Table 6-1^2–4 lists the isotopes most commonly used for in vivo studies of thyroid function. Isotopes with slower physical decay, such as ¹²⁵I and ¹³¹I, are particularly suitable for long-term studies. Conversely, isotopes with faster decay, such as ¹²³I and ¹³²I, usually deliver a lower radiation dose and are advantageous in short-term and repeated studies. Because the peak photon energy γ-emission differs among isotopes, simultaneous studies can be performed with two different isotopes.

Table 6-1

Commonly Used Isotopes for in Vivo Studies and Radiation Dose Delivered

^*Calculations take into account the rate of maximal uptake and the residence time of the isotope, as well as gland size. For the iodine isotopes, average data for adult euthyroid people used were a uptake of 5 hours, a biological of 50 days, maximal uptake of 20%, and gland size of 15 g (see also Quimby et al.⁴ and MIRD^2,3).

^†Dose used for early thyroid uptake studies.

Thyroidal Radioiodide Uptake

The iodide radioisotope usually is given orally in a capsule or in liquid form, and the quantity that is accumulated by the thyroid gland at various intervals is measured with a gamma scintillation counter. It is important to correct for the background activity of isotope circulating in the blood of the neck region (particularly during the early periods after administration). Background correction is achieved by subtracting counts obtained over the region of the thigh. A dose of the same radioisotope, usually 10%, placed in a neck “phantom” is also counted as a “standard.” The percentage of RAIU is calculated from the counts accumulated per constant time unit.

The percentage of RAIU 24 hours after the administration of radioiodide is most useful because in most instances, the thyroid gland has reached the plateau of isotope accumulation, and the best separation between high, normal, and low uptake is obtained at this time. Normal values for 24 hour RAIU in most parts of North America are 5% to 30%. In many other parts of the world, normal values range from 15% to 50%. Lower normal values are due to the increase in dietary iodine intake after the enrichment of foods, particularly mass-produced bread (150 µg of iodine per slice) containing this element. Over the past 3 decades, the mean ingestion of dietary iodine in the United States, although still within the recommended minimum for adults of 125 µg/day, has dramatically declined to approximately 240 to 300 µg/day for men and 190 to 210 µg/day for women.⁵ The inverse relationship between the daily dietary intake of iodine and the RAIU test is clearly illustrated in Fig. 6-1. Therefore, normal values of RAIU uptake will depend on the iodine content in a geographic region and also are related to age (with children having a higher iodine intake than adults). In Japan, the mean dietary iodine is six times higher than in the United States.

FIGURE 6-1 Relationship of 24-hour thyroidal radioiodide (¹³¹I) uptake (RAIU) to dietary content of stable iodine (¹²⁷I). Uptake increases with decreasing dietary iodine. If iodine intake is below the amount provided from thyroid hormone degradation, the latter contributes a larger proportion of the total iodine taken up by the thyroid. With dietary habits in the United States, the average 24 hour thyroidal RAIU is below 20%. (Data from DeGroot LJ, Reed Larsen P, Hennemann G, et al: The Thyroid and Its Diseases. New York, John Wiley & Sons, 1984.)

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 RAIU values to a level that is hardly detectable with the usual equipment and doses of isotope. Depending on 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 can suppress RAIU. It therefore is important to inquire about individual dietary habits and sources of excess iodide intake. Because dietary assessment of iodine ingestion can be somewhat inaccurate owing to the variable content of iodine added to various foods, measurement of iodine excretion is a more accurate assessment of the iodine balance. Spot urine iodine measurements were compiled from 1971 to 1974 and from 1988 to 1994 in the National Health and Nutrition Examination Surveys I and III, respectively, and have been found to be decreasing, in accordance with what was stated.⁶ Clinically, if one suspects that the patient had a large iodine load prior to an RAIU, a urine iodine measurement can be obtained. Urine iodine concentrations greater than 100 µg/day usually are associated with RAIU of 20% or less. Therefore, urine iodine can be useful in determining the feasibility of using RAIU.

