Techniques for analysis of glucose binding by human erythrocyte membranes

Two methods of measuring the binding of glucose by a component of the human erythrocyte membrane are described and compared, namely: (1) retardation of d-[³H]glucose with respect to l-[¹⁴C]glucose on columns of 'Celite'-ghost mixtures and (2) a new technique of ultrafiltration of a solution of d-[³H]glucose, l-[¹⁴C]-glucose and cell membrane extract. The binding is shown to be closely associated with the glucose transport system by the following criteria for the identification of a facilitated diffusion system: (a) The binding is specific for d-glucose in preference to the l form. (b) Phloretin, a competitive inhibitor of the monosaccharide transport system as studied by kinetic experiments, is shown to inhibit the binding. (c) Saturation of binding is indicated at high levels of glucose.