Glycocalyx Scaffolding to Control Cell Surface Glycan Displays.

Abstract

This article describes a protocol for remodeling cells with synthetic glycoprotein and glycolipid mimetics that are functionalized with lipid anchors, allowing for cell surface display of specific glycan structures in predefined nanoscale arrangements. The complex chemical heterogeneity of glycans found on the cell surface or the glycocalyx renders analysis of the individual contributions of glycans difficult. This technique allows for the precise study of individual glycans at different regions of the glycocalyx, and may be useful for interrogating glycan interactions in infection or immunity or in stem cell differentiation. CHO-Lec2 cells are prepared as adherent monolayers and, after reaching confluence, are incubated with the glycomaterials. Synthetic glycopolymers bearing α-2,3-sialyllactose glycans are used to decorate cellular surfaces in the form of 3D multivalent ligands projecting away from the cell surface, while α-2,6-sialyllactose glycolipid conjugates are used to anchor glycans in dynamic 2D arrays proximal to the cell membrane. Following washing, mimetic incorporation and glycan display can be analyzed using lectins with specificity for α-2,3- or α-2,6-linked sialic acids. Flow cytometry data reveals that cell surface remodeling with either glycoconjugate mimetic occurs efficiently in a dose-dependent manner. Combinations of glycoconjugates can also be employed simultaneously to generate a mixed glycocalyx with tunable composition and organization. © 2018 by John Wiley & Sons, Inc.