RAIU is a measure of the avidity of the thyroid gland for iodide and its rate of clearance relative to the kidney, but results of this test do not equate with hormone production or release. Disease states resulting in excessive production of thyroid hormone most often are associated with increased thyroidal RAIU, and those causing hormone underproduction generally are associated with decreased thyroidal RAIU (Fig. 6-2). Some important exceptions to these rules include the high uptake values that are seen in certain hypothyroid patients and the low values noted in some hyperthyroid patients. Increased thyroidal RAIU with hormonal insufficiency can be caused by severe iodide deficiency and by most inborn errors of hormonogenesis. Lack of substrate in the former and specific enzymatic block of hormone synthesis in the latter cause hypothyroidism that is poorly compensated by TSH-induced thyroid gland overactivity. The increase in serum TSH, in response to the low circulating level of thyroid hormone, stimulates thyroidal iodine uptake by the NIS and hence increases RAIU. This can be a point of confusion for the clinician who is confronted with an increased RAIU in a patient who is suspected to have thyroiditis on the basis of blood tests. Alternatively, decreased thyroidal RAIU with hormonal excess typically is encountered in the syndrome of transient thyrotoxicosis (both deQuervain’s and painless thyroiditis) after the ingestion of exogenous hormone (thyrotoxicosis factitia), with iodide-induced thyrotoxicosis (Jod-Basedow disease), rarely in patients with metastatic functioning thyroid carcinoma or struma ovarii, and in patients with thyrotoxicosis who have a moderately high intake of iodide. High or low thyroidal RAIU as a result of low or high dietary iodine intake, respectively, might not be associated with significant changes in thyroid hormone secretion.

FIGURE 6-2 Examples of thyroidal radioiodide uptake curves under various pathologic conditions. Note the prolonged uptake in renal disease caused by 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. (Data from DeGroot LJ, Reed Larsen P, Hennemann G, et al: The Thyroid and Its Diseases. New York, John Wiley & Sons, 1984.)

Various factors, including diseases that affect the value of the 24 hour thyroidal RAIU, are listed in Table 6-2. Several variations of the RAIU test have been devised that have particular value under special circumstances. Some of these variations are briefly described.

Table 6-2

Diseases and Other Factors That Affect 24 Hour Thyroidal RAIU

Increased RAIU

Hyperthyroidism (Graves’ disease, Plummer’s disease, toxic adenoma, trophoblastic disease, resistance to thyroid hormone, TSH-producing pituitary adenoma)

Nontoxic goiter (endemic, inherited biosynthetic defects, generalized resistance to thyroid hormone, Hashimoto’s thyroiditis)

Excessive hormonal loss (nephrosis, chronic diarrhea, hypolipidemic resins, diet high in soybean)

Decreased renal clearance of iodine (renal insufficiency, severe heart failure)

Recovery of the suppressed thyroid (withdrawal of thyroid hormone and antithyroid drug administration, subacute thyroiditis, iodine-induced myxedema)

Iodine deficiency (endemic or sporadic dietary deficiency, excessive iodine loss as in pregnancy or in the dehalogenase defect)

TSH administration

Decreased RAIU

Hypothyroidism (primary or secondary)

Syndromes of TSH resistance

Thyroid dysgenesis (hypoplasia, ectopy, or agenesis)

Na/I symporter defect

Defect in iodide concentration (inherited trapping defect, early phase of subacute thyroiditis, transient hyperthyroidism)

Suppressed thyroid gland caused by thyroid hormone (hormone replacement, thyrotoxicosis factitia, struma ovarii)

Iodine excess (dietary, drugs, and other iodine contaminants)

Miscellaneous drugs and chemicals (see Tables 6-10 and 6-13)

RAIU, Radioactive iodine uptake; TSH, thyroid-stimulating hormone.

Early Thyroid Radioiodide Uptake and ^99mTc Uptake Measurements

The combination of severe thyrotoxicosis and a low intrathyroidal iodine concentration may result in an accelerated turnover rate of iodine in some patients. This produces a rapid initial uptake of radioiodide, which reaches a plateau before 6 hours, followed by a decline through release of the isotope in hormonal or other forms (see Fig. 6-2). Although this phenomenon is rare, some laboratories choose to routinely measure early RAIU, usually at 2, 4, or 6 hours. As was mentioned above, early measurements require accurate determination of the background activity contributed by circulating isotope. Radioisotopes with a shorter half-life, such as ¹²³I and ¹³²I, are more suitable in this context.

