An analysis of the limits of resolution of binding experiments as assays for affinity heterogeneity

An analysis is presented of the limits of resolution of Scatchard and Sips plots as methods for detecting affinity heterogeneity in receptor populations. We estimate using statistical criteria for defining resolution that Scatchard plots can resolve two groups present at equal concentration, provided they differ by more than a factor of about six in affinity, and that resolution drops rapidly as the concentration difference between the groups increases. In addition, because of the non linear relation between affinity and free energy, and because the Sips analysis must necessarily be formulated in terms of free energies, the Scatchard plot is the more sensitive of the two methods for detecting heterogeneity.