1. Thyroid Function Tests
Hasan AYDIN, MD
Yeditepe University Medical Faculty
Endocrinology and Metabolism

2. Tests of Thyroid Function:
Tests of thyroid hormones in the blood
Evaluation of hypothalamo-pituitary-thyroid axis
Assessment of iodine metabolism
Estimation of gland size
Thyroid biopsy
Observation of the effects thyroid hormones on peripheral tissues
Measurement of thyroid autoantibodies

3. Tests of Thyroid Hormones in Blood
Total serum T4
Free T 4 level
Total serum T3
FT4-I
FT3 and rT3
Thyroglobulin

4. Total T3 and T4 Protein bound + free hormone
Measured by radioimmunoassay(RIA) or immunufluorescent assay.
Provide reliable index of thyroid gland activity if levels of thyroid binding proteins normal.
Changes in serum concentration of thyroid-binding proteins(TBP) or the presence of drugs that effects binding of T3 and T4 to TBP will modify total hormone levels, but not the biologically active free T3 and T4 levels.

5. Total T3 (Normal: 95-190 ng/dl)
The T3 is increased in almost all cases of hyperthyroidism and usually goes up before the T4 does.
T3 is a more sensitive indicator of hyperthyroidism than the Total T4.
In hypothyroidism the T3 is often normal even when the T4 is low.
The T3 is decreased during acute illness and starvation, and is affected by several medications including beta blocker, steroids and amiodarone.

6. Total T4 (Normal: 5-12 ng/dl)
In circulation most T4 (99.8%) is bound to specific plasma proteins. TBG (60-75%), Prealbumin/transthyretin 15-30%, albumin 10%
In the absence of thyroid dysfunction, abnormal results are usually due to an abnormal level of serum TBG.
The TT4 level may not always correspond to the FT4 concentration
Free hormone is only 0.03 % of the total.

7. Factors Influencing Concentration of Protein Bound Thyroid Hormones
A- Increased TBG concentration
Congenital
Hyperestrogenic states: Pregnancy, Estrogen therapy
Diseases: Acute infectious hepatitis, hypothyroidism
B- Decreased TBG concentration
Drugs: Androgenic steroids, glucocorticoids
Major systemic illness: Protein malnutrition, nephrotic syndrome, cirrhosis, hyperthyroidism
C- Drugs affecting thyroid hormone binding to TBG
Phenytoin Salycilates Diazepam......

8. Free T3 and Free T4 Free T4 (Normal: 0.9 - 2 ng/dl)
Free T3 (N: ng/dl) and T4 hormone levels can be
measured directly by dialysis method
by chemiluminescent assay
estimated using free thyroxine index
Free T4 (Normal: ng/dl)
The FT4 measures the concentration of free thyroxine, the only biologically active fraction in the serum.
The free thyroxine is not affected by changes in concentrations of binding proteins Thus such conditions as pregnancy, or estrogen and androgen therapy do not affect the FT4.

9. Thyroid Stimulating Hormone
A glycoprotein secreted by the anterior pituitary
Structurally similar to FSH, LH and chorionic gonadotropin (hCG)
Composed of two dissimilar non-covalently bound α-subunit and β-subunits
The α-subunit is hormone non-specific
The β-subunit is distinct for each hormone and confers biological and immunological specificity
Antibodies used in immunoassays of TSH target the β-subunit

10. Thyroid Stimulating Hormone
Measures the response of the pituitary gland to circulating T4 and T3.
Elevated in hypothyroidism due to primary thyroid disease, iodine deficient goiter.
In hyperthyroidism, TSH will be suppressed.
This is probably the most important test that can be performed to assess thyroid function.
Highly sensitive TSH test is available. This method utilizes monoclonal antibodies directed against the beta subunit of the molecule and is therefore specific for TSH.

11. The effect of serum TSH assay sensitivity on the discrimination of euthyroid subject (Euth) from those with thyrotoxicosis

12. Measurement of TSH Methods Principle Comments
Radioimmunoassay Competitive binding of
(RIA) radiolabeled TSH and non- Currently being
labeled TSH to limited binding phased out
sites on antibody
Immunoradiometric Binding of TSH to radiolabeled Most sensitive
assay antibody and widely used
ELISA Binding of TSH to enzyme-
labeled antibody
ILMA Binding of TSH to luminescent
labeled-labeled antibody Sensitive

13. Evaluation of Hypothalamo-Pituitary-Thyroid Axis
The integrity of the hypothalamo-pituitary-thyroid axis can be evaluated by
the response of the pituitary gland to thyroid hormone excess or deficiency;
the ability of the thyroid gland to respond to thyrotropin (TSH);
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; primary (thyroid), secondary (pituitary), or tertiary (hypothalamic) malfunction.

