Microbead-Based Immunoassay for Simultaneous Detection of Shiga Toxins and Isolation of Escherichia
coli O157 in Foods
Shiga toxin–producing Escherichia coli (STEC) is a significant foodborne pathogen with great economic consequences. There has been an increased food safety concern with this organism since outbreaks of human illnesses caused by this pathogen were first reported in 1982.
Therefore, developing a reliable, sensitive, and rapid assay capable of detecting E. coli O157 and the main toxins produced by STEC (i.e., Shiga toxins 1 [Stx1] and 2 [Stx2]) will directly benefit regulatory agencies by minimizing analysis time. Here, we use Luminex
technology to detect multiple analytes in a single 50-ml sample. Using commercially available monoclonal antibodies coupled to carboxylated magnetic microbeads, we developed an immunoassay capable of simultaneously serotyping E. coli O157 and detecting Stx1 and/or Stx2.
The specificity and sensitivity of this immunoassay was tested against a collection of 34 E. coli isolates belonging to various O serogroups phenotypically different for Stx. The results were compared with microplate sandwich enzyme-linked immunosorbent assay (ELISA), and no cross-reactivity
was observed for any of the monoclonal antibodies used. An increased sensitivity up to 1,000 times was observed in the microbead-based immunoassay when compared with the microplate sandwich ELISA. The results indicate that Luminex technology has the potential to simultaneously detect multiple
targets without loss of specificity and/or sensitivity. A blind experiment was conducted with 48 samples of ground beef, lettuce, and milk spiked with ≤2 CFU/g E. coli. All the samples were correctly identified, with no false positives or false negatives. This microbead-based immunoassay
could be extended to simultaneously detect additional foodborne pathogens and their toxic markers.
Document Type: Research Article
Western Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, Albany, California 94710
U.S. Food and Drug Administration, Alameda, California 94502, USA
Western Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, Albany, California 94710. firstname.lastname@example.org
Publication date: March 1, 2011
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