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United States Department of Agriculture

Agricultural Research Service

Research Project: Technologies for Detecting and Determining the Bioavailability of Bacterial Toxins

Location: Foodborne Toxin Detection and Prevention

Title: A monoclonal antibody based capture ELISA for botulinum neurotoxin serotype B: toxin detection in food

Authors
item Stanker, Larry
item Scotcher, Miles -
item Cheng, Luisa Wai Wai
item Ching, Kathryn -
item McGarvey, Jeffery
item Hodge, David -
item Hnasko, Robert

Submitted to: Toxins
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 7, 2013
Publication Date: November 15, 2013
Citation: Stanker, L.H., Scotcher, M.C., Cheng, L.W., Ching, K., McGarvey, J.A., Hodge, D., Hnasko, R.M. 2013. A monoclonal antibody based capture ELISA for botulinum neurotoxin serotype B: toxin detection in food. Toxins. 5(11):2212-2226. DOI:10.3390/toxins5112212.

Interpretive Summary: Botulism is a serious, often fatal neuroparalytic disease in humans and animals caused by a protein toxin (botulinum toxin, BoNT) produced by the bacterium Clostridium botulinum. Of the seven different types of BoNT, four cause human disease, serotypes A, B, E, and F. The standard test for BoNT is the mouse bioassay (MBA). While being very sensitive, the MBA requires between 2-7 days to complete, uses death as an end point, is expensive and only available in a few laboratories. A rapid, monoclonal antibody-based immunoassay has been developed. This assay has a sensitivity comparable to that observed with the MBA but can be completed in a few hours versus a few days. Using this test botulinum toxin added to ground beef and milk was readily detected. Thus, the test described here furthers our ability to monitor for toxin and improves the safety of the US food supply

Technical Abstract: Botulism is a serious foodborne neuroparalyic disease caused by botulinum neurotoxin (BoNT) produced by the anaerobic bacterium Clostridium botulinum. Seven toxin serotypes (A-H) have been described. The majority of human cases of botulism are caused by serotypes A and B followed by E and F. We report here a group of serotype B specific monoclonal antibodies (mAbs) capable of binding toxin under physiological conditions. Thus, they serve as capture antibodies for a sandwich (capture) ELISA. The antibodies were generated using recombinant peptide fragments corresponding to the receptor-binding domain of the toxin heavy chain as immunogen. Their binding properties suggest that they bind a complex epitope with KD ranging from 10–48 x 1011 M. The assay performance using all possible combinations of capture-detector antibody pairs was evaluated and the antibody pair resulting in the lowest level of detection (L.O.D.), ~ 20 pg/mL was identified. Toxin was detected in spiked dairy samples with good recoveries at concentrations as low as 0.5 pg/mL and in ground beef samples at levels as low as 2 ng/g. Thus, the sandwich ELISA described here uses mAb for both the capture and detector antibodies (binding different epitopes on the toxin molecule) and readily detects toxin in those food samples tested.

Last Modified: 9/22/2014
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