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Toxins 2018, 10(9), 360; https://doi.org/10.3390/toxins10090360

Cell-Based Reporter Release Assay to Determine the Potency of Proteolytic Bacterial Neurotoxins

1
Department of Nutritional Biochemistry, Institute of Nutritional Science, University of Potsdam; Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany
2
Croma-Pharma GmbH, Cromazeile 2, A-2100 Leobendorf, Austria
*
Author to whom correspondence should be addressed.
Received: 16 August 2018 / Revised: 31 August 2018 / Accepted: 3 September 2018 / Published: 5 September 2018
(This article belongs to the Section Bacterial Toxins)
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Abstract

Despite the implementation of cell-based replacement methods, the mouse lethality assay is still frequently used to determine the activity of botulinum toxin (BoNT) for medical use. One explanation is that due to the use of neoepitope-specific antibodies to detect the cleaved BoNT substrate, the currently devised assays can detect only one specific serotype of the toxin. Recently, we developed a cell-based functional assay, in which BoNT activity is determined by inhibiting the release of a reporter enzyme that is liberated concomitantly with the neurotransmitter from neurosecretory vesicles. In theory, this assay should be suitable to detect the activity of any BoNT serotype. Consistent with this assumption, the current study shows that the stimulus-dependent release of a luciferase from a differentiated human neuroblastoma-based reporter cell line (SIMA-hPOMC1-26-GLuc cells) was inhibited by BoNT-A and-C. Furthermore, this was also inhibited by BoNT-B and tetanus toxin to a lesser extent and at higher concentrations. In order to provide support for the suitability of this technique in practical applications, a dose–response curve obtained with a pharmaceutical preparation of BoNT-A closely mirrored the activity determined in the mouse lethality assay. In summary, the newly established cell-based assay may represent a versatile and specific alternative to the mouse lethality assay and other currently established cell-based assays. View Full-Text
Keywords: botulinum toxin; BoNT; tetanus toxin; RRR; replacement botulinum toxin; BoNT; tetanus toxin; RRR; replacement
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).
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Pathe-Neuschäfer-Rube, A.; Neuschäfer-Rube, F.; Haas, G.; Langoth-Fehringer, N.; Püschel, G.P. Cell-Based Reporter Release Assay to Determine the Potency of Proteolytic Bacterial Neurotoxins. Toxins 2018, 10, 360.

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