Endocrine Laboratory Pearls: Thyroid Tests
Physicians are comfortable ordering and interpreting familiar laboratory tests, but they might not recognize some of the tests’ limitations, resulting in overutilization or in misguided confidence in the validity of the results. In addition, less-familiar laboratory tests that could improve patient care or reduce costs might be ordered too infrequently because of uncertainty about testing indications or result interpretation.
Free Thyroid Hormone Tests
Stefan K. Grebe, MD, of the Division of Clinical Biochemistry and Immunology and the Department of Laboratory Medicine and Pathology at Mayo Clinic in Rochester, Minnesota, says: “Tests for free thyroid hormone are among the most frequently ordered laboratory tests, despite consensus that measurement of thyrotropin (TSH) should usually suffice.” Dr Grebe explains that peripheral thyroid hormone testing should be limited to a few clinical scenarios:
- Confirmation of hypothyroidism or hyperthyroidism in cases with borderline abnormal serum TSH concentrations
- Assessment of severity of hyperthyroidism or hypothyroidism
- Rapidly changing thyroid hormone levels (eg, during treatment of thyrotoxicosis)
- An unreliable pituitary-thyroid feedback loop (eg, pituitary disease)
- TSH assay interferences due to TSH autoantibodies or heterophilic antibodies
Dr Grebe notes: “In many such cases, measurement of total thyroid hormones is just as informative as testing for free thyroid hormones while being analytically more reliable (Figure 1). Moreover, free thyroid hormone assays are only marginally less susceptible to interferences from drugs or nonthyroidal illness than total thyroid hormone assays. For example, heparin (and low-molecular-weight heparin) elevates lipoprotein lipase levels, creating increased circulating concentrations of free fatty acid. These fatty acids displace thyroid hormones from binding proteins, elevating free thyroxine (FT4) and free triiodothyronine (FT3) levels, in some cases more than 2-fold above the upper limit of the reference ranges.”
Figure 1. Thyroid function tests. Range of result differences of thyroid function tests on aliquots of the same sample, measuredwith 15 different immunoassays. This sample levels of thyroid hormones and thyroid hormone-binding proteins within the normal
reference range. In samples with abnormal concentrations of thyroid hormones or thyroid hormone-binding proteins, even larger differences
are observed between the assays for free thyroxine (FT4) and free triiodothyronine (FT3).
TT4 indicates total thyroxine; TT3, total triiodothyronine
Ravinder J. Singh, PhD, in the Division of Clinical Biochemistry and Immunology and the Department of Laboratory Medicine and Pathology at Mayo Clinic in Minnesota, explains: “Even in the absence of interferences, many FT4 and FT3 assays give inaccurate results in some cases. Most of these assays use thyroid hormone analogues, designed to not displace thyroid hormone from binding proteins while competing with the patient’s free thyroid hormone for assay antibodies. This assay design works only over a relatively narrow range of concentrations of binding proteins and thyroid hormones. Reference methodologies using physical separation of bound thyroid hormone from free thyroid hormone solve this problem, but they are only available for FT4, have longer turnaround times, and continue to be susceptible to interferences related to drugs or illness.”
Measurement of Circulating Thyroid-Stimulating Immunoglobulins
Thyrotoxicosis affects about 3 million new patients in the United States each year. More than 60% of cases are caused by Graves’ disease, a disorder characterized by production of autoantibodies (thyroid-stimulating immunoglobulins [TSIs]) that stimulate the TSH receptor. Since TSIs are disease specific and are detectable in more than 90% of patients with Graves’ disease, they reliably distinguish Graves’ disease from other causes of thyrotoxicosis.
Dr Singh says: “There are 2 different types of clinical assays for TSI detection: TSH receptor autoantibody–binding (TRAB) assays and TSI bioassays. In TRAB assays, labeled TSH competes with TSI in patient serum for binding to assay TSH receptors. TSI bioassays use cell lines that express the TSH receptor and a cyclic adenosine monophosphate (cAMP)-controlled luciferase gene. When these cells are exposed to TSIs, cAMP is produced and thus drives luciferase production, which in turn leads to light production upon cell lysis and substrate addition.”
