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Applied and Environmental Microbiology, September 1999, p. 3787-3792, Vol. 65, No. 9
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Development of an In Vitro Bioassay for Clostridium botulinum Type B Neurotoxin in Foods That Is More Sensitive than the Mouse Bioassay

Matthew Wictome,1,* Kirsti Newton,1 Karen Jameson,1 Bassam Hallis,1 Paul Dunnigan,2 Eric Mackay,2 Sally Clarke,3 Richard Taylor,3 Joy Gaze,3 Keith Foster,1 and Clifford Shone1

Centre for Applied Microbiology and Research, Porton Down, Salisbury, Wiltshire, SP4 OJG,1 Rhône Diagnostics Technologies, West of Scotland Science Park, Glasgow, G20 OSP,2 and Campden and Chorleywood Food Research Association, Chipping Campden, GL55 6LD,3 United Kingdom

Received 11 January 1999/Accepted 12 June 1999

A novel, in vitro bioassay for detection of the botulinum type B neurotoxin in a range of media was developed. The assay is amplified by the enzymic activity of the neurotoxin's light chain and includes the following three stages: first, a small, monoclonal antibody-based immunoaffinity column captures the toxin; second, a peptide substrate is cleaved by using the endopeptidase activity of the type B neurotoxin; and finally, a modified enzyme-linked immunoassay system detects the peptide cleavage products. The assay is highly specific for type B neurotoxin and is capable of detecting type B toxin at a concentration of 5 pg ml-1 (0.5 mouse 50% lethal dose ml-1) in approximately 5 h. The format of the test was found to be suitable for detecting botulinum type B toxin in a range of foodstuffs with a sensitivity that exceeds the sensitivity of the mouse assay. Using highly specific monoclonal antibodies as the capture phase, we found that the endopeptidase assay was capable of differentiating between the type B neurotoxins produced by proteolytic and nonproteolytic strains of Clostridium botulinum type B.

* Corresponding author. Mailing address: Centre for Applied Microbiology and Research, Porton Down, Salisbury, Wiltshire, SP4 OJG, United Kingdom. Phone: 44 1980 612626. Fax: 44 1980 611310. E-mail: matt.wictome{at}camr.org.uk.

Applied and Environmental Microbiology, September 1999, p. 3787-3792, Vol. 65, No. 9
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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