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Maturation of the rat fetal thyroid was studied with the aid of 1131 and of fluorescence and electron microscopy. The I ~31 concentration of the fetal gland increased exponentially from day 17 to day 20 of gestation and was related to the weight of the fetus (and presumably the weight of the thyroid) and also to the quantity of 113t accumulated by the fetus. In the 17-day gland, thyroglobulin or immunologically similar material was sparsely present in the incipient lumens of some cell clusters. With maturation, this material increased and was also observed within follicular cells on days 18 to 19 of gestation. On day 20, the specifically reacting material was present in the follicular lumens and was absent from the cytoplasm of follicular epithelium. Ultrastructurally, the earliest thyroid cells examined were replete with all the organelles found in the more mature epithelium. No direct correlation could be made between the cytoplasmic structures and the presence of thyroglobulin, although the granular endoplasmic reticulum was most likely the organelle responsible for synthesis of thyroglobulin. Thyroglobulin or a precursor was found in fetal thyroid cells before measurable quantities of 113~ were concentrated and before cytoplasmic droplets appeared.Between day 18 and day 19 of gestation, the fetal thyroid of the rat acquires the capacity to concentrate easily measurable quantities of I t3t (1). Based on this property of isotope accumulation by the fetal thyroid, and with the aid of electron and fluorescence microscopy, the following study was designed to seek answers to three questions: First, does an intracellular organelle appear or become singularly prominent at the time the thyroid gland begins to trap iodide in high concentration? Second, during the maturation of the fetal thyroid, is it possible to localize the site of synthesis of specific thyroid protein, thyroglobulin? Third, is thyroglobulin or a precursor produced before the advent of iodide concentration by the gland? MATERIALS AND METHODSPregnant rats (Holtzman) with known mating times were injected subcutaneously on the 16th to 19th day of gestation a with 50 to 60 #c of 1131. 24 hours later the fetuses were removed from their etherized dams and their thyroids were visualized under a stereomicroscope. The glands to be processed for electron microscopy were flooded with 2 per cent osmium tetroxide in situ as quickly as possible after removal of the fetus from the uterus, usually within less than 1 minute. After freeing from the trachea, they were placed in 1 per cent buffered osmium tetroxide containing sucrose for 1 to 2 hours and then processed for Vestopal W embedding. Other fetal glands were embedded in a small piece of maternal 1 24 hours after a vaginal smear positive for sperm was counted as 1 day of gestation.