FIGURE 4.
Binding of HRG to γ′-peptide. A, the binding of 1.1 μm HRG to fluorescein-labeled γ′-peptide (50 nm) was monitored in the presence of ZnCl2 (0–20 μm) at λex = 492 nm and λem = 532 nm. Initial fluorescence was determined in the presence of HRG but in the absence of ZnCl2 (Io). Aliquots of ZnCl2 were then added, and the fluorescence intensity (I) was measured after each addition. I/Io values are plotted versus ZnCl2 concentrations. B, binding of HRG to immobilized γ′-peptide was measured by SPR. Biotinylated γ′-peptide was adsorbed to a streptavidin-immobilized CM4 chip. Binding of HRG (0–700 nm) to γ′-peptide in the presence of 20 μm ZnCl2 was then determined in the absence (open symbols) or presence (closed symbols) of the γ′-peptide-directed IgG. Corrected Req values are plotted against the input HRG concentrations. Symbols represent the mean ± S.D. of two experiments, and lines represent nonlinear regression analyses of the data.