Development of an in vitro bioassay for Clostridium botulinum type B neurotoxin in foods that is more sensitive than the mouse bioassay

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 a ctivity of the neurotoxin's light chain and includes the following three st ages: first, a small, monoclonal antibody-based immunoaffinity column captu res the toxin; second, a peptide substrate is cleaved by using the endopept idase activity of the type B neurotoxin; and finally, a modified enzyme-lin ked immunoassay system detects the peptide cleavage products. The assay is highly specific for type B neurotoxin and is capable of detecting type B to xin 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 det ecting botulinum type B toxin in a range of foodstuffs with a sensitivity t hat exceeds the sensitivity of the mouse assay. Using highly specific monoc lonal antibodies as the capture phase, we found that the endopeptidase assa y was capable of differentiating between the type B neurotoxins produced by proteolytic and nonproteolytic strains of Clostridium botulinum type B.