A technique used to determine the degree of ion bonding by protein; the protein solution, placed in a bag impermeable to protein but permeable to small ions, is immersed in a solution containing the diffusible ion whose binding is being studied; after equilibration of the ion across the membrane, the concentration of ion in the protein-free solution is determined; the concentration of ion in the protein solution is determined by subtraction; if binding has occurred, the concentration of ion in the protein solution must be greater.