Because thyroidal uptake in the very early period after administration of radioiodide reflects mainly iodide trapping activity, ^99mTc as the pertechnetate ion (^99mTcO₄⁻) may be used. In euthyroid patients, thyroid trapping is maximal at about 20 minutes and is approximately 1% of the administered dose. This test, when coupled with the administration of triiodothyronine (T₃), has been used to evaluate thyroid gland suppressibility in thyrotoxic patients who are treated with antithyroid drugs.

Perchlorate Discharge Test

The perchlorate discharge test is used to detect defects in intrathyroidal iodide organification. It is based on the following physiologic principle. Iodide is “trapped” in the thyroid gland by an active transport mechanism that is mediated by NIS.⁷ Once in the gland, iodine is rapidly bound to thyroglobulin (Tg), and retention no longer requires active transport. Several ions, such as thiocyanate (SCN⁻) and perchlorate (ClO₄⁻), inhibit NIS-mediated iodide transport and cause release of the intrathyroidal iodide that is not bound to thyroid protein. Thus, intrathyroidal radioiodine loss after the administration of an inhibitor of iodide trapping measures intrathyroidal iodide that is not protein bound and indicates the presence of an iodide-binding defect.

In the standard test, epithyroid counts are obtained every 10 or 15 minutes after the administration of radioiodide. Two hours later, 1 g of KClO₄ is administered orally, and repeated epithyroid counts continue to be obtained for an additional 2 hours. We generally give this dose to patients 6 years or older and give 500 mg to children between the ages of 2 and 6 years. In normal individuals, radioiodide accumulation in the thyroid gland ceases after administration of the iodide transport inhibitor, and little or no loss of the accumulated thyroidal radioactivity occurs after induction of the “trapping” block. An organification defect is indicated if a loss of 5% or more is noted. The severity of the defect is proportional to the extent of radioiodide discharged from the gland and is complete when virtually all the activity accumulated by the gland is lost. The test is positive in the inborn defect of iodide organification caused by thyroid peroxidase (TPO) defects or by mutations in the chloride/iodide transport protein (pendrin) when associated with sensorineural deafness known as Pendred’s syndrome and in defects of the H₂O₂-generating enzyme, dual oxidase 2 (DUOX2), or its maturation factor, DUOXA2. The test may also be positive during the administration of iodide organification–blocking agents or after treatment with radioactive iodide.

Iodine Saliva-to-Plasma Ratio Test

An abnormal iodine (I⁻) saliva-to-plasma (S/P) ratio is pathognomonic of the iodine-trapping defect. The test can be carried out without interruption of thyroid hormone treatment. Furthermore, the measurement of I⁻ S/P can distinguish between a trapping defect and thyroid agenesis, which cannot be determined by RAIU. The I⁻ S/P ratio can be measured in a medical center without access to a gamma camera. The test is based on the observation that all tissues that normally concentrate iodide are affected by the trapping defect.⁸ The presence of an I⁻ transport defect in the parietal cells of the stomach and the choroid plexus of these patients has been used diagnostically by measurement of the gastric fluid-to-plasma and cerebrospinal fluid (CSF)-to-plasma ratios of radioiodide, following the administration of isotope.

One hour after the oral administration of 5 µCi of Na¹²⁵I, saliva is collected without stimulation over a period of 5 to 10 minutes. At the same time, a venous blood sample is obtained. After defrothing of the saliva and removal of cell debris by centrifugation (approximately 10 minutes at 500 × g) and separation of the serum, the S/P ratio of radioiodide is determined by counting equal volumes of these fluids in a gamma scintillation counter. A normal I⁻ S/P is 25. Affected individuals who are not able to concentrate iodide and therefore cannot secrete it into their saliva have a very low S/P. An I⁻ S/P of approximately 1 is diagnostic of a complete trapping defect, and a value between 1 and 20 is consistent with a partial defect.

