Equilibrium gel filtration liquid chromatography is a technique used for the quantitation of ligand binding. We have designed an experiment for the senior undergraduate laboratory using this technique for the measurement of the binding of ATP to cytochrome c. This experiment introduces the student to the use of liquid chromatography and absorbance spectrometry and permits a quantitative result within a three-hour period. The experiment complements classroom discussions regarding liquid chromatography, spectroscopy, dynamic equilibrium, and kinetic parameters.