FIG. 4.
Binding of CON6 gp120 and gp140CF to sCD4 and anti-Env MAbs. (A and B) Each of the indicated MAbs and sCD4 were covalently immobilized to a CM5 sensor chip (BIAcore), and CON6 gp120 (A) or gp140CF (B) was injected over each surface (100 and 300 μg/ml, respectively). (C and D) To determine induction of 17b MAb binding to CON6 gp120 and gp140CF, CON6 gp120 (C) or gp140CF (D) proteins were captured (400 to 580 response units) on individual flow cells immobilized with sCD4 or MAb A32 or T8. Following stabilization of each of the surfaces, MAb 17b was injected and allowed to flow over each of the immobilized flow cells. (E) To determine binding of CON6 gp120 and gp140CF to human MAbs in ELISA, titers of stock solutions of 20 μg of MAbs 447-52D, F39F, A32, IgG1b12, and 2F5 were determined with CON6 gp120 and gp140CF glycoproteins. MAbs 447-52D (V3), F39F (V3) A32 (gp120), and IgG1b12 (CD4 binding site) each bound to both CON6 gp120 and gp140CF well, while 2F5 (anti-gp41 ELDKWAS) bound only CON6 gp140CF. The concentrations at the end point titer (end titer where the experimental versus the control value was ≥3.0) with gp120 for MAb 447-52D and F39F binding were <0.003 and 0.006 μg/ml, respectively; that for MAb A32 was <0.125 μg/ml; that for IgG1b12 was <0.002 μg/ml; and that for 2F5 with gp140CF was 0.016 μg/ml.