Figure 4.

Electrophysiological approach to measure the kinetics of 8B6 scFv- and antiSERT-EL4 antibody-binding to SERT.A and B, representative peak current traces recorded with variable intervals of preincubation (1, 2, 5, 10, and 15 s in A and 0.2, 0.5, 1, 2, 5, and 10 s in B) for 100 nM 8B6 scFv (A) and 20 nM antiSERT-EL4 antibody (B). C and D, normalized peak currents for the indicated concentrations of 8B6 scFv (C) and of antiSERT-EL4 antibody (D) were plotted as a function of time. The data points were fitted to the equation for a monoexponential decay to a basal level to estimate the apparent association rates (k_app) for each concentration. These k_app values were plotted over the corresponding (E) 8B6 scFv and (F) antiSERT-EL4 antibody concentrations to yield a straight line. The data are mean ± SD from four to eight independent recordings. Association (k_on) and dissociation (k_off) rate constants were calculated from the slope and the y-intercept, respectively: k_on-values for 8B6 scFv and antiSERT-EL4 were 1.93 ± 0.36 nM × 10⁶ M⁻¹ s⁻¹ and 5.05 ± 0.61 nM × 10⁷ M⁻¹ s⁻¹ and k_off-values were 0.20 ± 0.07 s⁻¹ and 1.20 ± 0.07 s⁻¹, respectively.