This chapter has been superceeded by 4 newer chapters, 6a, 6b, 6c, and 6d. However this Chapter, written originally by Dr Samuel Refetoff and updated by Drs Franklyn and Shephard, remains a treasure trove of information on many now-obscure thyroid tests, and references. For that reason we maintain it as a part of our Archive for use of MDs who may wish to investigate a bit of the history of thyroid testing. L De Groot, MD
The possibility of thyroid disease is considered when signs or symptoms suggest hyper- or hypothyroidism or some physical abnormality of the thyroid gland. Evaluation of the patient should include a thorough history and physical examination. Since most thyroid diseases require prolonged periods of treatment, it is crucial that a firm diagnosis be established before embarking on such a program. Further, a number of medications, in particular those used in the treatment of thyroid disease, may alter the results of thyroid function tests in such a way that reinvestigation after therapy has begun may provide ambiguous results.
During the past three decades, clinical thyroidology has witnessed the introduction of a plethora of diagnostic procedures. These laboratory procedures provide greater choice, sensitivity, and specificity which have enhanced the likelihood of early detection of occult thyroid diseases presenting with only minimal clinical findings or obscured by coincidental nonthyroid diseases. They also assist in the exclusion of thyroid dysfunction when symptoms and signs closely mimic a thyroid ailment. On the other hand, the wide choice of complementary and overlapping tests indicates that each procedure has its limitations and that no single test is always reliable.
Thyroid tests can be classified into broad categories according to the information they provide at the functional, etiologic, or anatomic levels ( Table 6-1 ).
1. Tests that directly assess the level of the gland activity and integrity of hormone biosynthesis. These tests such as thyroidal radioiodide uptake and perchlorate discharge are carried out in vivo.
2. Tests that measure the concentration of thyroid hormones and their transport in blood. They are performed in vitro and provide indirect assessment of the level of the thyroid hormone dependent metabolic activity.
3. Another category of tests attempts to more directly measure the impact of thyroid hormone on peripheral tissues. Unfortunately, tests available to assess this important parameter are nonspecific, since they are often altered by a variety of nonthyroidal processes.
4. The presence of several substances, such as thyroid autoantibodies, usually absent in healthy individuals, are useful in establishing the etiology of some thyroid illnesses.
5. Invasive procedures, such as biopsy, for histological examination or enzymatic studies are occasionally required to establish a definite diagnosis. Gross abnormalities of the thyroid gland, detected by palpation, can be assessed by scintiscanning and by ultrasonography.
6.The integrity of the hypothalamo-pituitary-thyroid axis can be evaluated by (a) the response of the pituitary gland to thyroid hormone excess or deficiency; (b) the ability of the thyroid gland to respond to thyrotropin (TSH); and (c) the pituitary responsiveness to thyrotropin-releasing hormone (TRH). These tests are intended to identify the primary organ affected by the disease process that manifests as thyroid dysfunction; in other words, primary (thyroid), secondary (pituitary), or tertiary (hypothalamic) malfunction.
7.Lastly, a number of special tests will be briefly described. Some are valuable in the elucidation of the rare inborn errors of hormone biosynthesis, and others are mainly research tools.
Each test has inherent limitations, and no single procedure is diagnostically adequate for the entire spectrum of possible thyroid abnormalities. The choice, execution, application and interpretation of each test requires the understanding of thyroid physiology and biochemistry dealt with in the preceding chapters. Thyroid tests serve not only in the diagnosis and management of thyroid illnesses but also to better understand the pathophysiology underlying a specific disease.
