Detection of botulinum neurotoxin-A activity in food by peptide cleavage assay.
The World Health Organization (WHO) and U.S. Centers for Disease Control and Prevention (CDC) have labeled botulinum toxins as a high priority biological agent that may be used in terrorist attacks against food supplies. Due to this threat there is an increased need to develop fast and effective methods to detect active botulinum neurotoxins (BoNTs). This study reports the successful use of an enzymatic assay employing an internally quenched fluorogenic peptide as a fast, simple and inexpensive alternative to the mouse bioassay. In less than 15 min the assay can detect 0.25 nM BoNT-A in liquid food samples. The detection level is far below the adult human lethal oral dose of 70 microg of toxin. Immunomagnetic beads coated with IgG monoclonal antibodies that target the toxin heavy chain can concentrate the toxin without neutralizing its enzymatic activity, overcoming matrix effects caused by endogenous protease inhibitors and peptidases. This fast and effective assay system could be used for large scale screening to detect BoNT-A.
Western Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, Albany, California 94710, United States. reuven.rasooly@ars.usda.gov
Reuven
R
Rasooly
Larry H
LH
Stanker
John Mark
JM
Carter
Paula M
PM
Do
Luisa W
LW
Cheng
Xiaohua
X
He
David L
DL
Brandon
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D023482:Bioterrorism; D019274:Botulinum Toxins, Type A / Q000032:analysis* / Q000276:immunology / Q000633:toxicity; D003014:Clostridium botulinum / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005504:Food Analysis / Q000379:methods / Q000592:standards; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods* / Q000592:standards; D012680:Sensitivity and Specificity; D013997:Time Factors
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D019274:Botulinum Toxins, Type A