title: Extraction and inhibition of enzymatic activity of botulinum neurotoxins/A1, /A2, and /A3 by a panel of monoclonal anti-BoNT/A antibodies.
abstract: Botulinum neurotoxins (BoNTs) are extremely potent toxins that are capable of causing death or respiratory failure leading to long-term intensive care. Treatment includes serotype-specific antitoxins, which must be administered early in the course of the intoxication. Rapidly determining human exposure to BoNT is an important public health goal. In previous work, our laboratory focused on developing Endopep-MS, a mass spectrometry-based endopeptidase method for detecting and differentiating BoNT/A-G serotypes in buffer and BoNT/A, /B, /E, and /F in clinical samples. We have previously reported the effectiveness of antibody-capture to purify and concentrate BoNTs from complex matrices, such as clinical samples. Because some antibodies inhibit or neutralize the activity of BoNT, the choice of antibody with which to extract the toxin is critical. In this work, we evaluated a panel of 16 anti-BoNT/A monoclonal antibodies (mAbs) for their ability to inhibit the in vitro activity of BoNT/A1, /A2, and /A3 complex as well as the recombinant LC of A1. We also evaluated the same antibody panel for the ability to extract BoNT/A1, /A2, and /A3. Among the mAbs, there were significant differences in extraction efficiency, ability to extract BoNT/A subtypes, and inhibitory effect on BoNT catalytic activity. The mAbs binding the C-terminal portion of the BoNT/A heavy chain had optimal properties for use in the Endopep-MS assay.
authors.affiliation: Centers for Disease Control and Prevention, National Center for Environmental Health, Division of Laboratory Sciences, Atlanta, GA, USA.
authors.forename: Suzanne R
authors.initials: SR
authors.lastname: Kalb
authors.forename: Jianlong
authors.initials: J
authors.lastname: Lou
authors.forename: Consuelo
authors.initials: C
authors.lastname: Garcia-Rodriguez
authors.forename: Isin N
authors.initials: IN
authors.lastname: Geren
authors.forename: Theresa J
authors.initials: TJ
authors.lastname: Smith
authors.forename: Hercules
authors.initials: H
authors.lastname: Moura
authors.forename: James D
authors.initials: JD
authors.lastname: Marks
authors.forename: Leonard A
authors.initials: LA
authors.lastname: Smith
authors.forename: James L
authors.initials: JL
authors.lastname: Pirkle
authors.forename: John R
authors.initials: JR
authors.lastname: Barr
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D019274:Botulinum Toxins, Type A / Q000037:antagonists & inhibitors* / Q000145:classification / Q000276:immunology / Q000302:isolation & purification*; D001906:Botulism / Q000175:diagnosis / Q000628:therapy; D047013:Clostridium botulinum type A / Q000276:immunology / Q000472:pathogenicity; D006801:Humans; D051379:Mice; D012703:Serotyping; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D019274:Botulinum Toxins, Type A