title: A monoclonal antibody based capture ELISA for botulinum neurotoxin serotype B: toxin detection in food.
abstract: Botulism is a serious foodborne neuroparalytic disease, caused by botulinum neurotoxin (BoNT), produced by the anaerobic bacterium Clostridium botulinum. Seven toxin serotypes (A-H) have been described. The majority of human cases of botulism are caused by serotypes A and B followed by E and F. We report here a group of serotype B specific monoclonal antibodies (mAbs) capable of binding toxin under physiological conditions. Thus, they serve as capture antibodies for a sandwich (capture) ELISA. The antibodies were generated using recombinant peptide fragments corresponding to the receptor-binding domain of the toxin heavy chain as immunogen. Their binding properties suggest that they bind a complex epitope with dissociation constants (KD's) for individual antibodies ranging from 10 to 48 × 10-11 M. Assay performance for all possible combinations of capture-detector antibody pairs was evaluated and the antibody pair resulting in the lowest level of detection (L.O.D.), ~20 pg/mL was determined. Toxin was detected in spiked dairy samples with good recoveries at concentrations as low as 0.5 pg/mL and in ground beef samples at levels as low as 2 ng/g. Thus, the sandwich ELISA described here uses mAb for both the capture and detector antibodies (binding different epitopes on the toxin molecule) and readily detects toxin in those food samples tested.
authors.affiliation: Foodborne Toxin Detection and Prevention Unit, United States Department of Agriculture, Agricultural Research Service, 800 Buchanan Street, Albany, CA 94510, USA. larry.stanker@ars.usda.gov.
authors.forename: Larry H
authors.initials: LH
authors.lastname: Stanker
authors.forename: Miles C
authors.initials: MC
authors.lastname: Scotcher
authors.forename: Luisa
authors.initials: L
authors.lastname: Cheng
authors.forename: Kathryn
authors.initials: K
authors.lastname: Ching
authors.forename: Jeffery
authors.initials: J
authors.lastname: McGarvey
authors.forename: David
authors.initials: D
authors.lastname: Hodge
authors.forename: Robert
authors.initials: R
authors.lastname: Hnasko
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000302:isolation & purification; D001905:Botulinum Toxins / Q000737:chemistry / Q000302:isolation & purification*; D019274:Botulinum Toxins, Type A; D002417:Cattle; D003014:Clostridium botulinum; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D000939:Epitopes / Q000737:chemistry; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology; D006801:Humans; D008460:Meat / Q000382:microbiology; D008892:Milk / Q000382:microbiology; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000737:chemistry
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D011993:Recombinant Fusion Proteins; C096323:rimabotulinumtoxinB; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A