COMPARISON OF THE WESTERN-BLOT ASSAY WITH THE NEUTRALIZING-ANTIBODY AND ENZYME-LINKED IMMUNOSORBENT ASSAYS FOR MEASURING ANTIBODY TO VEROCYTOTOXIN-1

A Western blot (immunoblot) assay (WBA) was developed to detect immuno globulin G (IgG) antibodies against Escherichia coli Verocytotoxin 1 ( VT1) by using a chemiluminescence detection system. The assay was comp ared with a VT1-neutralizing-antibody (VT1-NAb) assay and an anti-VT1 IgG enzyme-linked immunosorbent assay (ELISA). When four human serum s amples that were known to be positive by VT1-NAb assay and ELISA were titrated to the endpoint by the three assays, the WBA gave endpoint ti ters that were up to 8-fold higher than those by ELISA and up to 256-f old higher than those by the VT1-NAb assay. Of 32 serum samples that w ere known to be positive by VT1-NAb assay and ELISA, 31 (97%) were pos itive by WBA; the one sample with a discrepant result gave borderline results by the VT1-NAb assay and ELISA. Of 52 serum samples that were known to be negative by the VT1-NAb assay and ELISA, 50 (96%) were neg ative and 2 (4%) were positive by WBA. Of 44 serum samples that gave d iscrepant results by the VT1-NAb assay and ELISA, neither of the latte r correlated with the results of WBA. In an investigation of 19 pairs of acute- and convalescent-phase serum samples from patients with hemo lytic-uremic syndrome, 10 pairs that were positive by the VT1-NAb assa y were also WBA positive, while 9 pairs that were NAb negative were al so WBA negative. The WBA is inherently more specific and sensitive tha n either the NAb assay or the ELISA and may be used as a ''gold standa rd'' to detect IgG antibodies to VT1. Like the NAb assay and the ELISA for detecting antibodies to VT1, the WBA has little to offer in the d iagnostic setting but is expected to play an important role in seroepi demiological studies.