General Conditions for Interpretable Results
In principle, any technique that separates bound from unbound ligand, or occupied from unoccupied protein, can be used to monitor binding. As a general rule, highest accuracy is achieved when the concentration of the protein in binding studies is approximately equal to K d (within about one order of magnitude either way), this makes it easier to determine the concentration difference between bound and total ligand. Since the supply of protein is usually a limiting factor in research, binding studies are expensive and the sample size must be reduced as much as possible. Considerable progress has been made in this direction over the years.