We optimized an RIA for measurement of arginine⁸-vasopressin (AVP) in plasma by use of 100-mg Isolute C₁₈ columns for extraction, addition of a preincubation step, and use of maximal dilution of a commercially available antiserum. The detection limit was 0.06 ng/L when 0.5 mL of acidified plasma was extracted. The within- and between-run CVs (n = 16) at physiological concentrations were 5.8–10.2% and 6.5–11.7%, respectively. Prolonged storage of blood at 25 °C, but not at 4 °C, led to a significant increase in measured plasma AVP concentrations. When plasma samples were prepared at several centrifugation speeds, plasma AVP was significantly correlated with the platelet count (r = 0.899; P <0.001). This emphasizes the need for careful sample preparation to avoid contamination of plasma with platelet-bound AVP. Basal plasma AVP in 203 children and adolescents (105 males and 98 females; ages, 1 day to 18 years) averaged 1.1 ± 0.6 ng/L. There was no significant difference between males and females and no correlation with age. In 16 healthy adult controls, plasma AVP averaged 1.0 ± 0.5 ng/L.