Measurement of Hormone Concentration and Other Iodinated Compounds and Their Transport in Blood

The tests most commonly used for evaluating thyroid hormone–dependent metabolic status consist of measurements of free thyroid hormone concentrations. This approach is used because of the development of simple, sensitive, and specific methods for measuring these iodothyronines, and because of the lack of specific tests for direct measurement of the metabolic effects of these hormones on target tissues. Other advantages are the requirement of only a small blood sample and the large number of determinations that can be completed by a laboratory during a regular workday. In fact, although a clinician may entertain a diagnosis of thyroid dysfunction, the certainty of the diagnosis is confirmed only after measurement of the thyroid hormone concentrations and thyrotropin (TSH, see later).

The principal source of all hormonal iodine-containing compounds or their precursors is the thyroid gland, whereas peripheral tissues are the source of the products of their degradation. Their chemical structures and normal concentrations in serum are given in Fig. 6-3. It is important to note that the concentration of each substance is dependent not only on the amount synthesized and secreted by the thyroid gland, but also on its affinity for carrier serum proteins, distribution in tissues, rate of degradation, and, finally, clearance.

FIGURE 6-3 Iodine-containing compounds in the serum of healthy adults. a, Iodothyronine concentrations in the euthyroid population are not normally distributed. Therefore, calculation of the normal range on the basis of 95% confidence limits for a Gaussian distribution is accurate. b, Significant decline with old age. c, Probably an overestimation because of cross-reactivity by related substances.

Quantitatively, the major secretory product of the thyroid gland is thyroxine (T₄), with T₃ being next in relative abundance. They are synthesized and stored in the thyroid gland as part of a larger molecule, Tg, which is degraded to release the two iodothyronines in a ratio favoring T₄ by 10- to 20-fold. Under normal circumstances, only minute amounts of Tg escape into the circulation. On a molar basis, it is the least abundant iodine-containing compound in blood. With the exception of T₄, Tg, and small amounts of diiodotyrosine (DIT) and monoiodotyrosine (MIT), all other iodine-containing compounds that are found in normal human serum are produced mainly in extrathyroidal tissues by a stepwise process of deiodination of T₄. An alternative pathway of T₄ metabolism that involves deamination and decarboxylation but retention of the iodine residues gives rise to tetraiodothyroacetic acid (TETRAC) and triiodothyroacetic acid (TRIAC).9,10 Conjugation to form sulfated iodoproteins also occurs. Sulfoconjugates of T₄, T₃, and reverse T₃ (rT₃) have been identified in human biological fluids. Additionally, maternal serum levels of 3,3′-diiothyronine sulfate (T₂S) may reflect on the status of fetal thyroid function. Circulating iodalbumin is generated by intrathyroidal iodination of serum albumin. Small amounts of iodoproteins may be formed in peripheral tissues or in serum by covalent linkage of T₄ and T₃ to soluble proteins. The physiologic function of circulating iodine compounds other than T₄ and T₃ remains unknown, with the exception of rT₃. rT₃ levels are elevated during fasting and during significant nonthyroidal illness. In such instances, measurement of rT₃ can help the clinician to distinguish between these conditions and central hypothyroidism.

Measurement of Total Thyroid Hormone Concentration in Serum

Iodometry

Because iodine is an integral part of the thyroid hormone molecule, it is not surprising that determination of the iodine content in serum was the first method used over 6 decades ago for the identification and quantitation of thyroid hormone.¹¹ Measurement of protein-bound iodine was the earliest method used routinely for the estimation of thyroid hormone concentration in serum. This test measured the total quantity of iodine precipitable with serum proteins, 90% of which is T₄. The normal range was 4 to 8 mg of iodine per deciliter of serum.

Efforts to measure serum thyroid hormone levels with greater specificity and with lesser interference from nonhormonal iodinated compounds led to the development of measurement of butanol-extractable iodine and T₄ iodine by column techniques. All such chemical methods for the measurement of thyroid hormone in serum have been replaced by ligand assays, which are devoid of interference by even large quantities of nonhormonal iodine-containing substances.

Table 6-3

Conditions Associated With Changes in Serum TT₄ Concentration and Relationship to Clinical Status

T₃, Triiodothyronine; T₄, thyroxine; TBG, thyroxine-binding globulin; TSH, thyroid-stimulating hormone; TT₄, total T₄.

