Optimization of SNAP-25 and VAMP-2 Cleavage by Botulinum Neurotoxin Serotypes A–F Employing Taguchi Design-of-Experiments

Berg, Laura von; Stern, Daniel; Weisemann, Jasmin; Rummel, Andreas; Dorner, Martin B.; Dorner, Brigitte G.

doi:10.3390/toxins11100588

Cited by 3 publications

(1 citation statement)

References 73 publications

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“…A collaborative study organized by the FDA and the CDC (Centers for Disease Control and Prevention) to assess the performance of ELISA kits to detect BoNT toxinotypes A, B, E, and F in various food matrices has established the suitability of ELISA kits to detect BoNTs in food samples [ 28 , 29 ]. The most recent and sophisticated version of the ELISA method, the Luminex assay, uses microsphere beads conjugated to antibodies; it has shown better limits of detection than the MBA [ 30 , 31 ].…”

Section: Bont Detectionmentioning

confidence: 99%

Recent Developments in Botulinum Neurotoxins Detection

Rasetti-Escargueil

Popoff

2022

Microorganisms

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Botulinum neurotoxins (BoNTs) are produced as protein complexes by bacteria of the genus Clostridium that are Gram-positive, anaerobic and spore forming (Clostridium botulinum, C. butyricum, C. baratii and C. argentinense spp.). BoNTs show a high immunological and genetic diversity. Therefore, fast, precise, and more reliable detection methods are still required to monitor outbreaks and ensure surveillance of botulism. The botulinum toxin field also comprises therapeutic uses, basic research studies and biodefense issues. This review presents currently available detection methods, and new methods offering the potential of enhanced precision and reproducibility. While the immunological methods offer a range of benefits, such as rapid analysis time, reproducibility and high sensitivity, their implementation is subject to the availability of suitable tools and reagents, such as specific antibodies. Currently, the mass spectrometry approach is the most sensitive in vitro method for a rapid detection of active or inactive forms of BoNTs. However, these methods require inter-laboratory validation before they can be more widely implemented in reference laboratories. In addition, these surrogate in vitro models also require full validation before they can be used as replacement bioassays of potency. Cell-based assays using neuronal cells in culture recapitulate all functional steps of toxin activity, but are still at various stages of development; they are not yet sufficiently robust, due to high batch-to-batch cell variability. Cell-based assays have a strong potential to replace the mouse bioassay (MBA) in terms of BoNT potency determination in pharmaceutical formulations; they can also help to identify suitable inhibitors while reducing the number of animals used. However, the development of safe countermeasures still requires the use of in vivo studies to complement in vitro immunological or cell-based approaches.

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Section: Bont Detectionmentioning

confidence: 99%

Recent Developments in Botulinum Neurotoxins Detection

Rasetti-Escargueil

Popoff

2022

Microorganisms

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Neutralizing chimeric heavy-chain antibody targeting the L-HN domain of Clostridium botulinum neurotoxin type F

Sun,

Luo,

Jiang

et al. 2024

Arch Toxicol

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A versatile immunoassay based on functionalized nanoparticles for botulinum neurotoxin detection and sensor development

Nguyen,

et al. 2024

Discov Appl Sci

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Botulinum neurotoxins (BoNTs) serotypes A and B are the two most common of the four BoNTs that cause the high mortality botulism disease in individuals consuming contaminated foods. The gold standard assay for BoNT detection is the live mouse bioassay, which has several major disadvantages, including tedious procedures and animal sacrifice requirements. In this study, we developed an immuno-based assay using magnetic streptavidin nanoparticles (mSNP) functionalized with specific synthetic biotinylated, 6xHis-tagged peptide substrates (Peptides A, PA and B, PB) designed for BoNT/A and BoNT/B proteolytic reactions, respectively. In the presence of active toxins that possess endopeptidase activity, upon cleavage, the released fragments with His-tag were dotted on a blotting membrane, ultimately producing color signals after incubation with anti-His antibody, alkaline phosphatase (AP)-conjugated antibody, and then AP substrates. The results showed that the efficiency of peptide-mSNP complex formation reached up to 81%, and the dot blot immunoassay allowed peptide detection from 10 ng of His-tagged peptides. Preliminary testing with the extracellular extracts from the isolated Clostridium botulinum strains indicated that the botulinum toxin in the 2020 botulism outbreak in Vietnam belonged to serotype A, the most potent BoNT. The established assay could be applied to construct a portable biosensor for BoNT detection and a high throughput device to screen potential BoNT inhibitors for drug development.

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Optimization of SNAP-25 and VAMP-2 Cleavage by Botulinum Neurotoxin Serotypes A–F Employing Taguchi Design-of-Experiments

Cited by 3 publications

References 73 publications

Recent Developments in Botulinum Neurotoxins Detection

Recent Developments in Botulinum Neurotoxins Detection

Neutralizing chimeric heavy-chain antibody targeting the L-HN domain of Clostridium botulinum neurotoxin type F

A versatile immunoassay based on functionalized nanoparticles for botulinum neurotoxin detection and sensor development

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