Interactions of Form X with variable concentrations of HMG1. Form X, end-labelled to the highest possible specific activity, was incubated with serial dilutions of protein HMG1. (A) The interactions were performed at three different concentrations of Form X (16 pM, 1.6 pM, and 0.16 pM). For each concentration the quantification of the radioactivity in the bands was performed with a phosphorimager, allowing us to determine the HMG1 concentration necessary to bind 50% of the Form X. (B) The results were plotted on a double logarithmic scale, showing that the protein concentration at half saturation (expressed as a dilution factor of a protein stock at ~ 20 μg/mL i.e. ~ 8 × 10-7 M) decreases with the concentration of Form X in the samples, and therefore that the KD of the interaction is lower than 0.16 pM.