Figure 2

Saturation binding characteristics of [¹²⁵I-Tyr]CGRP(8–37). Increasing amounts of [¹²⁵I-Tyr]CGRP(8–37) were used on a crude HEK293T-RAMP1 membrane preparation that had been transiently transfected with wild type CLR, in the absence and presence of 1μM CGRP to determine total and nonspecific binding, respectively. Non-linear regression analysis was used to best fit a specific binding curve to each individual binding experiment. From this best fit curve an equilibrium dissociation constant (K_d) for the peptide radioligand and total number of specific binding sites (B_max) were estimated. The K_d of [¹²⁵I-Tyr]CGRP(8–37) was calculated to be 0.9 ± 0.2 nM with a B_max estimated at 285 ± 206 fmols/mg protein. The data presented represents the mean ± S.E. for n = 3 experiments performed in duplicate.