Production and characterisation of monoclonal antibodies to Verotoxins 1 and 2 from Escherichia coli of serotype O 157:H7

Abstract

Fourteen hybridoma cell lines were isolated that produced monoclonal antibodies (MAbs) to purified Verotoxins 1 and 2 (VT1, VT2) of Escherichia coli of serotype O157:H7. Of these MAbs, eight were obtained by immunisation of BALB/c mice with purified VT1, and six were obtained from BALB/c mice immunised with purified VT2. With the exception of MAb 1C5, with a heavy chain of IgG2b class, antibodies produced from mice immunised with heat-treated toxin were of IgM class. MAbs produced from mice immunised with heat-treated VT1 or VT2 reacted with both verotoxins in ELISA, and Western-blot analysis revealed that they reacted with subunit A and the A1 fragment of nicked subunit A of both toxins, but not with subunit B; furthermore, none of them neutralised Vero cytotoxicity or mouse lethality of either toxin. In contrast, MAbs produced from mice immunised with heat-treated and formalin-treated VT1 reacted in Western blots with subunits A and B of VT1 and subunit A, but not subunit B, of VT2, reacted in ELISA with VT1 only, and neutralised Vero cytotoxicity and mouse lethality of VT1 but not of VT2. Results indicate the existence of a common epitope on subunit A of VT1 and VT2 that is not responsible for the biological activity of these toxins, and that subunit B is essential for the biological activity of VT1. MAbs capable of reacting with both verotoxins from E. coli of serotype O157:H7 may be useful reagents for screening bacterial isolates capable of producing one or both of these toxins.