Extraction and Inhibition of Enzymatic Activity of Botulinum Neurotoxins/A1, /A2, and /A3 by a Panel of Monoclonal Anti-BoNT/A Antibodies

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. Rapid determination of human exposure to BoNT is an important public health goal. Previous work in our laboratory focused on the development of Endopep-MS, a mass spectrometry-based endopeptidase method for detecting and differentiating /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 monoclonal anti-BoNT/A antibodies (mAbs) for their ability to inhibit the in vitro activity of BoNT/A1, /A2, and /A3 complex as well as the recombinant light chain of A1. We also evaluated the same antibody panel for the ability to extract BoNT/A1, /A2, and /A3. MAbs differed significantly with respect to their extraction efficiency, their ability to extract BoNT/A subtypes, and their inhibitory effect on BoNT catalytic activity. MAbs binding the C-terminal portion of the BoNT/A heavy chain were found to have the optimal properties for use in the Endopep-MS assay.
Published in the Public Library of Science (PLoS ONE), v4 i4 e5355, Apr 2009. Prepared in cooperation with Army Medical Research Institute of Infectious Diseases, Ft. Detrick, MD. Prepared in collaboration with University of California at San Francisco. The original document contains color images.