Rapid Detection of Botulinum Neurotoxins—A Review

Hobbs, Robert J.; Thomas, C.; Halliwell, Jennifer; Gwenin, Christopher

doi:10.3390/toxins11070418

Cited by 43 publications

(34 citation statements)

References 126 publications

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“…When selecting a BoNT detection assay, it is important to consider the end goal, such as laboratory diagnosis of botulism, detection in food or environmental samples, regulatory actions, pharmaceutical activity and properties, and toxicity evaluations of novel BoNTs [83,91].…”

Section: Considerations For Botulinum Neurotoxin Detection Methodsmentioning

confidence: 99%

“…However, these assays may not detect novel BoNTs and BoNTs for which the assays have not been validated, and require careful and thorough validation, ideally between independent laboratories. Several recent reviews provide detailed and comprehensive information on in vitro detection methods for BoNTs [83,84,87,88,91].…”

Section: Considerations For Botulinum Neurotoxin Detection Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Critical Analysis of Neuronal Cell and the Mouse Bioassay for Detection of Botulinum Neurotoxins

Pellett

Tepp

Johnson

2019

Toxins

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Botulinum Neurotoxins (BoNTs) are a large protein family that includes the most potent neurotoxins known to humankind. BoNTs delivered locally in humans at low doses are widely used pharmaceuticals. Reliable and quantitative detection of BoNTs is of paramount importance for the clinical diagnosis of botulism, basic research, drug development, potency determination, and detection in clinical, environmental, and food samples. Ideally, a definitive assay for BoNT should reflect the activity of each of the four steps in nerve intoxication. The in vivo mouse bioassay (MBA) is the 'gold standard' for the detection of BoNTs. The MBA is sensitive, robust, semi-quantitative, and reliable within its sensitivity limits. Potential drawbacks with the MBA include assay-to-assay potency variations, especially between laboratories, and false positives or negatives. These limitations can be largely avoided by careful planning and performance. Another detection method that has gained importance in recent years for research and potency determination of pharmaceutical BoNTs is cell-based assays, as these assays can be highly sensitive, quantitative, human-specific, and detect fully functional holotoxins at physiologically relevant concentrations. A myriad of other in vitro BoNT detection methods exist. This review focuses on critical factors and assay limitations of the mouse bioassay and cell-based assays for BoNT detection. Key Contribution:This manuscript reviews two widely used botulinum neurotoxin detection assays, the mouse bioassay and cell-based assays, and the critical factors involved in their methodology as well as interpretation of results. Despite our ability to now reduce animal use and replace the mouse bioassay for certain aspects of botulinum neurotoxin detection, the mouse bioassay is sensitive and is the only in vivo assay considering all aspects of botulinum neurotoxin intoxication on a physiological level as well as pharmacokinetic aspects of the toxins, and thus remains important assay for botulinum neurotoxin detection.Toxins 2019, 11, 713 2 of 23 cells, with preferential entry into motor-neurons. Inside a neuron, the toxins block neurotransmitter release, leading to the long-lasting descending bilateral paralysis characteristic of botulism [6]. Even if applied intramuscularly for therapeutic purposes, a portion of the injected BoNTs, in particular at high doses, can diffuse away from the local injection site leading to distal neuronal effects and at very high doses systemic distribution [7][8][9][10]. BoNTs can also enter human or vertebrate circulation and cause botulism by different routes, including ingestion of contaminated foods, by wound infection with neurotoxigenic clostridia, and by the colonization of the intestinal tract by neurotoxigenic clostridia, causing infant botulism or adult intestinal botulism [11,12]. The latter is rare, as C. botulinum usually does not colonize a healthy intestine with a mature microbiota. The severe symptoms of botulism last from several days to up to a year, depending ...