Serum TT₄ levels are low in conditions that are associated with decreased TBG concentrations, in the presence of abnormal TBGs with reduced binding affinity, and when the available T₄-binding sites on TBG are partially saturated by competing drugs present in blood in high concentration (Table 6-4). Conversely, TT₄ levels are high when the serum TBG concentration is high. In this situation, the person remains euthyroid provided that feedback regulation of the thyroid gland is intact.

Table 6-4

Compounds That Affect Thyroid Hormone Serum Transport Proteins

NSAID, Nonsteroidal antiinflammatory drug; o,p′-DDD, 2,4′-dichlorodiphenyldichloroethane (mitotane); TBG, thyroxine-binding globulin; TTR, transthyretin.

Although changes in transthyretin (TTR) concentration rarely give rise to significant alterations in TT₄ concentration, the presence of a variant serum albumin with high affinity for T₄ or antibodies against T₄ produce apparent elevations in the measured TT₄ concentration, whereas the metabolic status remains normal. The variant albumin is inherited as an autosomal dominant trait termed familial dysalbuminemic hyperthyroxinemia (FDH).

Another possible cause of discrepancy between the observed serum TT₄ concentration and the metabolic status of the patient is divergent changes in serum total T₃ (TT₃) and TT₄ concentrations with alterations in the serum T₃/T₄ ratio. The most common situation is that of an elevated TT₃ concentration. The source of T₃ may be endogenous, as in T₃ thyrotoxicosis, or exogenous, as during ingestion of T₃. In the former situation, contrary to the common variety of thyrotoxicosis, elevation in the serum TT₃ concentration is not accompanied by an increase in the TT₄ level. In fact, the serum TT₄ level is normal and occasionally low. This finding indicates that the pathogenesis of T₃ thyrotoxicosis is the direct secretion of T₃ from the thyroid gland rather than the peripheral conversion of T₄ to T₃. Ingestion of pharmacologic doses of T₃ results in thyrotoxicosis associated with severe depression of the serum TT₄ concentration. Moderate hypersecretion of T₃ can be associated with euthyroidism and a low serum TT₄ concentration. This situation, occasionally referred to as T₃ euthyroidism, may be more prevalent than T₃ thyrotoxicosis. It is believed to constitute a state of compensatory T₃ secretion as a physiologic adaptation of the failing thyroid gland, such as after treatment of thyrotoxicosis, in some cases of chronic thyroiditis, or during iodine deprivation. The serum TT₄ concentration is also low in normal persons receiving replacement doses of T₃. Conversely, serum TT₄ levels are above the upper limit of normal in 15% to 50% patients treated with exogenous T₄ and having normal serum TSH. Because of the relatively slow rate of metabolism and the large extrathyroidal T₄ pool, the serum concentration of the hormone varies little with the time of sampling in relation to ingestion of the daily dose.

Serum Total T₃

Triiodothyronine is principally responsible for the effects of thyroid hormones on the target organs. T₃ is formed extrathyroidally via 5′ deiodination of T₄. Thus, serum T₃ concentration reflects the functional state of the peripheral tissue rather than the secretory performance of the thyroid gland. Like T₄, 99% of T₃ is present in protein-bound form. However, the affinity of T₃ to TBG is 10-fold lower than that of T₄.

Normal serum TT₃ concentrations in the adult range from 80 to 190 ng/dL (1.2 to 2.9 nmol/L). Sex differences are small, but age differences are more dramatic. In contrast to serum TT₄, the TT₃ concentration at birth is low, about half the normal adult level. It rises rapidly within 24 hours to about double the normal adult value, followed by a decrease over the subsequent 24 hours to a level in the upper adult range, which persists for the first year of life. A decline in the mean TT₃ level has been observed in old age, although not in healthy subjects,23,24 which suggests that a fall in TT₃ might reflect the prevalence of nonthyroidal illness rather than an effect of age alone. Although a positive correlation between serum TT₃ level and body weight has been observed, this might be related to overeating.²⁵ Rapid and profound reductions in serum TT₃ can be produced within 24 to 48 hours of total calorie or carbohydrate-only deprivation.