14. TRH Test Procedure: IV injection of
TRH µg. Time-course measurement of TSH -15, 0, 15 and 30 mn.
Normal and abnormal responses to thyrotropin-releasing hormone. (Top panel) The delayed thyroid-stimulating hormone (TSH) response to thyrotropin-releasing hormone (TRH) in a patient with hypothalamic disease. Peak plasma TSH values are reached at approximately 60min following exogenous TRH, compared to the normal, where the peak occurs at 20-30min. In pituitary hypothyroidism, TSH reserve may be severely reduced and there may be only a slight, or no response, to exogenous TRH. (Bottom panel) The normal rise in circulating plasma triiodothyronine (T3) following an injection of TRH. The T3 response lags behind the TSH response, reflecting the fact that T3 rises secondary to stimulation of the thyroid gland by the released TSH.
Diminished resp to TRH is observed with pharmacologic doses of glucocorticoid use, levodopa, bromocriptine, dopamine; resp is augmented with dopaminergic antagonists domperidone, metoclopropamide
examines interrelationship of TRH and TSH secretions by administering exogenous TRH

15. Clinical Utility of Thyroid-Related Laboratory Tests
Name of Test Abbreviation Clinical Utility
Tests for Evaluation of Thyroid Status
Thyrotropin (sensitive IA) sTSH Best general test
(Conventional RIA) TSH phased out
Free thyroxine (appropriate method) FT Second best general test
Free (3,5,3') triiodothyronine FT Adjunct test, diagnosis of
T3 toxicosis
Total (3,5,3') triiodothyronine T Used in lieu of FT3
Total thyroxine T Inadequate as general test
Thyroglobulin Tg Valuable in thyroid cancer
Free T4 index FT4/FTI
T4 x T3-BR Replaced by FT4
T4 x T4-BR

16. Thyroglobulin (Tg) In serum by double antibody immunoassay
The normal range vary with method and laboratory ,
Normal range is less than 40 ng/mL ( < 40 mikrogram/L) in the euthyroid individual
<5 ng/mL in a thyroidectomized individual.
The major problem endogenous Tg Ab
Spuriously low or spuriously high values.

17. Thyroglobulin (Tg) Elevations
Graves ' disease
Toxic MNG
Acute or chronic thyroiditis
Papillary or follicular thyroid carcinoma.
Following thyroidectomy and 131-I therapy,
serum thyroglobulin greater than 10 ng/dL (> 10 ~g/L) indicates the presence of metastatic disease
rise in a patient with known metastases indicates progression of the disease.

18. Thyroid Autoantibodies
Thyroid peroxidase (TPO) (or antimicrosomal (TMAb) and antithyroglobulin (TgAb) antibodies are measured to confirm or rule out autoimmune thyroid disease. High titers of these antibodies are present in 97 % of patients with Grave’s disease or Hashimoto’s thyroiditis. Not specific for the type of disease.
Generally these are measured using RIA, ELISA or IRMA techniques
Reported prevalence of thyroid antibodies in population is highly variable ( %2-30)
Thyroid receptor stimulating antibody is characteristic of Grave’s disease (Present in 90%). Helpful in diagnosis of euthyroid Grave’s ophtalmopathy and in predicting possible occurrence of thyroid dysfunction in neonates
Thyroid receptor blocking antibody is characteristic of autoimmune thyroid disease

19. Thyroid peroxidase (TPO) and Antithyroglobulin (TgAb)
Measured to confirm or rule out autoimmune thyroid disease.
High titers of these antibodies are present in 97 % of patients with Grave’s disease or Hashimoto’s thyroiditis.
Not specific for the type of disease.
Generally these are measured using RIA, ELISA or IRMA techniques
Reported prevalence of thyroid antibodies in population is highly variable (2-30%)