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Section: Considerations For Botulinum Neurotoxin Detection Methodsmentioning

confidence: 99%

Section: Considerations For Botulinum Neurotoxin Detection Methodsmentioning

confidence: 99%

Critical Analysis of Neuronal Cell and the Mouse Bioassay for Detection of Botulinum Neurotoxins

Pellett

Tepp

Johnson

2019

Toxins

View full text Add to dashboard Cite

show abstract

“…The mouse bioassay (MBA) is still considered to be the gold standard for laboratory testing due to its high sensitivity [27]. However, this method is costly, labor intensive and time-consuming, requires trained personnel and has ethical implications due to the use of live animals [29]. In cases of inhalational botulism, BoNT would not be detectable in serum or stool samples and the best approach then would be an ELISA test on nasal or broncho-alveolar lavage within 24 h of inhalation [28].…”

Section: Inhalational Botulismmentioning

confidence: 99%

Biosecurity Threat Posed by Botulinum Toxin

Cenciarelli

Riley

Baka

2019

Toxins

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The deliberate release of biological agents with terrorist or criminal intent continues to pose concerns in the current geopolitical situation. Therefore, attention is still needed to ensure preparedness against the potential use of pathogens as unconventional weapons. Botulinum neurotoxin (BoNT) is one such biological threat, characterized by an extremely low lethal dose, high morbidity and mortality when appropriately disseminated, and the capacity to cause panic and social disruption. This paper addresses the risks of a potential release of the botulinum neurotoxin and summarizes the relevant aspects of the threat.

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“…1f). Further optimization and characterization of these sensors would enable their use in multiple applications, such as rapid and low-cost detection of highly toxic botulinum neurotoxins in the food industry, which currently relies heavily on live-animal bioassays 18 , or detection of high serological levels of soluble Her2 (>15 ng/mL) associated with metastatic breast cancer 19 , levels that could be detected with the current sensitivity of lucCageHer2.…”

mentioning

confidence: 99%

De novodesign of modular and tunable allosteric biosensors

Quijano‐Rubio

Yeh

Park

et al. 2020

Preprint

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Naturally occurring allosteric protein switches have been repurposed for developing novel biosensors and reporters for cellular and clinical applications, but the number of such switches is limited, and engineering them is often challenging as each is different. Here, we show that a very general class of allosteric protein-based biosensors can be created by inverting the flow of information through de novo designed protein switches in which binding of a peptide key triggers biological outputs of interest. Using broadly applicable design principles, we allosterically couple binding of protein analytes of interest to the reconstitution of luciferase activity and a bioluminescent readout through the association of designed lock and key proteins. Because the sensor is based purely on thermodynamic coupling of analyte binding to switch activation, only one target binding domain is required, which simplifies sensor design and allows direct readout in solution. We demonstrate the modularity of this platform by creating biosensors that, with little optimization, sensitively detect the anti-apoptosis protein Bcl-2, the hIgG1 Fc domain, the Her2 receptor, and Botulinum neurotoxin B, as well as biosensors for cardiac Troponin I and an anti-Hepatitis B virus (HBV) antibody that achieve the sub-nanomolar sensitivity necessary to detect clinically relevant concentrations of these molecules. Given the current need for diagnostic tools for tracking COVID-19, we use the approach to design sensors of antibodies against SARS-CoV-2 protein epitopes and of the receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein. The latter, which incorporates a de novo designed RBD binder, has a limit of detection of 15pM with an up to seventeen fold increase in luminescence upon addition of RBD. The modularity and sensitivity of the platform should enable the rapid construction of sensors for a wide range of analytes and highlights the power of de novo protein design to create multi-state protein systems with new and useful functions.

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Rapid Detection of Botulinum Neurotoxins—A Review

Cited by 43 publications

References 126 publications

Critical Analysis of Neuronal Cell and the Mouse Bioassay for Detection of Botulinum Neurotoxins

Critical Analysis of Neuronal Cell and the Mouse Bioassay for Detection of Botulinum Neurotoxins

Biosecurity Threat Posed by Botulinum Toxin

De novodesign of modular and tunable allosteric biosensors

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