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Antibodies 2018, 7(4), 37; https://doi.org/10.3390/antib7040037

A Novel Panel of Rabbit Monoclonal Antibodies and Their Diverse Applications Including Inhibition of Clostridium perfringens Epsilon Toxin Oligomerization

1
Brain and Mind Research Institute, Weill Cornell Medical College of Cornell University, New York, NY, 10065, USA
2
Rockefeller University, New York, NY 10065, USA
3
Biogen, Cambridge, MA 02142, USA
*
Author to whom correspondence should be addressed.
Received: 28 August 2018 / Revised: 18 October 2018 / Accepted: 22 October 2018 / Published: 25 October 2018
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Abstract

The pore-forming epsilon toxin (ETX) produced by Clostridium perfringens is among the most lethal bacterial toxins known. Sensitive antibody-based reagents are needed to detect toxin, distinguish mechanisms of cell death, and prevent ETX toxicity. Using B-cell immuno-panning and cloning techniques, seven ETX-specific monoclonal antibodies were generated from immunized rabbits. ETX specificity and sensitivity were evaluated via western blot, ELISA, immunocytochemistry (ICC), and flow cytometry. ETX-neutralizing function was evaluated both in vitro and in vivo. All antibodies recognized both purified ETX and epsilon protoxin via western blot with two capable of detecting the ETX-oligomer complex. Four antibodies detected ETX via ELISA and three detected ETX bound to cells via ICC or flow cytometry. Several antibodies prevented ETX-induced cell death by either preventing ETX binding or by blocking ETX oligomerization. Antibodies that blocked ETX oligomerization inhibited ETX endocytosis and cellular vacuolation. Importantly, one of the oligomerization-blocking antibodies was able to protect against ETX-induced death post-ETX exposure in vitro and in vivo. Here we describe the production of a panel of rabbit monoclonal anti-ETX antibodies and their use in various biological assays. Antibodies possessing differential specificity to ETX in particular conformations will aid in the mechanistic studies of ETX cytotoxicity, while those with ETX-neutralizing function may be useful in preventing ETX-mediated mortality. View Full-Text
Keywords: Clostridium perfringens; epsilon toxin; epsilon protoxin; neutralizing; antibodies; oligomerization; pore formation Clostridium perfringens; epsilon toxin; epsilon protoxin; neutralizing; antibodies; oligomerization; pore formation
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Linden, J.R.; Telesford, K.; Shetty, S.; Winokour, P.; Haigh, S.; Cahir-McFarland, E.; Antognetti, G.; Datta, A.; Wang, T.; Meier, W.; Vartanian, T. A Novel Panel of Rabbit Monoclonal Antibodies and Their Diverse Applications Including Inhibition of Clostridium perfringens Epsilon Toxin Oligomerization. Antibodies 2018, 7, 37.

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