Figure 1.

Panel A: Screenshot of the SEDPHAT ITC visualization tool of the differential heats dQ/dc_A,tot for the titration of a component A into another component B (K_D = 0.1 µM, ΔH = −10 kcal/mol), which are shown in the form of a two-dimensional heat map (large negative enthalpy changes in gray/purple/magenta/blue, and small heat changes in red to black). The screenshot was modified to enhance the axis labels. In the plot, the plane of total concentrations (ranging logarithmically from 0.01 to 100 µM in each dimension) is cropped to the region that reflect experimentally observable concentrations if the maximum stock concentration of each component is 100 µM, provided a cell volume of 1400 µl and a total syringe volume of 300 µl. A left-to-right swipe with the mouse allows to delineate an experimental titration trajectory, indicated by the yellow circles connected by the black line, here reflecting 30 injections at 10 µl each of 45.6 µM A into the cell with initially 6.2 µM B – information that is provided after any trajectory is graphically entered. Additionally, the small inset shows the shape of heat changes of a putative experiment along this titration. The bank of buttons on the right allows depicting any fractional population of free or complex species in any component (light blue), as well as the fractional contribution of each species to the total heat changes (green). The bank of buttons on the left allows to switch the titration type, crop the display, pick another experimental trajectory, change binding parameters, switch to a differencing mode, and exit this display to simulate data for the trajectory shown. Panel B: Analogous isotherm for the same system for the injection of B into A, with an experimental trajectory of titrating 56.9 µM B into 5.7 µM A. Panel C: ITC data generated along the trajectories shown in Panel A and B.