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Detection of shiga toxins by lateral flow assay.

Ching KH, He X, Stanker LH, Lin AV, McGarvey JA, Hnasko R - Toxins (Basel) (2015)

Bottom Line: The contamination of food products with STEC represents a food safety problem that necessitates rapid and effective detection strategies to mitigate risk.This LFA provides a rapid and sensitive test for the detection of Stxs directly from STEC culture supernatants or at risk food samples with a 0.1 ng/mL limit of detection (LOD) for Stx2a.This Stx LFA is applicable for use in the rapid evaluation of Stx production from cultured E. coli strains or as a tool to augment current methods as part of food safety testing.

View Article: PubMed Central - PubMed

Affiliation: Produce Safety & Microbiology Research Unit, Agricultural Research Service, U.S. Department of Agriculture, 800 Buchanan St, Albany, CA 94710, USA. kathryn.ching@ars.usda.gov.

ABSTRACT
Shiga toxin-producing Escherichia coli (STEC) produce shiga toxins (Stxs) that can cause human disease and death. The contamination of food products with STEC represents a food safety problem that necessitates rapid and effective detection strategies to mitigate risk. In this manuscript, we report the development of a colorimetric lateral flow assay (LFA) for the rapid detection of Stxs in <10 min using a pair of monoclonal antibodies that bind epitopes common to Stx1 and six Stx2 variants. This LFA provides a rapid and sensitive test for the detection of Stxs directly from STEC culture supernatants or at risk food samples with a 0.1 ng/mL limit of detection (LOD) for Stx2a. This Stx LFA is applicable for use in the rapid evaluation of Stx production from cultured E. coli strains or as a tool to augment current methods as part of food safety testing.

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Related in: MedlinePlus

Detection of shiga toxins from STEC serotypes by lateral flow assay (LFA). Supernatant from Stx1 and seven variant Stx2-producing STEC cultures were diluted 1:5 in PBS and evaluated using the Stx LFA. −/−, a non-Stx-producing E. coli serotype; LB, Luria broth; T, test line; C, control line.
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toxins-07-01163-f001: Detection of shiga toxins from STEC serotypes by lateral flow assay (LFA). Supernatant from Stx1 and seven variant Stx2-producing STEC cultures were diluted 1:5 in PBS and evaluated using the Stx LFA. −/−, a non-Stx-producing E. coli serotype; LB, Luria broth; T, test line; C, control line.

Mentions: Stx LFA specificity: To determine the binding specificity of our Stx LFA, we evaluated nine independent STEC strains (Table 1). Genetic analysis confirmed that each strain carried only one Stx variant gene, and toxin expression was evaluated by the Vero cell assay [22]. Culture supernatants from each STEC strain along with non-STEC E. coli controls were diluted and positive LFA test lines resolved for those producing Stx1 and Stx2 variants (Figure 1). The supernatant from Stx1-, Stx2a- and Stx2g-producing cultures gave a strong positive result on the LFA with a dense test line resolving <1 min after initiation of the assay. Culture supernatants from Stx2c-, 2d-, 2e- and 2f-producing E. coli strains were slower to resolve by LFA and resulted in test lines of weaker intensity. The detection of the Stx2b variant in the supernatant of the O118:H2 clinical serotype was inconclusive. The Stx2b test line resolved at an intensity that was equivalent to, or slightly above, the background established by non-Stx producing E. coli culture supernatant (−/−). Luria-Bertani broth (LB) served as a negative test line control. All LFA test strips displayed a visible control line, validating the proper function of the assay. The presence of these toxins in the culture supernatant was confirmed by a sandwich ELISA using a polyclonal anti-Stx antibody (unpublished data).


Detection of shiga toxins by lateral flow assay.

Ching KH, He X, Stanker LH, Lin AV, McGarvey JA, Hnasko R - Toxins (Basel) (2015)

Detection of shiga toxins from STEC serotypes by lateral flow assay (LFA). Supernatant from Stx1 and seven variant Stx2-producing STEC cultures were diluted 1:5 in PBS and evaluated using the Stx LFA. −/−, a non-Stx-producing E. coli serotype; LB, Luria broth; T, test line; C, control line.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4417961&req=5

toxins-07-01163-f001: Detection of shiga toxins from STEC serotypes by lateral flow assay (LFA). Supernatant from Stx1 and seven variant Stx2-producing STEC cultures were diluted 1:5 in PBS and evaluated using the Stx LFA. −/−, a non-Stx-producing E. coli serotype; LB, Luria broth; T, test line; C, control line.
Mentions: Stx LFA specificity: To determine the binding specificity of our Stx LFA, we evaluated nine independent STEC strains (Table 1). Genetic analysis confirmed that each strain carried only one Stx variant gene, and toxin expression was evaluated by the Vero cell assay [22]. Culture supernatants from each STEC strain along with non-STEC E. coli controls were diluted and positive LFA test lines resolved for those producing Stx1 and Stx2 variants (Figure 1). The supernatant from Stx1-, Stx2a- and Stx2g-producing cultures gave a strong positive result on the LFA with a dense test line resolving <1 min after initiation of the assay. Culture supernatants from Stx2c-, 2d-, 2e- and 2f-producing E. coli strains were slower to resolve by LFA and resulted in test lines of weaker intensity. The detection of the Stx2b variant in the supernatant of the O118:H2 clinical serotype was inconclusive. The Stx2b test line resolved at an intensity that was equivalent to, or slightly above, the background established by non-Stx producing E. coli culture supernatant (−/−). Luria-Bertani broth (LB) served as a negative test line control. All LFA test strips displayed a visible control line, validating the proper function of the assay. The presence of these toxins in the culture supernatant was confirmed by a sandwich ELISA using a polyclonal anti-Stx antibody (unpublished data).

Bottom Line: The contamination of food products with STEC represents a food safety problem that necessitates rapid and effective detection strategies to mitigate risk.This LFA provides a rapid and sensitive test for the detection of Stxs directly from STEC culture supernatants or at risk food samples with a 0.1 ng/mL limit of detection (LOD) for Stx2a.This Stx LFA is applicable for use in the rapid evaluation of Stx production from cultured E. coli strains or as a tool to augment current methods as part of food safety testing.

View Article: PubMed Central - PubMed

Affiliation: Produce Safety & Microbiology Research Unit, Agricultural Research Service, U.S. Department of Agriculture, 800 Buchanan St, Albany, CA 94710, USA. kathryn.ching@ars.usda.gov.

ABSTRACT
Shiga toxin-producing Escherichia coli (STEC) produce shiga toxins (Stxs) that can cause human disease and death. The contamination of food products with STEC represents a food safety problem that necessitates rapid and effective detection strategies to mitigate risk. In this manuscript, we report the development of a colorimetric lateral flow assay (LFA) for the rapid detection of Stxs in <10 min using a pair of monoclonal antibodies that bind epitopes common to Stx1 and six Stx2 variants. This LFA provides a rapid and sensitive test for the detection of Stxs directly from STEC culture supernatants or at risk food samples with a 0.1 ng/mL limit of detection (LOD) for Stx2a. This Stx LFA is applicable for use in the rapid evaluation of Stx production from cultured E. coli strains or as a tool to augment current methods as part of food safety testing.

Show MeSH
Related in: MedlinePlus