Improved curve fitting procedures to determine equilibrium binding constants

Frank H. Stootman*, Dianne M. Fisher, Alison Rodger, Janice R. Aldrich-Wright

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

109 Citations (Scopus)


For ligand-biomacromolecule titration experiments it has been traditional practice to extract parameters such as the equilibrium binding constant K and the number of bases per ligand binding site n with relatively labour intensive methods, usually based on single wavelength data, such as the difference method by Rodger and Nordén coupled together with a Scatchard plot. Presented in this paper are both the theory and a least squares fitting method to derive parameters such as K and n more directly from all spectral non-linear experimental data. Both the case of non competitive binding of a metal complex ligand to DNA and the case of displacement by a metal complex ligand of an ethidium marker attached to the DNA are considered. This work may be applied directly to reduce experimental data produced by a spectropolarimeter (for circular or linear dichroism) or a spectrophotometer (for fluorescence or UV-Vis spectroscopy).

Original languageEnglish
Pages (from-to)1145-1151
Number of pages7
Issue number10
Publication statusPublished - 2006
Externally publishedYes


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