Zinc metabolism was studied in Zn-deficient and control Holstein calves over a 2-mo period following a single oral or i.v. 65Zn dose. In both orally and i.v. dosed animals, all gastrointestinal tissue sections from Zn-deficient animals contained more 65Zn than comparable tissues of controls. Contents of proximal small intestinal sections of Zn-deficient calves contained more 65Zn 8 to 10 wk after dosing than did those from controls; however, the reverse occurred in the distal small intestine, cecum, and large intestine. With both dosing methods, Zn-deficient calves retained more 65Zn throughout the study. Daily 65Zn excretion rate as a percentage of that retained declined for 6 wk after dosing, indicating a constantly increasing biological half-life. For deficient calves, the biological half-life was about 500 d in the later weeks of the experiments. In orally dosed, Zn-deficient animals, specific activity of fecal 65Zn exceeded that of serum Zn throughout the study. This shows a shortcoming in the basic assumption of measuring endogenous Zn loss from fecal and serum specific activities and total fecal stable Zn. Thus, endogenously excreted Zn is not representative of that remaining.