K. S. Morey and Gerald Litwackf abstract: Two proteins which bind the two most negatively charged metabolites of radioactive cortisol formed in 45 min were isolated from rat liver cytosol. Protein-bound radioactivity was separated initially fromunbound radioactivity by column chromatography on Bio-Gel P-100. It then was fractionated into two protein-radioactivity com-plexes uponchromatography on DEAE-Sephadex A-50. Each binding protein was purified to homogeneity. The large binder was purified by chromatography on Cellex-phosphate followed bySephadex G-75. The small binder was purified by chromatography and rechromatography on Sephadex G-75. The large binder had an estimated molecular weight of 37,000 calculated from amino acid analysis and from sedi-mentation velocity and diffusion data, although a value of 50,000±6,000 was obtained by chromatography on Sephadex G-75. The sedimentation coefficient was 3.47 S, a value