Urinary excretion of retinol-binding protein (RBP) in rabbit chronically poisoned with cadmium. Isolation and partial characterization of rabbit RBP.

Studies were conducted to induce urinary excretion of retinol-binding protein (RBP) in the rabbit by long-term administration of cadmium, and to isolate and characterize rabbit RBP. Two rabbits were exposed to Cd at a dose of 0.8 and/or 1.5mg/kg of body weight by means of subcutaneous injections five times per week. One rabbit excreted large amounts of protein (0.8 to 1.6 g/day) from days 80 to 118. The electrophoretic patterns of the urinary protein showed two fluorescent bands specific for protein-bound retinol in the alpha-region; these were presumed to be RBP. Rabbit RBP was isolated from the pooled urine (10.3 liters) by a sequence of procedures which included gel filtration on Sephadex G-100 and chromatography on DEAE- and SP-Sephadex. Purified RBP (20 mg) was found to be homogeneous by physical and immunological criteria. The RBP had alpha-mobility, with a molecular weight of approximately 20,000. The properties of rabbit RBP resembled those of human RBP simultaneously isolated from the urine of patients with "Itai-Itai" disease in many ways: ultraviolet and fluorescence spectra, and amino acid compositions. A monospecific anti-rabbit RBP antiserum was raised in a goat. There was no immunological cross-reactivity between rabbit and human RBP. The molecular size of the retinol-containing protein in fresh rabbit serum was estimated to be about 60,000 to 70,000 by gel filtration on Sephadex G-200. Rabbit RBP in the serum was also shown to be immunologically identical with purified RBP from the urine.