Interpretation of Double Reciprocal Plots Used to Determine the Spectroscopic Parameters of Bound Ligand for Binding Assays

The spectroscopic determination of binding data usually requires measurement of the spectral characteristics of the chromophore when all sites on the acceptor are occupied by the ligand. The double reciprocal plot employed for this purpose has been interpreted in diverse ways. This paper examines the theoretical and practical limitations which must be considered in making such interpretations, and shows that the linear extrapolation of the double reciprocal plot which is normally practised cannot be justified.