Monoclonal antibodies and reagents for botulinum research

Abstract

Botulinum neurotoxin (BoNT) is produced by Clostridium botulinum as a dichain protein of ~ 150 kDa that blocks acetylcholine release resulting in muscular paralysis. Diagnosis of BoNT often relies on the mouse bioassay that has a detection limit of 10-20 pg/mL and can take up to four days to complete. Rapid in vitro methods for toxin detection are needed and to date, most rely on either immunoassay or endopeptidase activity. In the latter, many also use specific antibodies to concentrate the toxin. We have developed panels of monoclonal antibodies (mAbs) to purified toxin, serotypes A, B and E, as well as mAbs to the non-toxic associated proteins. Application of these mAbs in sandwich ELISAs and assay performance in milk and other foods is discussed. The assays described here are able to detect toxin in a few hours, at levels lower than the mouse bioassay.

Detection of Botulinum Neurotoxin Serotypes A and B Using a Chemiluminescent versus Electrochemiluminescent Immunoassay in Food and Serum

“…A combination (1:1:1) of biotinylated mAbs F1-40, F1-41 and F2-43 each at 1 μg/mL gave the best detection sensitivities for BoNT/A compared to single or a sets of two mAbs. 19 For BoNT/B detection, mAb MCS6-27 was used as the capture mAb and biotinylated mAb BoB92-32 was used the detector mAb. Only a single mAb was available for BoNT/B detection at this time.…”

“…To simulate the effect of polyclonal antibodies, we tested the use of a mixture of mAbs as detector antibodies. A combination (1:1:1) of biotinylated mAbs F1-40, F1-51, and F2-43 each at 1 μg/mL gave the best detection sensitivities for BoNT/A as compared to single or a sets of two mAbs . For BoNT/B detection, mAb MCS6-27 was used as the capture mAb, and biotinylated mAb BoB92-32 was used the detector mAb.…”

“…Fresh aliquots were used for each experiment. mAbs for BoNT/A F1-2, F1-40, F2-43, and F1-51 were described previously. ,, Anti-BoNT/B mAb MCS 6-27 was described previously, and mAb BoB92-32 is a newly developed antibody from our laboratory (unpublished results).…”

Botulinum Neurotoxin Detection Methods for Public Health Response and Surveillance

“…Sensitive and reliable immunoassay platforms could be ideally used for detecting the presence of toxin antigens in in food samples for surveillance purpose or in clinical specimens alongside clinical diagnosis, to substantially reduce false negative or inconclusive results obtained from clinical specimens. Unlike the MLB, which relies on sample dilution (Stanker and Cheng, 2012 ), immunoassays have an excellent dynamic range of quantification (Worbs et al, 2015 ). Immunoassays are more cost effective, requiring fewer dedicated personnel, less instrumentation and are not skill intensive (Lindstrom and Korkeala, 2006 ; Cai et al, 2007 ; Capek and Dickerson, 2010 ).…”