We coat beads with proteins of interest and measure association of a receptor and ligand by flow cytometry. For instance we have produced beads coated with pairs of LFA-1 molecules (a b 2 integrin expressed by neutrophils). Because LFA-1 is thought to be expressed in pairs (i.e. dimerized) on the surface of a neutrophil, this configuration is relevant to neutrophil recruitment. We measure association of fluorescently labeled ICAM-1 (the primary ligand of LFA-1) with the LFA-1 coated beads by flow cytometry. The more fluorescent ICAM-1 binds to the beads, the brighter each bead will appear during flow cytometric analysis. We found that “dimeric” LFA-1 binds 10 times more strongly to paired ICAM-1 than monomers of ICAM-1, indicating that the density of ICAM-1 on the endothelial surface an important determinant of neutrophil adhesion. We continue to use this technique to characterize the binding rate of novel inhibitors of LFA-1 function; a precursor to the development of anti-inflammatory drugs.

The above image is a schematic of the surface chemistry of an integrin coated bead.