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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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Dose-dependent detection of purified Stx2a by LFA. Purified Stx2a was serially diluted in PBS from 500 to 0.1 ng/mL and assessed by LFA (top panel); Density measurements were obtained at the test line from three independent experiments and the data plotted as the mean (±SEM) at the test line for each Stx2a dilution (bottom panel). Regression analysis revealed a linear relationship between the density of the test line and the concentration of Stx2a between 20 and 2.5 ng/mL (R2 = 0.99). T, test line; C, control line.
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toxins-07-01163-f002: Dose-dependent detection of purified Stx2a by LFA. Purified Stx2a was serially diluted in PBS from 500 to 0.1 ng/mL and assessed by LFA (top panel); Density measurements were obtained at the test line from three independent experiments and the data plotted as the mean (±SEM) at the test line for each Stx2a dilution (bottom panel). Regression analysis revealed a linear relationship between the density of the test line and the concentration of Stx2a between 20 and 2.5 ng/mL (R2 = 0.99). T, test line; C, control line.

Mentions: Stx LFA sensitivity: Purified Stx2a was used to establish the limit of detection (LOD) for the LFA device. Stx2a was serially diluted in buffer from 500 ng/mL to 0.1 ng/mL, and three independent test strips at each dilution were evaluated. The performance of the LFA was validated by resolution of the control line with a total test time of 5 min. Qualitative detection of Stx2a by visual inspection revealed a dose-dependent increase in test line density with an LOD of 0.1 ng/mL (Figure 2A). To further evaluate the relationship between Stx2a concentration and LFA performance, we performed densitometric analysis of the test line at each dilution in triplicate. The density plot resulted in a sigmoidal-shaped curve that revealed a plateau in test line density at concentrations >100 ng/mL (Figure 2B). Regression analysis for Stx2a test line densities <100 ng/mL showed an optimal linear relationship (R2 = 0.99) of this LFA at Stx2a concentrations between 25 ng/mL and 2.5 ng/mL (Figure 2B; solid line). These results suggest that this LFA could be used to make a quantitative determination of Stx2a concentration from a test line density read from a Stx2a standard curve established between 20 ng/mL and 2.5 ng/mL.


Detection of shiga toxins by lateral flow assay.

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

Dose-dependent detection of purified Stx2a by LFA. Purified Stx2a was serially diluted in PBS from 500 to 0.1 ng/mL and assessed by LFA (top panel); Density measurements were obtained at the test line from three independent experiments and the data plotted as the mean (±SEM) at the test line for each Stx2a dilution (bottom panel). Regression analysis revealed a linear relationship between the density of the test line and the concentration of Stx2a between 20 and 2.5 ng/mL (R2 = 0.99). T, test line; C, control line.
© Copyright Policy
Related In: Results  -  Collection

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

toxins-07-01163-f002: Dose-dependent detection of purified Stx2a by LFA. Purified Stx2a was serially diluted in PBS from 500 to 0.1 ng/mL and assessed by LFA (top panel); Density measurements were obtained at the test line from three independent experiments and the data plotted as the mean (±SEM) at the test line for each Stx2a dilution (bottom panel). Regression analysis revealed a linear relationship between the density of the test line and the concentration of Stx2a between 20 and 2.5 ng/mL (R2 = 0.99). T, test line; C, control line.
Mentions: Stx LFA sensitivity: Purified Stx2a was used to establish the limit of detection (LOD) for the LFA device. Stx2a was serially diluted in buffer from 500 ng/mL to 0.1 ng/mL, and three independent test strips at each dilution were evaluated. The performance of the LFA was validated by resolution of the control line with a total test time of 5 min. Qualitative detection of Stx2a by visual inspection revealed a dose-dependent increase in test line density with an LOD of 0.1 ng/mL (Figure 2A). To further evaluate the relationship between Stx2a concentration and LFA performance, we performed densitometric analysis of the test line at each dilution in triplicate. The density plot resulted in a sigmoidal-shaped curve that revealed a plateau in test line density at concentrations >100 ng/mL (Figure 2B). Regression analysis for Stx2a test line densities <100 ng/mL showed an optimal linear relationship (R2 = 0.99) of this LFA at Stx2a concentrations between 25 ng/mL and 2.5 ng/mL (Figure 2B; solid line). These results suggest that this LFA could be used to make a quantitative determination of Stx2a concentration from a test line density read from a Stx2a standard curve established between 20 ng/mL and 2.5 ng/mL.

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