Figure 2.

Competition binding assay between plasminogen and angiostatin. HUVEC were plated at a density of 10,000 cells/well and incubated with 1.0 μM ¹²⁵I-labeled plasminogen in the presence of 100-fold molar excess of unlabeled angiostatin for 1 h at 4°C. Cells were washed, and the remaining radioactivity was quantified by γ-counting. (A) Total binding of 1.0 μM ¹²⁵I-labeled plasminogen was designated as 100%. (B) Plasminogen binding is inhibited by ≈80% in the presence of a 25-fold molar excess of unlabeled plasminogen. (C) Plasminogen binding was not inhibited in the presence of a 100-fold molar excess of unlabeled angiostatin, suggesting distinct binding sites for each on the cells. Similar experiments by using ¹²⁵I-labeled angiostatin (D) showed no inhibition of binding in the presence of a 2-fold molar excess unlabeled plasminogen (E). Error bars represent SD.