Characterization of rabbit polyclonal sera against recombinant Shiga toxin and its subunits for detection of Stx-producing E. coli.

Abstract

Shiga toxin (Stx) is the principal virulence factor of Shigatoxigenic Escherichia coli (STEC), a food-born pathogen associated disease with uncomplicated diarrhea or the hemolytic-uremic syndrome. In this study, rabbit polyclonal anti recombinant A, B subunits of Shiga toxin and holotoxin antisera were raised and employed for immunological purpose. By immunoblotting, these antisera recognized respective subunit and the holotoxin antiserum recognized both subunits, equally. The raised antisera could also neutralize the cytotoxicity of the shiga toxin on vero cells. The neutralizing ability of the prepared sera was compared for different subunits. The neutralization of toxicity was observed by incubation of raised sera with the rStx or Shiga toxin from wild type strains. The inhibition of cell toxicity was shown by anti-A, anit-B and anti-AB antisera, separately. It was shown that anti-A antibody, more significantly recognized Stx producing strains, comparing to anti-B antibody. These sera from immunized rabbits were also used as specific antibodies in Enzyme-Linked Immunosorbant Assay (ELISA) for detection of Shiga toxin. It was demonstrated that the raised antibodies especially antibody against A subunit could be a useful tool for immunological diagnosis of STEC induced infection.