期刊
TOXICOLOGY IN VITRO
卷 21, 期 8, 页码 1641-1649出版社
PERGAMON-ELSEVIER SCIENCE LTD
DOI: 10.1016/j.tiv.2007.06.015
关键词
tetanus neurotoxin; endopeptidase assay; botulinum neurotoxin type B; synaptobrevin-2; vesicle associated membrane protein; cleavage site specific antibody; ELISA
类别
Tetanus neurotoxin (TeNT1) is a bacterial protease which specifically cleaves the vesicle protein synaptobrevin-2 (vesicle associated membrane protein-2, VAMP-2). This proteolytic feature of the toxin has been used to develop a sensitive endopeptidase assay for the detection of TeNT activity as an alternative to the in vivo assay for TeNT toxicity. Recombinant synaptobrevin-2 (rSyb2) is immobilized onto a microtiter plate, and the cleavage of immobilized rSyb2 by TeNT is detected with a polyclonal antibody directed against the newly generated C-terminus of the cleavage product. This antibody is shown to be a highly specific tool for detecting rSyb2 proteolysis by TeNT. The method reaches a detection limit of less than 1 pg TeNT/ml. To our knowledge, this is the most sensitive in vitro assay for the detection of TeNT activity, and it is easy to perform. Besides, the assay can also detect the activity of botulinum neurotoxin type B (BoNT/B). The method can be applied to examine the toxicity of TeNT or BoNT/B preparations as well as the influence of chemicals on TeNT and BoNT/B activity. In the future, the assay may also serve as a basis for the replacement of the in vivo safety control of tetanus vaccines. (C) 2007 Elsevier Ltd. All rights reserved.
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