FIG. 4.

Immunoblot analysis of Stx2 expression in E. coli culture supernatants. Samples of cell-free E. coli culture supernatants (35 μl) and various amounts of purified Stx2 were separated on a sodium dodecyl sulfate-10 to 20% polyacrylamide gradient gel, transferred onto nitrocellulose, and probed with the mouse monoclonal anti-Stx2 antibody VT 135/6-B9 (Sifin GmbH, Berlin, Germany), which recognizes the 32-kDa A subunit and the 27-kDa A1 fragment of Stx2 (10). Molecular masses of the protein standards are shown to the left. Lanes: 1, Stx1- and Stx2-producing E. coli O157 strain RM1697; 2, non-Stx-producing E. coli O157 strain RM4876; 3, nonpathogenic E. coli strain RM5034; 4, 0.25 μg of Stx2; 5, 0.125 μg of Stx2; 6, 0.06 μg of Stx2; and 7, 0.03 μg of Stx2.