A toxin-coregulated pilus (TCP), that is important for intestinal colonization of Vibrio cholerae O1, may be produced by vibrios of both classical and EI Tor biotypes. By comparing TCP produced by various strains of the two biotypes in immunoblotting and enzyme-linked immunosorbent assays (ELISA) using monoclonal antibodies (mAbs) and polyclonal antisera against TCP from classical vibrios, we have found biotype-related epitope differences in TCP. Our results indicate that TCP of classical strains has an epitope in the TcpA-subunit (20.5 kDa) that is missing in EI Tor TcpA, and an additional epitope that is more strongly expressed in classical TcpA. A polyclonal antiserum reacted strongly with TcpA from strains of both biotypes in immunoblotting suggesting both the presence of major shared TcpA epitopes and that the low or absent reactivity of EI Tor TcpA with the mAbs was not due to lower production of TcpA by EI Tor strains. Whereas all the TcpA-positive classical strains inhibited the binding of polyclonal antiserum and mAbs to solid phase-bound TCP-positive bacteria in an inhibition ELISA, practically no inhibition was observed with TcpA-positive EI Tor strains. This together with findings in immunoelectron microscopy studies that TCP 'bundles' were only detected on classical strains, suggest that TCP is poorly expressed on EI Tor vibrios.