Quantification of IdoA present at the at cell surface staining was performed after detachment of the cells using PBS supplemented with 0.5 mM EDTA. Sequential application of antibodies to DS (GD3A12; 1:80), rabbit anti-tag VSV-G (1:400), and donkey anti-rabbit IgG 488 (1:200; Jackson) was performed for 30 min at 4 °C. Human, biotinylated HGF binding experiments were performed after 24 h starvation in 0.1% serum followed by cell detachment in 2 mM EDTA/PBS and analyzed according to the protocol provided by the manufacturer. A FACS-Calibur instrument integrated with Cell-Quest software (BD Biosciences), and Flowjo were used for analysis.
