High-throughput screening of monoclonal antibodies against plant cell wall glycans by hierarchical clustering of their carbohydrate microarray binding profiles.
Isabel Moller, Susan E Marcus, Ash Haeger, Yves Verhertbruggen, Rene Verhoef, Henk Schols, Peter Ulvskov, Jørn Dalgaard Mikkelsen, J Paul Knox, William Willats
Antibody-producing hybridoma cell lines were created following immunisation with a crude extract of cell wall polymers from the plant Arabidopsis thaliana. In order to rapidly screen the specificities of individual monoclonal antibodies (mAbs), their binding to microarrays containing 50 cell wall glycans immobilized on nitrocellulose was assessed. Hierarchical clustering of microarray binding profiles from newly produced mAbs, together with the profiles for mAbs with previously defined specificities allowed the rapid assignments of mAb binding to antigen classes. mAb specificities were further investigated using subsequent immunochemical and biochemical analyses and two novel mAbs are described in detail. mAb LM13 binds to an arabinanase-sensitive pectic epitope and mAb LM14, binds to an epitope occurring on arabinogalactan-proteins. Both mAbs display novel patterns of recognition of cell walls in plant materials.
