E. coli Verotoxin 1+2 Ag ELISA
The DRG E. coli Verotoxin 1+2 Ag ELISA is an in vitro device for direct detection of verotoxin 1 and 2 (shiga-toxin 1 and 2) in faecal specimens and stool culture supernatants.
Invasive and toxigenic Escherichia coli strains cause dirrhoea in infants and adults. Among pathogenic E. coli strains the group of enterohaemorrhagic E. coli (EHEC) can cause lifethreatening haemorrhagic colitis and haemolytic uraemic syndrome (HUS) leading to acute renal failure and haemolytic anaemia with thrombocytopenia (1, 2, 3). Strains like E. coli O:157; O:26; O:111 and other serovars are characterized by the production of cytotoxins (verotoxin 1 and 2 or shigatoxin 1 and 2, shiga-toxin variants). The diagnosis of an EHEC infection is initially done by detection of the shiga-toxins. Diagnostic methods can be direct toxin detection by cytotoxicity test on vero-cells and subsequent neutralization test or the detection of the encoding genes with probes or polymerase chain reaction (PCR). These methods are time-consuming and not suited for a routine diagnostic laboratory. Immunological methods like enzyme immunoassay enable a fast and specific shiga-toxin detection in stool specimens. It is commonly recommended to enrich the EHEC bacteria in selective broth media prior to the test run to enhance the sensitivity of the method (4, 5, 6).
The DRG E. coli Verotoxin 1+2 Ag ELISA is an indirect two-site-immunoassay for the qualitative determination of verotoxin 1 and 2 based on polyclonal and monoclonal antibodies.
Verotoxin 1 and/or 2 of specimens and the positive control react with polyclonal anti-verotoxin 1 and 2 antibodies coated on the solid phase of the microplate. After incubation for 60 minutes at 22-25°C non-bound material is removed by a washing step.
Subsequently bound toxins specifically react with biotinylated monoclonal anti-verotoxin 1 and anti-verotoxin 2 antibodies during a second incubation period of 30 min at 22-25°C. Non-bound material is separated from the solid-phase immune complexes by a subsequent washing step.
During the next incubation period of 30 min at 22-25°C horseradish peroxidase (HRP) conjugated streptavidin reacts with the bound biotinylated antibodies. Unbound conjugate is removed by a washing step.
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