Congenital hypothyroidism (CH) can be due to an absent gland (athyrosis), an ectopic gland (usually lingual), or an inborn metabolic error (goiter). In this study, radionuclide scanning was compared with ultrasound (US) and serum thyroglobulin (Tg) concentration as a method of determining the cause of CH in 50 newborns. Mean serum Tg values were significantly lower in athyrotic children (7.9 pmol/L) than in either those with goitrous (149.1 pmol/L) or ectopic (60.5 pmol/L) glands, but there was marked overlap among all three groups. With the interpreter "blinded" to the radionuclide diagnosis, a cervical gland could be delineated reliably on US. All 12 goiters and five of five normal glands were identified. In contrast, the correlation between US and radionuclide scanning was poor in patients in whom there was no cervical thyroid tissue. Ultrasound failed to identify 13 lingual glands and was reported as normal in four of 12 children with no radionuclide uptake in the neck and biochemical hypothyroidism. Two children with absent thyroid glands on scanning were biochemically euthyroid. The serum of two others who had normal results of radionuclide studies showed persistent hypothyroidism. Thyroid scanning remains the most accurate diagnostic modality to determine the cause of CH. Serum Tg and cervical US have not been established as reliable alternatives.