Glycobiology Advance Access originally published online on December 21, 2005
Glycobiology 2006 16(4):349-357; doi:10.1093/glycob/cwj071
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Immunoglobulin G specifically binding plant N-glycans with high affinity could be generated in rabbits but not in mice
2 Department of Chemistry and 3 Department of Biotechnology, University of Natural Resources and Applied Life Sciences (BOKU), 1190 Vienna, Austria; 4 Institute of Pathophysiology and 5 Clinical Institute of Pathology, Medical University Vienna, 1090 Vienna, Austria
1 To whom correspondence should be addressed; e-mail: friedrich.altmann{at}boku.ac.at
Received on November 8, 2005; revised on December 13, 2005; accepted on December 16, 2005
Xylosylated and core 
1,3-fucosylated N-glycans from plants are immunogenic, and they play a still obscure role in allergy and in the field of plant-made protein pharmaceuticals. We immunized mice to generate monoclonal antibodies (mAbs) binding plant N-glycans specifically via the epitope containing either the xylose or the core 1,3-fucose residue. Splenocytes expressing N-glycan-specific antibodies derived from C57BL/6 mice previously immunized with plant glycoproteins were preselected by cell sorting to generate hybridoma lines producing specific antibodies. However, we obtained only mAbs unable to distinguish fucosylated from xylosylated N-glycans and reactive even with the pentasaccharide core Man3GlcNAc2. In contrast, immunization of rabbits yielded polyclonal sera selectively reactive with either fucosylated or xylosylated N-glycans. Purification of these sera using glyco-modified neoglycoproteins coupled to a chromatography matrix provided polyclonal sera suitable for affinity determination. Surface plasmon resonance measurements using sensor chips with immobilized glyco-modified transferrins revealed dissociation constants of around 10–9 M. This unexpectedly high affinity of IgG antibodies toward carbohydrate epitopes has repercussions on our conception of the binding strength and significance of antiglycan IgE antibodies in allergy.
Key words: binding affinity / carbohydrate epitope / glycoprotein / immunogenic N-glycan
* These authors contributed equally to this work.
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