Polyacrylamide glycoconjugates, Glyc-PAA, having various tags or labels are convenient tools for analysis of cellular
lectins. Adaptation of such glycoprobes for flow cytometry allows us to reveal
lectins expressed on cell surface and analyze their
carbohydrate specificity as well as functionality. Localization of
lectins is visualized by labeling of cells with
fluorescein-tagged glycoprobes, Glyc-PAA-fluo, in combination with fluorescent microscopy techniques. Additionally, biotinylated glycoprobes can be immobilized on magnetic particles making it possible to separate a cell population according to its
carbohydrate-binding profile. Here, we exemplify application of glycoprobes in the study of cellular
siglecs and
galectins, as well as
lectin patterning of
tumor cells. The specificity of
sialic acid binding membrane-anchored
lectins, siglecs-1, -5, -7, -8 and -9 was determined using this methodology. To study the
carbohydrate-binding profile of soluble galactoside-binding
lectins, galectins-1 or -3, these were loaded on (initially
galectin free) Raji cells and probed using Glyc-PAA-fluo. Lessons learned from this model system allowed us to study the
galectin distribution pattern of
tumors: cells obtained from mice carrying mammary
adenocarcinoma or
lymphoma were probed with Glyc-PAA-fluo using flow cytometry.
Disaccharide 6OSuLacdiNAc was shown to be the most potent probe for
adenocarcinoma cells, demonstrating that 6OSuLacdiNAc-binding molecules accumulate on cell surface in a patch-wise distribution.