20. Iodine Metabolism & Biosynthetic Activity
Radioactive iodine uptake
RAIU supression test

21. RAIU
Radioactive iodine allows assessment of the turnover of iodine by the thyroid gland in vivo.
Iodine-123 is the ideal isotope for this purpose
Administered orally at a dose of micCi,
Radioactivity over the thyroid area is measured with a scintillation counter at 4 or 6 hours and again at 24 hours

22. RAIU Vary with the iodide intake.
In areas of low iodide intake and endemic goiter, the 24-hour RAIU may be as high as 60-90%.
The normal uptake at 6 hours is 5-15% and at 24 hours 8-30%.
In thyrotoxicosis due to Graves' disease or toxic nodular goiter, the 24- hour RAIU is markedly elevated

23. RAIU Supression Test
microgram of T3 in divided doses daily for 5 days
Reduce the 24-hour RAIU by more than 50% (suppression test).
Failure of the thyroid to suppress on this treatment indicates autonomous thyroid function, as in Graves' disease, or autonomously functioning thyroid nodules.

24. Thyrotoxicosis with a Very Low Thyroidal RAIU
Subacute thyroiditis
During the active phase of Hashimoto's thyroiditis,
Thyrotoxicosis factitia due to oral ingestion of a large amount of thyroid hormone
As a result of excess iodide intake (amiodarone therapy), inducing thyrotoxicosis in a patient with latent Graves ' disease or multinodular goiter, the low uptake being due to the huge iodide pool;
Struma ovarii
Ectopic functioning metastatic thyroid carcinoma after total thyroidectomy.

25. Thyroid Imaging Radionuclide imaging Radioisotope Scanning
Fluorescent Scanning
Thyroid Ultrasonography
Magnetic Resonance Imaging

26. Radionuclide Imaging 123-Iodine
oraIly in dose of microCi, scan of the thyroid at 8-24 hours
Technetium Tc 99 m pertechnetate (99mTc as 4) intravenously in a dose of 1-10 mCi scan is obtained at minutes.
Images can be obtained either a rectilinear scanner or a gamma camera.

27. Radionuclide Imaging Both the size and shape of the thyroid gland
The geographic distribution of functional activity in the gland. Functioning thyroid nodules ''hot'' nodules, Non-functioning ones ''cold'' nodules.
The incidence of malignancy in hot nodules is about 1%, but they may become toxic, producing enough hormone to suppress the rest of the gland and induce thyrotoxicosis.

28. Fluorescent Scanning
Advantage of this procedure patient receives no radioisotope gland can be imaged even when it is loaded with iodine ( after IV contrast media)
Disadvantage requires specialized equipment that may not be generally available.

29. Thyroid Ultrasonography
Thyroid size and nodularity much better detail
Useful for measuring the size of the gland or individual nodules
Evaluating the results of therapy.
Differentiating solid from cystic lesions
Guide the operator to a deep nodule during fine- needle thyroid aspiration biopsy.
Limited to thyroid tissue in the neck, ie, it cannot be used for substernal lesions.

30. Magnetic Resonance Imaging
Thyroid size and nodularity much better detail
Excellent image of the thyroid gland, including posterior or substernal extension of a goiter or malignancy .
Both transverse and coronal images
Lymph nodes as small as 1 cm
Invaluable for the demonstration of tracheal compression from a large goiter, tracheal invasion or local extension of a thyroid malignancy , or metastases to local or mediastinal lymph nodes.

31. Fine-Needle Aspiration Biopsy
The best method for differentiation of benign from malignant thyroid disease.
Outpatient procedure
Requires no preparation.
The skin over the nodule is cleansed with alcohol,
If desired, a small amount of 1 % lidocaine can be injected intracutaneously for local anesthesia.

32. Effects of Thyroid Hormones on Peripheral Tissues
A measurement of basal oxygen in the intact organism the basal metabolic rate (BMR).
However, this test is nonspecific and insensitive and is rarely used today
Photomotogram. The traction and relaxation time of the Achilles tendon

33. Algorithm for Diagnosis of Thyroid Dysfunction
TSH
Decreased
Normal
Increased
FreeT4&T3
Euthyroid No further testing
FreeT4
Decreased Nonthyroidal illness or drug effect Secondary Hypothyr.
Normal Subclin hypothyroidism
Increased Hyperthyroidism
Increased Thyroid Resistance or Pituitary Tm.
Normal Subclin hyperthyroidism Glucocort.
Decreased Primary Hypothyroidism