Most conditions that cause serum TT₄ levels to increase are associated with high TT₃ concentrations. Thus, serum TT₃ levels are usually elevated in thyrotoxicosis and reduced in hypothyroidism. However, in both conditions, the TT₃/TT₄ ratio is elevated relative to normal euthyroid persons. This elevation is due to the disproportionate increase in serum TT₃ concentration in thyrotoxicosis and a lesser diminution in hypothyroidism relative to the TT₄ concentration. Accordingly, measurement of the serum TT₃ level is a more sensitive test for the diagnosis of hyperthyroidism, and measurement of TT₄ is more useful in the diagnosis of hypothyroidism.

Under certain conditions, changes in the serum TT₃ and TT₄ concentrations are disproportionate or occur in the opposite direction (Table 6-5). Such conditions include the syndrome of thyrotoxicosis with normal TT₄ and FT₄ levels (T₃ thyrotoxicosis). In some patients, treatment of thyrotoxicosis with antithyroid drugs normalizes the serum TT₄ but not the TT₃ level and produces a high TT₃/TT₄ ratio. In areas of limited iodine supply and in patients with limited thyroidal ability to process iodide, euthyroidism can be maintained at low serum TT₄ and FT₄ levels by increased direct thyroidal secretion of T₃. Although these changes have a rational physiologic explanation, the significance of discordant serum TT₄ and TT₃ levels under other circumstances is less well understood.

Table 6-5

Conditions That May Be Associated With Discrepancies Between the Concentration of Serum TT₃ and TT₄

TT₃, Total triiodothyronine; TT₄, total thyroxine.

^*Artifactual values depend on the method of hormone determination in serum.

^†Hepatic and renal failure, diabetic ketoacidosis, myocardial infarction, infectious and febrile illness, cancers.

^‡Glucocorticoids, iodinated contrast agents, amiodarone, propranolol, propylthiouracil.

The most common cause of discordant serum concentrations of TT₃ and TT₄ is a selective decrease in serum TT₃ caused by decreased conversion of T₄ to T₃ in peripheral tissues. This reduction is an integral part of the pathophysiology of a number of nonthyroidal acute and chronic illnesses and calorie deprivation. In these conditions, the serum TT₃ level is often lower than that commonly found in patients with frank primary hypothyroidism. However, no clear clinical evidence of hypometabolism is found in this situation. In some individuals, decreased T₄-to-T₃ conversion is an inherited condition.²⁶ A combination of high TT₃ and low TT₄ is typical in subjects with loss-of-function mutations in the iodothyronine cell membrane transporter, MCT8.²⁷

A variety of drugs are responsible for producing changes in the serum TT₃ concentration without apparent metabolic consequences. Drugs that compete with hormone binding to serum proteins decrease serum TT₃ levels, generally without affecting the free T₃ (FT₃) concentration (see Table 6-4). Some drugs such as glucocorticoids²⁸ depress the serum TT₃ concentration by interfering with the peripheral conversion of T₄ to T₃. Others, such as phenobarbital,²⁹ depress the serum TT₃ concentration by stimulating the rate of intracellular hormone degradation and clearance. Most have multiple effects. These effects are combinations of those described above, as well as inhibition of the hypothalamic-pituitary axis or thyroidal hormonogenesis.

Changes in serum TBG concentration have an effect on the serum TT₃ concentration similar to that on TT₄. The presence of endogenous antibodies to T₃ can also result in apparent elevation of serum TT₃, but as in the case of high TBG, it does not cause hypermetabolism.

Administration of commonly used replacement doses of T₃, usually on the order of 75 µg/day or 1 µg/kg body weight per day,³⁰ results in serum TT₃ levels in the thyrotoxic range. Furthermore, because of rapid gastrointestinal absorption and a relatively fast degradation rate, the serum level varies considerably according to the time of sampling in relation to hormone ingestion.

Measurement of Total and Unsaturated Thyroid Hormone–Binding Capacity in Serum

The concentration of thyroid hormone in serum is dependent on its supply, as well as on the abundance of hormone-binding sites on serum proteins; therefore, estimation of the latter has proved useful in the correct interpretation of values obtained from measurement of the total hormone concentration. These results have been used to provide an estimate of the free hormone concentration, which is important in differentiating changes in serum total hormone concentrations caused by alterations in binding proteins in euthyroid patients from those caused by abnormalities in thyroid gland activity that give rise to hypermetabolism or hypometabolism.

