The influence of topical iodine-containing antiseptics on thyroid function test results of premature infants was determined in two separate studies. Thyroxine and thyrotropin levels were measured on blood-spotted filter paper. Samples were obtained from 128 premature infants on their tenth day of life; the infants were treated in two neonatal intensive care units. Both units used similar treatment protocols; however, one routinely used topical iodinated antiseptic agents (n = 73), whereas the other used chlorhexidine-containing antiseptics (n = 55). There was no difference in the mean T4 levels between the two groups. The mean thyrotropin levels were elevated in preterm babies exposed to iodine (15.4 vs 7.8 mIU/L, p < 0.01). Among the iodine-exposed infants, elevated thyrotropin levels (> 30 mIU/L) were found in 13.7% of infants, compared with none in the chlorhexidine-treated group (p < 0.01). We then studied an additional 46 premature infants who were treated in one neonatal intensive care unit. Iodine-containing solutions were used in 24 infants and chlorhexidine was used in 22 infants. T4 and thyrotropin levels were measured weekly during the first 28 days, one every 2 weeks until the age of 60 days, and at the age of 90 days. Among iodine-exposed infants, 20.8% had thyrotropin values > 30 mIU/L, whereas none of the infants in the chlorhexidine group had elevated thyrotropin values (p < 0.05). The elevated thyrotropin levels correlated positively with the area of disinfection. Elevated urine iodine levels were present reflecting an abnormally high iodine absorption. This study suggests that iodine absorption from topical iodine-containing antiseptics may cause disturbances in thyroid function test results in premature infants. We recommend that caution be exercised in the use of iodine-containing antiseptics in premature infants.