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A theoretical analysis of the stepwise equilibrium model for macromolecule-ligand interaction is presented. Conditions for the mathematical equivalence of this model and the Scatchard model are examined. The constants for the stepwise equilibrium model can be approximated from the parameters of the Scatchard model. This provides a practical means for fitting binding data to the stepwise equilibrium model.The data initially are analyzed in terms of an appro priate Scatchard model, and the constants for the stepwise equilibrium model are estimated from the results of this analysis. These estimates then are refined by a model-fitting procedure which analyzes the data directly in terms of stepwise equilibrium constants. A detailed application involving the

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