Examination of Whole Cell Galectin Binding by Solid Phase and Flow Cytometric Analysis

Abstract

We have utilized simple flow cytometric and fluorescence-based solid phase assays to study the interaction of glycan-binding proteins (GBP) to cell surface glycoconjugates. These methods utilize commonly employed flow cytometry techniques and commercially available streptavidin-coated microplates to immobilize various biotinylated ligands, such as glycopeptides, oligosaccharides, and whole cells. Using this approach, fluorescently labeled GBPs, in particular, members of the galectin family, can be interrogated for potential interactions with cell surface carbohydrates, including elucidation of the potential impact of alterations in glycosylation on carbohydrate recognition. Using these approaches, we present examples of flow cytometric and fluorescence-based solid phase assays to study galectin–carbohydrate interactions.

Galectin-9 recognizes and exhibits antimicrobial activity toward microbes expressing blood group–like antigens

“…Bacteria were washed twice more and resuspended in 300 µL of 1x PBS for analysis by flow cytometry using a FACSCalibur flow cytometer (BD Biosciences). Results were analyzed using CellQuest software (BD Biosciences) (65).…”