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

2 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.

LanguageEnglish
Pages912-919
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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