Figure 6.

Competition binding assay between angiostatin and the antibody against the α-subunit of ATP synthase from E. coli. HUVEC were plated at a constant density of 10,000 cells/well and incubated with 0.5 μM ¹²⁵I-labeled angiostatin in the presence of 1:10 dilution of antibody against the α-subunit of ATP synthase from E. coli for 1 h at 4°C. Cells were washed and remaining bound radioactivity was quantified by γ-counting. Nonspecific binding was measured in the presence of excess unlabeled angiostatin and was subtracted from total binding. (A) Total binding of 0.5 μM ¹²⁵I-labeled angiostatin was designated as 100%. (B) Angiostatin binding is inhibited by 59% in the presence of a 1:10 dilution of anti-α-subunit ATP synthase antibody. Competition studies also were performed simultaneously by using rabbit preimmune serum to account for nonspecific inhibition. Error bars represent SD. A one-tailed homoscedastic t test was used for statistical analysis; P < 0.10.