Fig. 3.

Binding of EFNB2 to HeV and NiV G. Two ELISA formats were conducted as detailed in Experimental Procedures. (A) EFNB2/Fc was used as the capture antigen for HeV and NiV sG, detected with anti-sG antisera. (B) HeV and NiV sG were used as capture antigens and detected with either anti-HeV or anti-NiV antisera or EFNB2/Fc. Anti-Tioman virus (TIV) was used as negative control. (C) HeV and NiV coprecipitations. Cell lysates containing myc-tagged metabolically labeled G were precipitated with 9E10 mAb (lanes 1-3) or EFNB2/Fc (lanes 4-6) and protein G (15). Precipitates were separated by SDS/PAGE, and visualized by autoradiography. WR, control vaccimia virus. (D) Interaction between EFNB2/Fc and sG by surface plasmon resonance using a BIACORE 1000. Two surfaces where sG was associated at two different concentrations in two independent experiments were used for test of reproducibility. In all of the experiments EFNB2 was stripped with 10 mM glycine·HCl (pH 2.0), and regeneration did not affect its binding at the same concentration in the next cycle. The arrow indicates the beginning of the dissociation phase.