A cadmium-saturation technique for quantifying metallothionein (MT) in mammalian tissues was evaluated for use in fish tissue. We administered 3 mg109cadmium/kg body weight by intraperitoneal injection over a 5-d period to adult brook trout Salvelinus fontinalis to induce MT in liver and kidney tissues. The cadmium-saturation technique was modified so the amount of cadmium bound to unsaturated and cadmium-saturated MT could be measured. The method gave precise measurements of MT concentrations when aliquots of liver supernatant, which were analyzed separately, were quantified by atomic absorption or radiometric measurements. Two to four times more cadmium and MT concentrated in the liver of treated fish than in the kidney. Intraperitoneal administration of cadmium completely displaced copper and zinc from MT in liver of treated fish; cadmium concentrations in liver determined by the quantitation of cadmium-saturated MT and of unsaturated MT were identical. However, exposure of brook trout to cadmium in water did not result in the complete saturation of MT with cadmium. Concentrations of MT and mortality were significantly increased in fish exposed to 5 μg Cd/L or more for 30 d.