Calibration of Size-Exclusion Chromatography: Use of a Double Gaussian Distribution Function to Describe Pore Sizes

Abstract A method for the calibration of size-exclusion chromatographic columns is proposed that takes into account the nonlinear dependence of the Stokes radius R s upon the partition coefficient K D . The method is based on the assumption that the pore size distribution of a molecular sieve column can be described by the sum of two Gaussian distributions and has been successfully tested with low-pressure chromatography gels (Sephacryl and Superose) and HPLC gels (TSK SW) over a wide range of protein sizes. An application of this method is described, in which aggregation states of the membrane protein prostaglandin H 2 synthase solubilized in nonionic detergents are estimated.