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ARS Home » Northeast Area » Wyndmoor, Pennsylvania » Eastern Regional Research Center » Molecular Characterization of Foodborne Pathogens Research » Research » Publications at this Location » Publication #313525

Title: Microtiter plate-based antibody microarrays for bacteria and toxins

Author
item Gehring, Andrew

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 2/1/2015
Publication Date: 4/21/2015
Citation: Gehring, A.G. 2015. Microtiter plate-based antibody microarrays for bacteria and toxins. Meeting Abstract. Meeting Abstract.

Interpretive Summary:

Technical Abstract: Research has focused on the development of rapid biosensor-based, high-throughput, and multiplexed detection of pathogenic bacteria in foods. Specifically, antibody microarrays in 96-well microtiter plates have been generated for the purpose of selective detection of Shiga toxin-producing E. coli (STEC) bacteria and associated toxins (Stx1 and Stx2). Several fluorescence and colorimetric immunoassay platforms were created for capture and labelling of various STEC and their toxins at the well bottoms of inexpensive, polystyrene microtiter plates (substrates) array-printed and with passively adsorbed antibodies. Centrifugation was frequently employed to significantly improve localization of bacteria to planar capture surfaces containing the printed antibodies. Fluorescence or enzymatic label was conferred among captured bacteria either using either nucleic acid intercalating dyes as universal labels or a “cocktail” of dye- or enzyme-conjugated antibodies. Quantitative detection of the antibody captured bacteria was affected through laser-induced fluorescence scanning or visible light flatbed scanning of colored enzymatic products. Optimized assays had demonstrated limits of detection of ~10**5 cells/mL and ~4.5 ng/mL for E. coli O157:H7 and Stx1 (or Stx2), respectively, in a total assay time of 2 h or less. In related research, a mixed culture enrichment broth was developed in support of this assay for the detection of additional bacterial pathogens, particularly Listeria monocytogenes. This new procedure may find use by food producers and testing laboratories for the high-throughput screening of food samples for pathogens or the efficacy of biorecognition elements.