Figure 1.

CX3CR1-M280 binds FKN with delayed kinetics. (a) Receptor expression on transfected HEK 293 cell lines. The name of the transfected receptor is at the top of each FACS plot. Thick lines: FITC-conjugated CX3CR1-specific mAb 2A9. Thin lines: isotype antibody controls. Data are representative of three independent experiments. (b) Equilibrium competition binding of ¹²⁵I-FKN to recombinant CX3CR1 variants. Cell lines analyzed in a were incubated with 0.25 nM ¹²⁵I-FKN in the presence of azide and the indicated amount of unlabeled FKN for 2 hours at 37°C. Inset: Scatchard analysis of the binding data. (c) Kinetic binding of ¹²⁵I-FKN to recombinant CX3CR1 variants. Cell lines analyzed in a were incubated with 0.25 nM ¹²⁵I-FKN at 37°C in the presence of azide for the indicated time period. Data in a and b are presented as mean ± SEM and are representative of three independent experiments. (d) Kinetic binding of ¹²⁵I-FKN to recombinant CX3CR1 variants. Cell lines analyzed in c were incubated with ¹²⁵I-FKN for 48 minutes at 37°C, at which time a 1,000-fold molar excess of unlabeled FKN was added. Serial samples of cells were then removed at the indicated timepoints after addition of the unlabeled ligand, and radioactivity was counted. Data are presented as mean ± SEM and are representative of three independent experiments.