Table 6-1. Tests of Thyroid Function and Aids in the Diagnosis of Thyroid Diseases
In Vivo Tests of Thyroid Gland Activity and Integrity of Hormone Synthesis and Secretion Thyroidal Radioiodide Uptake (RAIU) Early Thyroid RAIU and 99mPertechnetate Uptake Measurements Perchlorate Discharge Test Saliva to Plasma Radioiodide Ratio Measurement of Hormone Concentration and Other Iodinated Compounds and Their Transport in Blood Measurement of Total Thyroid Hormone Concentration in Serum Iodometry Radioligand and Immunometric Assays TT4 TT3 Measurement of Total and Unsaturated Thyroid Hormone-Binding Capacity in Serum In vitro Uptake Tests TBG Measurement Estimation of Free Thyroid Hormone Concentration Dialysable T4 and T3 by Isotopic Equilibrium Free T4 and T3 Index Methods Estimation of FT4 and FT3 by TBG Measurement Two-step Immunoassays Analogue (one-step) Immunoassays Measurements of Iodine-Containing Hormone Precursors and Products of Degradation 3.3′,5′-triiodothyronine of Reverse T3 (rT3) 3,5,-diiodothyronine (3,5-T2) 3,3′,-diiodothyronine (3,3′-T2) 3′,5′,-diiodothyronine (3′,5′,-T2) 3′-monoiodothyronine (3′-T1) 3-monoiodothyronine (3-T1) Tetra- and triiodothyroacetic acid (TETRAC and TRIAC) 3,5,3′-T3 sulfate (T3S) di- and monoiodityrosine (MIT and DIT) Thyroglobulin (Tg) Measurement of Thyroid Hormone and Its Metabolites in Other Body Fluids and in Tissues Urine Amniotic Fluid (AF) Cerebrospinal Fluid (CSF) Milk Saliva Effusions Tissues Tests Assessing the Effects of Thyroid Hormone on Body Tissues Basal Metabolic Rate (BMR) Deep Tendon Reflex Relaxation Time (Photomotogram)
Tests Related to Cardiovascular Function Miscellaneous Biochemical and Physiologic Changes Related to the Action of Thyroid Hormone on Peripheral Tissues Measurement of Substances Absent in Normal Serum Thyroid Autoantibodies Thyroid-Stimulating Immunoglobulins (TSI) Thyroid Stimulation Assays Standard in vivo Mouse Bioassay (LATS) In vitro Bioassays (animal or human tissue and recombinant TSH Receptor) Thyrotropin Binding Assays Thyroid Growth-Promoting Assay Other Substances with Thyroid-Stimulating Activity Exophthalmos-Producing Substance (EPS) Tests of Cell-Mediated Immunity (CMI) Anatomic and Tissue Diagnoses Thyroid Scintiscanning Radioiodide and 99mPertechnitate Scans Other Isotope Scans Fluorescent Scans Ultrasonography X-Ray and Related Procedures Computed Tomography (CT Scanning) Angiography Lymphography Thermography Magnetic Resonance Imaging (MRI) Biopsy of the Thyroid Gland Core Biopsy (Open od Closed) Percutaneous Fine-needle Aspiration (FNA) Evaluation of the Hypothalamic-Pituitary-Thyroid Axis Thyrotropin (TSH) Thyrotropin-Releasing Hormone (TRH) Test Other Tests of TSH Reserve Thyroid Stimulation Test Thyroid Suppression Test Specialized Thyroid Tests Iodotyrosine Deiodinase Activity Test for Defective Hormonogenesis Iodine Kinetic Studies Absorption of Thyroid Hormone Turnover Kinetics of T4 and T3 Metabolic Kinetics of Thyroid Hormones and Their Metabolites Measurement of the Production Rate and Metabolic Kinetics of Other Compounds Transfer of Thyroid Hormone from Blood to Tissues Applications of Molecular Biology in the Diagnosis of Thyroid Diseases
In Vivo Tests of Thyroid Gland Activity and Integrity of Hormone Synthesis and Secretion
Common to these tests is the administration to the patient of radioisotopes that cannot be distinguished by the body from the naturally occurring stable iodine isotope (127I). In contrast to all other tests, these procedures provide a means to directly evaluate thyroid gland function. Formerly these tests were used in the diagnosis of hypothyroidism and thyrotoxicosis, but this application has been supplanted by measurement of serum TSH and thyroid hormone concentrations in blood. Also, alterations of thyroid gland activity and in handling of iodine are not necessarily coupled to the amount of hormone produced and secreted. The tests are time consuming, relatively expensive and expose the patient to irradiation. Nevertheless, they still have some speccific applications including the diagnosis of inborn errors of thyroid hormonogenesis. Administration of isotopes is required for thyroid gland scanning used to demonstrate ectopic thyroid tissue and to establish the etiology of some forms of thyrotoxicosis. Finally, measurement of the thyroidal radioiodide uptake can be used as a means for estimating the dose of radioiodide to be delivered in the therapy of thyrotoxicosis and thyroid carcinoma.
