Hi All,

I am performing a metal-ligand binding experiment and want to determine the binding kinetics of the ligand, which is a protein. I understand I have to generate a saturation binding curve, which is a plot of the bound metal against the total metal ion concentration; then, the Kd and Bmax will be determined from the plot. I have done the experiment; however, couldn't get a saturation, as the plot was persistently linear. I explored many options such as decreasing the protein concentration by many folds, which would be expected to decrease the metal-binding sites, and hence can facilitate saturation as the metal concentration increases. However, the results remain the same.

Please I would really appreciate your suggestions on how to attain a saturation, or an alternative approach to determining the binding kinetics, if any.