Receptor binding properties of CXCL4 and CXCL4L1 on endothelial cells and CXCR3 transfectants. TAMRA-CXCL4L1 [0 ng/mL (no label) or 300 ng/mL (all other lanes)] were added to CXCR3A- or CXCR3B-transfected CHO cells in the absence (Co) or presence of 3000 ng/mL cold CXCL4L1, CXCL4, or CXCL10 (A,C). Cell-bound 8-kDa TAMRA-CXCL4L1 in lysates was analyzed by SDS-PAGE scanned for fluorescence. The dose-dependency of competition for binding to CHO-CXCR3A cells is demonstrated in panel B. Competition for binding of TAMRA-CXCL4L1 to CHO-CXCR3B is shown in panel C. To detect aspecific interaction, binding of 300 ng/mL TAMRA-CXCL4L1 to mock-transfected cells (D) was compared with binding to equal numbers of CXCR3A- and CXCR3B-transfected cells. Results shown are representative of 2 (C), 3 (A-B), or 5 (D) experiments. Confluent monolayers of HMVECs (E-G) were incubated with TAMRA-CXCL4L1 or TAMRA-CXCL10 in the presence or absence of competing unlabeled chemokines (CXCL4L1 or CXCL10). Cell-bound fluorescence present in the fixed cell cultures was quantified by a fluorescence plate reader. Either the concentration of labeled chemokine (E-F) or cold chemokine (G) was kept constant. Results shown are the mean of 2 independent experiments performed in triplicate (E-G).