Context: Thyroglobulin (Tg) may be a valuable indicator of improving thyroid function in children after salt iodization. A recently developed Tg assay for use on dried whole blood spots (DBS) makes sampling practical, even in remote areas. Objective: The study aim was to develop a reference standard for DBS-Tg, establish an international reference range for DBS-Tg in iodine-sufficient children, and test the standardized DBS-Tg assay in an intervention trial. Design, Participants, and Interventions: Serum Tg reference material of the European Community Bureau of Reference (CRM-457) was adapted for DBS and its stability tested over 1 yr. DBS-Tg was determined in an international sample of 5- to 14-yr-old children (n = 700) who were euthyroid, anti-Tg antibody negative, and residing in areas of long-term iodine sufficiency. In a 10-month trial in iodine-deficient children, DBS-Tg and other indicators of iodine status were measured before and after introduction of iodized salt. Results: Stability of the CRM-457 Tg reference standard on DBS over 1 yr of storage at -20 and -50 C was acceptable. In the international sample of children, the third and 97th percentiles of DBS-Tg were 4 and 40 μg/liter, respectively. In the intervention, before introduction of iodized salt, median DBS-Tg was 49 μg/liter, and more than two thirds of children had DBS-Tg values greater than 40 μg/liter. After 5 and 10 months of iodized salt use, median DBS-Tg decreased to 13 and 8 μg/liter, respectively, and only 7 and 3% of children, respectively, had values greater than 40 μg/liter. DBS-Tg correlated well at baseline and 5 months with urinary iodine and thyroid volume. Conclusions: The availability of reference material and an international reference range facilitates the use of DBS-Tg for monitoring of iodine nutrition in school-age children. Copyright © 2006 by The Endocrine Society.