Histochemistry of Single Molecules

Ellie-May Beaman, David R. F. Carter, Susan A. Brooks

Carlo Pellicciari, Marco Biggiogera

The glycans displayed on the cell surface are highly heterogeneous and their function in cell recognition, identity, signaling, adhesion, and behavior is increasingly recognized. Moreover, as it is yet incompletely understood, it is a topic of significant current interest. Lectins (naturally occurring carbohydrate-binding proteins) are very useful tools for exploring cellular glycosylation. Cell populations, within or between different tissues or species, and in development, health and disease, exhibit different glycosylation and thus distinct lectin-binding characteristics. Even monoclonal cell populations of established cell lines feature subpopulations with strikingly different glycosylation characteristics, and these differences may reflect differences in behavior or function. By separating cell populations on the basis of their cell surface glycosylation, the functional significance of glycosylation can be investigated in in vitro or in vivo models. Also, factors affecting glycosylation, which are also incompletely understood, can be explored or manipulated. In the protocol given here, cells can be separated into subpopulations on the basis of their recognition by a specific biotinylated lectin of choice immobilized on avidin-coated magnetic beads. Importantly, the protocol has been optimized such that lectin-binding and non-binding cells remain viable such that they can be further cultured, if necessary, for subsequent investigations.