Kinetic parameters of [2-14C]malonate uptake by the human erythrocyte membrane have been determined as Km, 24 mM and turnover number, 5 × 104 s−1. The translocation of this organic dianion is concentration, pH and temperature dependent. Competitive inhibition of malonate uptake by eosin and inorganic anions, strongly implies that a common route exists for both inorganic anions and organic dianions, namely the anion-exchange Band 3 protein. 14C-Malonate which is nonmetabolized in the erythrocyte, could be a useful probe for monitoring anion-exchange in reconstituted Band 3 systems.