Production of monoclonal antibodies against botulinum neurotoxin in Nicotiana benthamiana

Abstract

Botulism is a fatal neurologic disease caused by the botulinum toxin (BoNT) produced by Clostridium botulinum . It is a rare but highly toxic disease with symptoms, such as cramps, nausea, vomiting, diarrhea, dysphagia, respiratory failure, muscle weakness, and even death. Currently, two types of antitoxin are used: equine-derived heptavalent antitoxin and human-derived immunoglobulin (BabyBIG®). However, heptavalent treatment may result in hypersensitivity, whereas BabyBIG®, has a low yield. The present study focused on the development of three anti-BoNT monoclonal antibodies (mAbs), 1B18, C25, and M2, in Nicotiana benthamiana . The plant-expressed mAbs were purified and examined for size, purity and integrity by SDS-PAGE, western blotting and size-exclusion chromatography. Analysis showed that plant-produced anti-BoNT mAbs can fully assemble in plants, can be purified in a single purification step, and mostly remain as monomeric proteins. The efficiency of anti-BoNT mAbs binding to BoNT/A and B was then tested. Plant-produced 1B18 retained its ability to recognize both mBoNT/A1 and ciBoNT/B1. At the same time, the binding specificities of two other mAbs were determined: C25 for mBoNT/A1 and M2 for ciBoNT/B1. In conclusion, our results confirm the use of plants as an alternative platform for the production of anti-BoNT mAbs. This plant-based technology will serve as a versatile system for the development botulism immunotherapeutics.

Nicotiana benthamiana as a potential source for producing anti-dengue virus D54 neutralizing therapeutic antibody

“…The produced protein by this method is sufficiently supplied throughout clinical study. This method is also employed in the production of therapeutic proteins like anti-human PD-1 antibodies [ 48 ] and anti-botulinum neurotoxins [ 49 ] which are previously developed in our laboratory. Therefore, using agroinfiltration for the production of D54 mAb is more appropriate than stable transformation.…”