anti-ExoT polyclonal antiserum. Goat anti-rabbit IgG horseradish peroxidase conjugate was used as a secondary antibody, and detection was performed using the ECL system. Lane 1, PA103; lane 2, PA103ΔU; lane 3, PA103ΔUΔT; lane 4, PA103ΔU/T(R149G); lane 5, PA103ΔU/T(R149K); lane 6, PA103pscJ::Tn5; lane 7, PA103ΔUΔT+pUCP20; lane 8, PA103ΔUΔT+pExoT; lane 9, PA103ΔUΔT+pExoT(R149G); lane 10, PA103ΔUΔT+pExoT(R149K).