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Anal Chem. 2014 Mar 4;86(5):2279-84. doi: 10.1021/ac500262d. Epub 2014 Feb 24.

High sensitivity detection of active botulinum neurotoxin by glyco-quantitative polymerase chain-reaction.

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1
Department of Chemical and Biological Engineering, Department of Chemistry and Chemical Biology, and Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute , Troy, New York 12180, United States.

Abstract

The sensitive detection of highly toxic botulinum neurotoxin (BoNT) from Clostridium botulinum is of critical importance because it causes human illnesses if foodborne or introduced in wounds and as an iatrogenic substance. Moreover, it has been recently considered a possible biological warfare agent. Over the past decade, significant progress has been made in BoNT detection technologies, including mouse lethality assays, enzyme-linked immunosorbent assays, and endopeptidase assays and by mass spectrometry. Critical assay requirements, including rapid assay, active toxin detection, sensitive and accurate detection, still remain challenging. Here, we present a novel method to detect active BoNTs using a Glyco-quantitative polymerase chain-reaction (qPCR) approach. Sialyllactose, which interacts with the binding-domain of BoNTs, is incorporated into a sialyllactose-DNA conjugate as a binding-probe for active BoNT and recovered through BoNT-immunoprecipitation. Glyco-qPCR analysis of the bound sialyllactose-DNA is then used to detect low attomolar concentrations of BoNT and attomolar to femtomolar concentrations of BoNT in honey, the most common foodborne source of infant botulism.

PMID:
24506443
PMCID:
PMC3985614
DOI:
10.1021/ac500262d
[Indexed for MEDLINE]
Free PMC Article
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