To understand the physiological basis of this category of tests, one should remember the following facts. Iodine is an integral part of the thyroid hormone molecule. Although several other tissues (salivary glands, mammary glands, lacrimal glands, the choroid plexus, and the parietal cells of the stomach) can extract iodide from blood and generate a positive tissue to serum iodide gradient, only the thyroid gland stores iodine for an appreciable period of time. 1 Since the kidneys continually filter blood iodide, the final fate of most iodine atoms is either to be trapped by the thyroid gland or to be excreted in the urine. When a tracer of iodide is administered to the patient, it rapidly becomes mixed with the stable extrathyroidal iodide pool and is thereafter handled identically as the stable isotope. Thus, the thyroidal content of radioiodine gradually increases and that in the extrathyroidal body pool gradually declines, until virtually no free iodide is left. Normally this end point is reached between 24 and 72 hours.
From data of the radioiodide uptake by the thyroid gland and/or urinary excretion and/or stable iodide concentration in plasma and urine, the following parameters can be derived: (1) the rate of thyroidal iodine uptake (thyroid iodide clearance), (2) the fractional thyroid radioactive iodide uptake (RAIU), (3) the absolute iodide uptake (AIU) by the thyroid gland, and (4) the urinary excretion of radioiodide, or iodide clearance. After the complete removal of the administered radioiodide from the circulation, depletion of the radioisotope from the thyroid gland can be monitored by direct counting over the gland. Reappearance of the radioiodine in the circulation in protein-bound form can be measured and can be used to estimate the intrathyroidal turnover of iodine and the secretory activity of the thyroid gland.
The foregoing tests can be combined with the administration of agents known either to normally stimulate or to inhibit thyroid gland activity thus providing information on the control of thyroid gland activity. Administration of radioiodide followed by scanning allows us to examine the anatomy of functional tissue. The latter two applications of in vivo tests utilizing radioiodide will be discussed under their respective headings.
The potential hazard of irradiation resulting from the administered radioisotopes should always be kept in mind. Children are particularly vulnerable, and doses of X-rays as small as 20 rads to the thyroid gland are associated with increased risk of developing thyroid malignancies. 2 However, it must be noted that there is no proven danger from isotopes used for the diagnosis of thyroid diseases. 3 In vivo administration of radioisotopes is absolutely contraindicated during pregnancy and in breast-feeding mothers because of placental transport of isotope and excretion into breast milk.
A number of radioisotopes are now available. Furthermore, provision of more sophisticated and sensitive detection devices has substantially decreased the dose required for the completion of the studies. Table 6-2 5-7 lists the most commonly used isotopes for in vivo studies of the thyroid. Isotopes with slower physical decay, such as 125I and 131I, are particularly suitable for long-term studies. Isotopes with faster decay, such as 123I and 132I, usually deliver a lower irradiation dose and are advantageous in short-term and repeated studies. The peak photon energy gamma emission differs among isotopes, allowing the execution of simultaneous studies with two isotopes.
Table 6-2. Commonly Used Isotopes for In Vivo Studies and Radiation Dose Delivered
Principal Photon Energy (keV)
Estimated Radiation Dose (m rads/ÂµCi)
Average Dose Given for Scanning Purposes (ÂµCi)
ÃŸ (0.606 Mev)
ÃŸ (2.12 MeV)
aCalculations take into account the rate maximal uptake, and residence time of the isotope as well as gland size. For the iodine isotopes, average data for adult euthyroid persons used were: t-1/2 of uptake 5 hours, biologic t-1/2 50 days, maximal uptake 20%, and gland size 15 g (see also refs. [Quimby, 1970 #628;MIRD, 1975 #629;MIRD, 1976 #630]). bDose used for early thyroidal uptake studies.
