Affinity of monoclonal antibodies for Globo-series glycans

• Common monoclonal antibodies bind SSEA-4 and Globo H with nanomolar affinity.
• The antibodies exhibit negligible cross-reactivity.
• Binding is tighter to surface-bound than to solution-phase glycans.
• High specificity validates common monoclonal antibodies for biomedical applications.

Globo-series glycans are human cell-surface carbohydrates that include stem-cell marker SSEA-4 and cancer-cell antigen Globo H. These two hexasaccharides differ only in their terminal saccharide: N-acetylneuraminic acid in SSEA-4 and l-fucose in Globo H. Herein, we evaluated the affinity of the monoclonal antibodies α-SSEA-4 and α-GH for the glycans SSEA-4 and Globo H. Using fluorescence polarization, we find that the two monoclonal antibodies have affinity for their cognate glycan in the low nanomolar range, and have negligible affinity for the non-cognate glycan. Using surface plasmon resonance, we find that each cognate affinity is ∼20-fold greater if the glycan is immobilized on a surface rather than free in solution. We conclude that the terminal saccharide plays a dominant role in the ability of monoclonal antibodies to recognize these Globo-series glycans and that the extraordinary specificity of these antibodies supports their use for identifying and sorting stem-cells (α-SSEA-4) and as an agent in cancer immunotherapy (α-GH).

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