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Title: Renewable Surface Fluorescence Sandwich Immunoassay Biosensor for Rapid Sensitive Botulinum Toxin Detection in an Automated Fluidic Format

Abstract

A renewable surface biosensor for rapid detection of botulinum toxin is described based on fluidic automation of a fluorescence sandwich immunoassay, using a recombinant fragment of the toxin heavy chain as a structurally valid simulant. Monoclonal antibodies AR4 and RAZ1 bind to separate epitopes of both this fragment and the holotoxin. The AR4 antibody was covalently bound to Sepharose beads and used as the capture antibody. A rotating rod flow cell was used to capture these beads delivered as a suspension by the sequential injection flow system, creating a 3.6 microliter column. After perfusing the bead column with sample and washing away the matrix, the column was perfused with Alexa 647 dye-labeled RAZ1 antibody as the reporter. Optical fibers coupled to the rotating rod flow cell at a 90 degree angle to one another delivered excitation light from a HeNe laser and collected fluorescent emission light for detection. After each measurement, the used sepharose beads are released and replaced with fresh beads. In a rapid screening approach to sample analysis, the toxin simulant was detected to concentrations of 10 pM in less than 20 minutes.

Authors:
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Publication Date:
Research Org.:
Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
Sponsoring Org.:
USDOE
OSTI Identifier:
951823
Report Number(s):
PNNL-SA-63257
Journal ID: ISSN 0003-2654; ANALAO; 30400; 400904120; TRN: US200913%%66
DOE Contract Number:  
AC05-76RL01830
Resource Type:
Journal Article
Journal Name:
Analyst, 134(5):987 - 996
Additional Journal Information:
Journal Volume: 134; Journal Issue: 5; Journal ID: ISSN 0003-2654
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; SENSORS; DESIGN; AUTOMATION; TOXINS; DETECTION; FLUORESCENCE; IMMUNOASSAY; MONOCLONAL ANTIBODIES; OPTICAL FIBERS; EXTRACTION COLUMNS; MINIATURIZATION; Environmental Molecular Sciences Laboratory

Citation Formats

Grate, Jay W, Warner, Marvin G, Ozanich, Richard M, Miller, Keith D, Colburn, Heather A, Dockendorff, Brian P, Antolick, Kathryn C, Anheier, Norman C, Lind, Michael A, Lou, Jianlong, Marks, James D, and Bruckner-Lea, Cindy J. Renewable Surface Fluorescence Sandwich Immunoassay Biosensor for Rapid Sensitive Botulinum Toxin Detection in an Automated Fluidic Format. United States: N. p., 2009. Web. doi:10.1039/B900794F.
Grate, Jay W, Warner, Marvin G, Ozanich, Richard M, Miller, Keith D, Colburn, Heather A, Dockendorff, Brian P, Antolick, Kathryn C, Anheier, Norman C, Lind, Michael A, Lou, Jianlong, Marks, James D, & Bruckner-Lea, Cindy J. Renewable Surface Fluorescence Sandwich Immunoassay Biosensor for Rapid Sensitive Botulinum Toxin Detection in an Automated Fluidic Format. United States. https://doi.org/10.1039/B900794F
Grate, Jay W, Warner, Marvin G, Ozanich, Richard M, Miller, Keith D, Colburn, Heather A, Dockendorff, Brian P, Antolick, Kathryn C, Anheier, Norman C, Lind, Michael A, Lou, Jianlong, Marks, James D, and Bruckner-Lea, Cindy J. Thu . "Renewable Surface Fluorescence Sandwich Immunoassay Biosensor for Rapid Sensitive Botulinum Toxin Detection in an Automated Fluidic Format". United States. https://doi.org/10.1039/B900794F.
@article{osti_951823,
title = {Renewable Surface Fluorescence Sandwich Immunoassay Biosensor for Rapid Sensitive Botulinum Toxin Detection in an Automated Fluidic Format},
author = {Grate, Jay W and Warner, Marvin G and Ozanich, Richard M and Miller, Keith D and Colburn, Heather A and Dockendorff, Brian P and Antolick, Kathryn C and Anheier, Norman C and Lind, Michael A and Lou, Jianlong and Marks, James D and Bruckner-Lea, Cindy J},
abstractNote = {A renewable surface biosensor for rapid detection of botulinum toxin is described based on fluidic automation of a fluorescence sandwich immunoassay, using a recombinant fragment of the toxin heavy chain as a structurally valid simulant. Monoclonal antibodies AR4 and RAZ1 bind to separate epitopes of both this fragment and the holotoxin. The AR4 antibody was covalently bound to Sepharose beads and used as the capture antibody. A rotating rod flow cell was used to capture these beads delivered as a suspension by the sequential injection flow system, creating a 3.6 microliter column. After perfusing the bead column with sample and washing away the matrix, the column was perfused with Alexa 647 dye-labeled RAZ1 antibody as the reporter. Optical fibers coupled to the rotating rod flow cell at a 90 degree angle to one another delivered excitation light from a HeNe laser and collected fluorescent emission light for detection. After each measurement, the used sepharose beads are released and replaced with fresh beads. In a rapid screening approach to sample analysis, the toxin simulant was detected to concentrations of 10 pM in less than 20 minutes.},
doi = {10.1039/B900794F},
url = {https://www.osti.gov/biblio/951823}, journal = {Analyst, 134(5):987 - 996},
issn = {0003-2654},
number = 5,
volume = 134,
place = {United States},
year = {2009},
month = {3}
}