Escherichia coli, O157, BioAssay(TM) ELISA Kit

Strains of Escherichia coli that produce Shiga-like toxins (SLTs), also known as Verocytotoxins (VTs), are an important cause of human disease. Clinical manifestations of infection include diarrhea, hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). Although Shiga-like toxin producing E. coli (SLTEC) belong to many O serogroups, serotypes O157/H7 and O157/H are the predominant SLTEC associated with HC and HUS.\n\nOutbreaks of HC and HUS due to E. coli O157 have been linked to consumption of undercooked ground beef (Riley et al, 1983; Wells et al 1983; Padhye and Doyle, 1992). Prevalence studies indicate that up to 3.7% of retail meats may contain E. coli O157 (Doyle and Schoeni 1987). Although this may seem low compared to some other foodborne pathogens, the seriousness of disease caused by E. coli O157 has made this pathogen a major food safety concern.\n\nDetection of E. coli O157/H7 in meats by cultural methods is time consuming, requiring several days to obtain presumptive positive results. Typically, meat samples are cultured in primary enrichment broth which is plated onto sorbitol-MacConkey (SMAC) agar. Non-sorbitol fermenting colonies are then subcultured for identification by serotyping or related methods. As some E. coli O157 strains associated with HC and HUS ferment sorbitol (Karch et al 1993), this approach may give false negative results.\n\nThe United States Biological E. coli O157 ELISA is a rapid and reliable test which significantly reduces the time required to screen foods for the presence of E. coli O157. Primary enrichment cultures grown for 8-16 hours can be tested in less than one hour, allowing ELISA-negative product to be released within 24 hours. Enrichment broths presumptively positive for E. coli O157 on the basis of positive ELISA tests can be cultured further for confirmation by standard methods.\n\nIntended Use:\nThe United States Biological E. coli O157 assay is an enzyme-linked immunosorbent assay (ELISA) that may be used to screen food products for the presence of E. coli O157 antigen.\n\nTest Principle:\nThe United States Biological E. coli O157 ELISA is a double antibody (sandwich) ELISA utilizing specific anti-E. coli O157 antibodies coated to microwells. After addition of the sample and the enzyme conjugate, a positive reaction (indicating the presence of E. coli O157 antigen) produces a deep blue color. Addition of the Stop Solution ends the assay and turns the blue color to yellow. The results may be read visually or with an ELISA reader.\n\nKit Components:\n1. Microtiter plate 1x96 wells each.\n2. Test strip holder: 1.\n3. Enzyme Conjugate: 1x11ml of peroxidase polyclonal antibody.\n4. Positive control: 1x1ml of killed cells in a buffered base.\n5. Negative control: 1x1ml of buffered base.\n6. Chromogen: 1x11ml of the chromogen tetramethylbenzidine (TMB).\n7. Wash Concentrate 20x: 2x25 ml of concentrated buffer and surfactant with preservative.\n8. Stop solution: 1x11ml of 1M phosphoric acid.\n\nStorage and Stability:\nStore components at 4 degrees C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vial prior to removing the cap.

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SPECIFICATIONS

Catalog Number

E3500-03

Size

1Kit

Applications

ELISA

References

1. Doyle, M.P., and J.L. Schoeni. 1987. Isolation of Escherichia coli O157:H7 from retail fresh meats and poultry. Appl. Environ. Microbiol. 53:2394-2396. 2. Karch, H., H. Bohm, H. Schmidt, F. Gunzer, S. Aleksic, and J. Heesemann. 1993. Clonal structure and pathogenicity of Shiga-like toxin-producing, Sorbitol-fermenting Escherichia coli O157:H7. J. Clin. Microbiol. 24:1200-1205. 3. Karmali, M.A. Infection by verocytotoxin-producing Escherichia coli. J Clin. Microbiol. Rev. 2:15-38 1989. 4. Padhye, N.V., and M.P. Doyle. 1991. Production and characterization of a monoclonal antibody specific for enterohemorrhagic Escherichia coli of serotypes O157:H7 and 026:H11. J. Clin. Microbiol. 24:99-103. 5. Riley, L.W., R.S. Remis, S.D. Helgerson, H.B. McGee, J.G. Wells, B.R. Davis, R.J. Herbert, E.S. Olcott, L.M. Johnson, N.T. 6. Hargrett, P., A. Blake, and M.L. Cohen. 1983. Hermorrhagic colitis associated with a rare Escherichia coli serotype. N. Engl. J. Med. 308:681-685. 6. Wells, J.G., B.R. Davis, I.K. Wachsmuth, L.W. Riley, R.S. Remis, R. Sokolow, and G.K. Morris. 1983. Laboratory investigation of hemorrhagic colitis outbreaks associated with a rare Escherichia serotype. J. Clin. Microbiol. 18:512-520. 7. Okrend, A.J.G. and Rose, B.E. Isolation and Identification of E. coli O157:H7 from Meat. Lab Commication, Rev#3, USDA, FSIS 1989.