Microcystin-LR Enrichment from Freshwater by a Recombinant Plant-derived Antibody Using Sol-Gel-Glass Immunoextraction.

Eutrophication of water bodies can promote cyanobacterial (blue-green algae) blooms, which has become a source of increasing concern for both recreational and drinking water use. Many bacterial species can produce toxins that pose threats to wildlife, domestic animals and humans. Microcystin-leucine-arginine (MC-LR) is the most frequent and most toxic microcystin congener. For the first time, lab-scale investigations were performed to test the application of a recombinant plant-derived anti-MC-LR antibody immobilized on an immunoaffinity support material to selectively extract the toxin from spiked freshwater samples. As a comparison, its hybridoma-derived counterpart (murine monoclonal antibody) was evaluated. The antibody-doped material was prepared via an optimized sol-gel process; its stability and binding efficiency of MC-LR in spiked freshwater samples were thoroughly tested using the ELISA and orthogonal LC-MS methods. For removal, two column-based procedures with sequential or continuous cyclic sample addition and a suspension mode (moving adsorbent) were tested. Noteworthy the results obtained with a crude antibody fraction were fully compatible with the highly purified preparation. This study paves the way for further investigation being focused on novel applications of plant-derived anti-MC-LR antibodies in bioremediation to selectively deplete the toxin from freshwater: a green and promising technology without secondary pollution.

Functional analysis of the broadly neutralizing human anti-HIV-1 antibody 2F5 produced in transgenic BY-2 suspension cultures.

We report the production of an important human therapeutic antibody in plant cell suspension cultures and the functional analysis of that antibody, including a comparison with the same antibody produced in CHO cells. We established transgenic tobacco BY2 suspension cell cultures expressing the human monoclonal antibody 2F5, which shows broadly neutralizing activity against HIV-1. The antibody was directed to the endoplasmic reticulum of the plant cells and was isolated by cell disruption, followed by protein A chromatography. The plant-derived antibody was shown to be largely intact by SDS-PAGE and immunoblot. Antigen binding activity was investigated by electrophoretic mobility shift assay and quantitatively determined by ELISA and Biacore biosensor technology. Ligand binding properties were analyzed using the ectodomain of human Fc gammaRI for kinetic analysis. The plant-derived antibody showed similar kinetic properties and 89% of the binding capacity of its CHO-derived counterpart, but was only 33% as efficient in HIV-1 neutralization assays. Our results show that plant suspension cultures can be used to produce human antibodies efficiently and that the analysis methods used in this study, including biosensor technology, provide useful functional data about antibody performance. This highlights important issues raised by the use of plant systems to produce human biologics.