Zinc, as an essential trace element for health, plays various biological roles in human body functions. Serum zinc reference values are essential for assessing zinc-associated abnormalities and the prevalence of zinc deficiency. This study aims at determining age- and sex-specific reference values for serum zinc concentrations in adult Iranian subjects. Serum zinc concentration was measured by flame atomic absorption spectrometry in 4,698 adult subjects, aged 20-94 years, randomly selected from the population of the Tehran, Lipid, and Glucose Study. After application of exclusion criteria, reference values for serum zinc were determined in 2,632 apparently healthy subjects according to guidelines of the International Federation of Clinical Chemistry (nonparametric method). Dietary zinc was assessed in 2,906 individuals, of which 1,685 were healthy subjects, using a validated semiquantitative food frequency questionnaire. Reference values for serum zinc concentrations ranged between 9.6 and 31.6, 8.9 and 29.9, and 9.3 and 30.8 μmol/L in men, women, and the total population, respectively. Prevalence of serum zinc deficiency was 3.0 and 2.4 % in men and women, respectively (p = 0.267); in men, but not in women, the prevalence increased significantly with age (p for trend <0.001). Of the total participants, 10.3 % (6.5 men and 3.8 % women, p < 0.01) had lower zinc intake compared to dietary reference intakes. The zinc density of the population was 6.3 mg/1,000 kcal. In conclusion, this study presents reference values for serum zinc concentration in adult Iranian subjects for both sexes and different age groups. Prevalence of serum zinc deficiency and dietary zinc inadequacy seems to be lower in Iranians, compared to some other populations.