Dr Grebe explains: “TRAB assays and TSI bioassays show about 90% agreement in detecting TSIs, with the latter being somewhat more sensitive at low TSI concentrations and the former possibly giving more accurate results at high TSI concentrations (Figure 2). Either assay is more accurate (and cheaper) than a radioactive iodine uptake or scan, which are traditionally used to differentiate Graves’ disease from other causes of thyrotoxicosis. TRAB assays and TSI bioassays are also particularly useful in distinguishing hyperemesis gravidarum–related thyrotoxicosis from a first-trimester presentation of Graves’ disease.”
Figure 2. Serial dilution curves of international standard material (IS-90/672) of thyroid-stimulating immunoglobulins (TSIs). The 2 curves are parallel to each other along their linear portions, indicating that the 2 tests have similar responses to IS-90/672. The TSI bioassay curve is shifted to the right, suggesting better detection sensitivity than the thyroxine receptor autoantibody–binding (TRAB) assay. At very high TSI concentrations, the TSI bioassay might be less accurate than the TRAB assay because of a high-dose hook effect. The reference range for the TSI bioassay is a TSI index of <1.3; the reference range for the TRAB assay is <16% thyrotropin (TSH)-binding inhibition.
He continues: “Another key application during pregnancy is risk assessment for fetal/neonatal Graves’ disease. This disorder can occur in pregnant women who had previous thyroid-ablative treatment for Graves’ disease. These women have normal thyroid function test results, but might still be producing TSIs, which can pass through the placenta to the infant and cause fetal thyrotoxicosis. Results of maternal TRAB assay or TSI bioassay that are more than 2 or 3 times the upper limit of the reference ranges are correlated with fetal thyrotoxicosis, indicating a need for high-risk obstetric care and serial TRAB assays or TSI bioassays.”
Reprinted from Mayo Clinic Endocrinology Update 2011; 6(2): 6-7. Used with permission.
Ehrlichia: Emergence of a New Pathogenic Species
Ehrlichia are intracellular, gram-negative bacteria transmitted by ticks to humans and animals. In humans, Ehrlichia bacteria infect human leukocytes, and severe disease may be associated with gastrointestinal, renal, respiratory, and central nervous system involvement and, rarely, death.
A new species of Ehrlichia was identified in 2009 by Mayo Clinic in the DNA from 4 patients (3 from Wisconsin, 1 from Minnesota) using a polymerase chain reaction (PCR)-based assay. Since then, more than 25 patients from Minnesota and Wisconsin have been identified with this infection. A specimen from a North Dakota resident was also found to be positive for the new species, but epidemiologic investigations revealed that he likely acquired the infection in Minnesota. The initially identified patients had symptoms consistent with ehrlichiosis and recovered following treatment with doxycycline, an antibiotic used to treat Ehrlichia infection.
The unique nucleotide sequence of the new species is similar to Ehrlichia muris, a mouse pathogen not previously identified from humans in North America. This new species has also been found in deer ticks (Ixodes scapularis) and white-footed mice (Peromyscus leucopus) collected from Minnesota and Wisconsin.
Prior to this discovery, human disease caused by any Ehrlichia species was not thought to be present in Minnesota and Wisconsin. Therefore, physicians need to know to test for ehrlichiosis in patients who reside or have recently visited these states so the diagnosis is not missed. Commercially available serologic assays for Ehrlichia chaffeensis or Anaplasma phagocytophilum do not provide a specific diagnosis and may miss a positive case. Mayo Medical Laboratories offers #84319 Ehrlichia/Anaplasma, Molecular Detection, PCR, Blood, which detects DNA from the new Ehrlichia species, as well as Ehrlichia chaffeensis, Ehrlichia ewingii, and Anaplasma phagocytophilum. A positive identification for any of these organisms supports the diagnosis of ehrlichiosis or anaplasmosis. This is the preferred test for diagnosis of acute disease with any of these 4 pathogens.