Detection and characterization of the coli surface antigen 6 of enterotoxigenic Escherichia coli strains by using monoclonal antibodies.

We describe, for the first time, the production of monoclonal antibodies (MAbs) against coli surface antigen 6 (CS6) of enterotoxigenic Escherichia coli (ETEC) and their use for characterization and diagnosis of CS6. Two MAbs, MAbs CS6-20:11:9 and CS6-2A:14, were produced by immunizing mice with purified CS6 or CS6-containing bacterial extracts. The MAb specificity was demonstrated by enzyme-linked immunosorbent assay (ELISA), immunoblotting, and immunoelectron microscopy, which showed that the MAbs bound to CS6-expressing bacteria as well as to purified CS6 and CS6 structural subunits but not to CS6-negative bacteria or other purified ETEC colonization factors. By using bacterial recombinants, i.e., strains with a complete CS6 operon or parts thereof, it was found that both MAbs were specific for CssB, one of the two structural subunits of CS6. Although the MAbs bound specifically to the entire surface of CS6-expressing bacteria, no structure of CS6 could be identified. The usefulness of the MAbs for the detection of CS6 was evaluated in an inhibition ELISA and in a dot blot test. Ninety-two ETEC strains with known colonization factors were analyzed, and all CS6-positive strains were identified by either assay with MAb CS6-2A:14, whereas MAb CS6-20:11:9 failed to identify two CS6-positive strains; in no instance was any CS6-negative strain identified by either of the MAbs. Parallel analyses of 48 strains with a gene probe specific for the other structural subunit of CS6, i.e., CssA, and the MAb-based assays gave identical results, suggesting the simultaneous presence of both subunits.