A graphical method for determination of enzyme-inhibitor dissociation constants for non-competitive inhibitors, especially those that bind very tightly. Where I is the total inhibitor concentration, E is the total enzyme concentration and α is the ratio of inhibited to non-inhibited rates under the same conditions, Iα is plotted on the ordinate (y-axis) against 1/(Iα) on the abscissa (x-axis) to give a straight line that intersects the ordinate at E; its slope is K_d, the dissociation constant.