Thyroidal Radioiodide Uptake (RAIU)
This is the most commonly used thyroid test requiring the administration of a radioisotope. It is usually given orally in a capsule or in liquid form and the quantity accumulated by the thyroid gland at various intervals of time is measured using a gamma scintillation counter. Correction for the amount of isotope circulating in the blood of the neck region, by subtracting counts obtained over the thigh, is of particular importance during the early periods following its administration. A dose of the same radioisotope, usually 10%, placed in a neck “phantom” is also counted as a “standard”. The percentage of thyroidal radioactive iodide uptake (RAIU) is calculated from the counts cumulated per constant time unit.
The percentage of RAIU 24 hours after the administration of radioiodide is most useful, since in most instances the thyroid gland has reached the plateau of isotope accumulation, and because it has been shown that at this time, the best separation between high, normal, and low uptake is obtained. Normal values for 24-hour RAIU in most parts of North America are 5 to 30 percent. In many other parts of the world, normal values range from 15 to 50 percent. Lower normal values are due to the increase in dietary iodine intake following the enrichment of foods, particularly mass produced bread (150 Âµg of iodine per slice), with this element. 8 The inverse relationship between the daily dietary intake of iodine and the RAIU test is clearly illustrated in Figure 6-1. The intake of large amounts of iodide (>5 mg/day), mainly from the use of iodine-containing radiologic contrast media, antiseptics, vitamins, and drugs such as amiodarone, suppresses the RAIU values to a level hardly detectable using the usual equipment and doses of the isotope. Depending upon the type of iodine preparation and the period of exposure, depression of RAIU can last for weeks, months, or even years. Even external application of iodide may suppress thyroidal radioiodide uptake. The need to inquire about individual dietary habits and sources of excess iodide intake is obvious.
Figure 6-1. Relation of 24 hour thyroidal radioiodide (I131) uptake (RAIU) to dietary content of stable iodine (I12 7 ). The uptake increases with decreasing dietary iodine. With iodine intake below the amount provided from thyroid hormone degradation, the latter contributes a larger proportion of the total iodine taken up by the thyroid. Under current dietary habits in the United States, the average 24-hour thyroidal RAIU is below 20 percent.
The test does not measure hormone production and release but merely the avidity of the thyroid gland for iodide and its rate of clearance relative to the kidney. Disease states resulting in excessive production and release of thyroid hormone are most often associated with increased thyroidal RAIU and those causing hormone underproduction with decreased thyroidal RAIU (Figure 6-2, below). Important exceptions include high uptake values in some hypothyroid patients and low values in some hyperthyroid patients. Increased thyroidal RAIU with hormonal insufficiency co-occur in the presence of severe iodide deficiency and in the majority of inborn errors of hormonogenesis (see Chapter 20 and 16 ). In the former, lack of substrate, and in the latter, a specific enzymatic block of hormone synthesis cause hypothyroidism poorly compensated by TSH-induced thyroid gland overactivity. Decreased thyroidal RAIU with hormonal excess is typically encountered in the syndrome of transient thyrotoxicosis (both de Quervain’s and painless thyroiditis), 9 ingestion of exogenous hormone (thyrotoxicosis factitia), iodide-induced thyrotoxicosis (Jod-Basedow disease), 10 and in patients with thyrotoxicosis on moderately high intake of iodide (see Table 6-3 ). High or low thyroidal RAIU as a result of low or high dietary iodine intake, respectively, may not be associated with significant changes in thyroid hormone secretion.
Figure 6-2. Examples of thyroidal RAIU curves under various pathological conditions. Note the prolonged uptake in renal disease due to decreased urinary excretion of the isotope and the early decline in thyroidal radioiodide content in some patients with thyrotoxicosis associated with a small but rapidly turning over intrathyroidal iodine pool.
Various factors including diseases that affect the value of the 24-hour thyroidal RAIU are listed in Table 6-3 . Several variations of the test have been devised which have particular value under special circumstances. Some of these are briefly described.
Table 6-3. Diseases and Other Factors That Affect the 24-Hour Thyroidal RAIU