Functional fluorescence assay of botulinum neurotoxin A in complex matrices using magnetic beads

Nevena Klisara, Jeroen Peters, Willem Haasnoot, Michel W.F. Nielen, Alagappan Palaniappan, Bo Liedberg

Research output: Contribution to journalArticleAcademicpeer-review

3 Citations (Scopus)

Abstract

The extremely toxic botulinum neurotoxin poses a threat for health and food safety, requiring rapid and easy-to-use detection platforms. To meet these requirements, we have explored a novel functional assay format for detection of botulinum neurotoxin serotype A light chain (BoNT/A LC) in complex matrices. The proposed assay utilizes a synthetic peptide designed to mimic the SNAP-25 protein (synaptosomal-associated protein 25) as substrate bound to a superparamagnetic bead and a fluorescent dye. Cleavage of the peptide by BoNT/A LC yields a reduction in fluorescence signal revealing the presence of the BoNT/A LC in the sample matrices tested. The superparamagnetic beads enable efficient separation of the cleaved peptides from food matrices, thereby improving the reliability of responses. Herein, we demonstrate a protocol offering an assay time of 6 h and a LOD of 0.5 nM (25 ng/ml). The proposed protocol is validated using carrot juice and diluted milk pending further improvements in sensitivity and overall assay time. Robustness, cost-effectiveness, low sample volume requirements in conjunction with the possibility of multiplexing for other proteolytic enzymes makes the proposed protocol competitive in comparison with conventional BoNT assays reported elsewhere.

Original languageEnglish
Pages (from-to)912-919
JournalSensors and Actuators, B: Chemical
Volume281
DOIs
Publication statusPublished - 15 Feb 2019

Fingerprint

Type A Botulinum Toxins
beads
peptides
Neurotoxins
Assays
Fluorescence
food
fluorescence
Synaptosomal-Associated Protein 25
Peptides
matrices
SNAP
juices
proteins
requirements
cost effectiveness
milk
multiplexing
health
format

Keywords

  • Botulinum neurotoxin A
  • Fluorescence detection
  • Peptide
  • Superparamagnetic beads

Cite this

Klisara, Nevena ; Peters, Jeroen ; Haasnoot, Willem ; Nielen, Michel W.F. ; Palaniappan, Alagappan ; Liedberg, Bo. / Functional fluorescence assay of botulinum neurotoxin A in complex matrices using magnetic beads. In: Sensors and Actuators, B: Chemical. 2019 ; Vol. 281. pp. 912-919.
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Functional fluorescence assay of botulinum neurotoxin A in complex matrices using magnetic beads. / Klisara, Nevena; Peters, Jeroen; Haasnoot, Willem; Nielen, Michel W.F.; Palaniappan, Alagappan; Liedberg, Bo.

In: Sensors and Actuators, B: Chemical, Vol. 281, 15.02.2019, p. 912-919.

Research output: Contribution to journalArticleAcademicpeer-review

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AU - Klisara, Nevena

AU - Peters, Jeroen

AU - Haasnoot, Willem

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AU - Palaniappan, Alagappan

AU - Liedberg, Bo

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