Detection of botulinal neurotoxins A, B, E, and F by amplified enzyme-linked immunosorbent assay: collaborative study

An amplified enzyme-linked immunosorbent assay (amp-ELISA) was compared with the AOAC Official Method 977.26 for detection of Clostridium botulinum and its toxins in foods. Eleven laboratories participated and the results of 10 laboratories were used in the study. Two anaerobic culture media, tryptone peptone glucose yeast extract (TPGY) and cooked meat medium (CMM) were used to generate toxic samples with types A, B, E, and F botulinal strains. Nonbotulinal clostridia were also tested. The toxicity of each botulinal culture was determined by the AOAC method, and the cultures were then diluted, if necessary, to high (about 10,000 minimal lethal dose [MLD]/mL) and low (about 100 MLD/mL) test samples. The overall sensitivity of detection in TPGY and CMM cultures with the amp-ELISA was 94.7% at about 100 MLD/mL and 99.6% for samples with > or = 10,000 MLD/mL toxicity. The amp-ELISA detection sensitivity for low toxin samples was 92.3% in TPGY and 99.4% in CMM. The false-positive rate ranged from 1.5% for type A to 28.6% for type F in TPGY, and from 2.4% for type A to 11.4% for type F in CMM. Most of the cross-reactivity was due to detection of other botulinal types, especially in high toxin samples. The amp-ELISA could be used to screen suspect cultures for botulinal toxins. Positive amp-ELISA samples would be confirmed by the AOAC reference method.