Evaluation of ELISA tests specific for Shiga toxin 1 and 2 in food and water samples

Abstract

Two enzyme-linked immunosorbent assay (ELISA) kits were evaluated for their effectiveness in detecting and differentiating between Shiga toxin 1 and 2 (Stx1 and Stx2) produced by Shiga toxin-producing E.coli (STEC) inoculated into food and water samples. Each kit incorporated monoclonal antibodies previously determined to bind all known Stx1 or Stx2 subtypes with the exception of Stx2b. Four different sample types, including ground beef, Romaine lettuce, pond water, and pasteurized milk were inoculated with Stx1a-, Stx2a-, or Stx1a-and Stx2a-producing STEC strains, enriched using modified tryptic soy broth (containing mitomycin C) for 6, 16, and 22 h, and tested using the ELISA kits in the presence of a bacterial protein extraction reagent (B-PER™). The two Shiga toxin types were readily detected and distinguished for all tested sample types. There was good overall sensitivity, specificity, variance, and reproducibility for the two ELISA kits and they should prove useful for application in food testing.

Three Dimensional Vero Cell-Platform for Rapid and Sensitive Screening of Shiga-Toxin Producing Escherichia coli

“…This enrichment step for sample preparation is required for most, if not all detection assays including immunoassay and PCR (Gould et al, 2009). It is well documented that PCR and immunoassays have superior sensitivity in detecting STEC (10 CFU/ml) and Stx (0.5 ng/ml) (Il-Hoon et al, 2015; Gehring et al, 2017); however, there are also drawbacks in detection time, sample and reagent preparation, and information provided about functional activity of Stx and viability of STEC (Pimbley and Patel, 1998; Cui et al, 2003; Banerjee et al, 2013; Amani et al, 2015; Zeng et al, 2016). Vero cell assay can also detect picogram quantities of Stx, however, detection time must be extended to 24–72 h to acquire confirmatory results (Hughes et al, 1998; Rasooly and Do, 2010; Lentz et al, 2011).…”

Detection of Shiga Toxin 2 Produced by Escherichia coli in Foods Using a Novel AlphaLISA

“…Lettuce and ground beef samples were prepared following the procedures described previously [ 15 ]. Briefly, Romaine lettuce was purchased from a local grocery store, the outermost leaves were removed, while the remaining leaves were chopped into ~1 cm 2 pieces on a 70% ethanol sterilized surface.…”

“…Because culture-based STEC detection can be long and laborious, alternative methods for identifying strains that produce Shiga toxin have emerged. More rapid antibody-based detection tests, specifically enzyme-linked immunosorbent assays (ELISAs) [ 15 , 16 ], antibody-based lateral flow assays (LATs) [ 17 ], and immunomagnetic separation assays (IMS) [ 18 , 19 ] have been employed as common testing platforms for the detection of STECs and Shiga toxin in lieu of traditional microbiological detection methods such as colony plating on selective media. A new platform known as the AlphaLISA, or amplified luminescent proximity homogenous assay-linked immunosorbent assay, utilizes both bead- and antibody-based technologies for pathogen or toxin detection.…”