In Vitro Uptake Tests

In vitro uptake tests measure the unoccupied thyroid hormone–binding sites on TBG. They use labeled T₃ or T₄ and some form of synthetic absorbent to measure the proportion of radiolabeled hormone that is not tightly bound to serum proteins. Because ion exchange resins often are used as absorbents, the test became known as the resin T₃ or T₄ uptake test, which describes the technique rather than the entity measured.

The test is usually carried out by incubating a sample of the patient’s serum with a trace amount of labeled T₃ or T₄. The labeled hormone, not bound to available binding sites on TBG present in the serum sample, is absorbed onto an anion exchange resin and measured as resin-bound radioactivity. Values correlate inversely with the concentration of unsaturated TBG. Various methods use different absorbing materials to remove the hormone that is not tightly bound to TBG. Labeled T₃ generally is used because of its less firm, yet preferential, binding to TBG. Depending on the method, typical normal results for T₃ uptake are 25% to 35% or 45% to 55%. Thus, it is more valuable to express results of the uptake tests as a ratio of the result obtained in a normal control serum run in the same assay as the test samples. Normal values will then range on either side of 1.0, usually from 0.85 to 1.15.

Uptake of tracer by the absorbent is inversely proportional to the number of unsaturated binding sites (unoccupied by endogenous thyroid hormone) in serum TBG. Thus, uptake is increased when the amount of unsaturated TBG is reduced as a result of excess endogenous thyroid hormone or a decrease in the concentration of TBG. In contrast, uptake is decreased when the amount of unsaturated TBG is increased as a result of a low serum thyroid hormone concentration or an increase in the concentration of TBG. Because the test can be affected by either or both independent variables—serum total thyroid hormone and TBG concentrations—the results cannot be interpreted without knowledge of the hormone concentration. As a rule, parallel increases or decreases in serum TT₄ concentration and the T₃ uptake test indicate hyperthyroidism and hypothyroidism, respectively, whereas discrepant changes in serum TT₄ and T₃ uptake suggest abnormalities in TBG binding. However, abnormalities in hormone and TBG concentrations can coexist in the same patient. For example, a hypothyroid patient with a low TBG level will typically show a low TT₄ level and normal T₃ uptake results (Fig. 6-4). Several nonhormonal compounds, because of structural similarities, compete with thyroid hormone for its binding site on TBG. Some are used as pharmacologic agents and thus may alter the in vitro uptake test, as well as the total thyroid hormone concentration in serum. A list is provided in Table 6-4.

FIGURE 6-4 Graphic representation of the relationship between the serum total thyroxine (T₄) concentration, the resin triiodothyronine uptake (rT₃U) test, and the free T₄ (FT₄) concentration in various metabolic states and in association with changes in thyroxine-binding globulin (TBG). The principle of communicating vessels is used as an illustration. The height of fluid in the small vessel represents the level of FT₄; the total amount of fluid in the large vessel, the total T₄ concentration; and the total volume of the large vessel, the TBG capacity. Dots represent resin beads; black dots represent those carrying the radioactive T₃ tracer (T₃*). The rT₃U test result (black dots) is inversely proportional to the unoccupied TBG-binding sites represented by the unfilled capacity of the large vessel.

TBG and TTR Measurements

The concentrations of TBG and TTR in serum can be estimated by measurement of their total T₄-binding capacity at saturation or measured directly by immunologic techniques.31,32

The TBG concentration in serum can be determined by RIA,³² and both TBG and TTR can be measured by Laurell’s rocket immunoelectrophoresis, by radial immunodiffusion, or by enzyme immunoassay; commercial methods are available. The true mean value for TBG is 1.6 mg/dL (260 nmol/L), with a range of 1.1 to 2.2 mg/dL (180 to 350 nmol/L) in serum. In adults, the normal range for TTR is 16 to 30 mg/dL (2.7 to 5.0 mmol/L). Concentrations of TBG and TTR in serum vary with age, gender, pregnancy, and posture. Determination of the concentration of these proteins in serum is particularly helpful for evaluation of extreme deviations from normal, as in congenital abnormalities of TBG. In most instances, however, the in vitro uptake test, in conjunction with the serum TT₄ level, gives an approximate estimation of the TBG concentration.