2. Case 1
• History:
• A 50 year old housewife complains of progressive weight gain
of 20 pounds in 1 year, fatigue, slight memory loss, slow
speech, dry skin, constipation, and cold intolerance.
• Physical examination:
• Vital signs include a temperature 96.8oF, pulse 58/minute and
regular, BP 140/100. She is moderately obese and speaks
slowly and has a puffy face, with pale, cool, dry, and thick
skin. The thyroid gland is slightly enlarged, firm, not nodular,
mobile, and not tender. The deep tendon reflex time is
delayed.
• Laboratory studies:
• CBC and differential WBC are normal. The serum T4
concentration is 3.8 ug/dl (N=4.5-12.5), the serum TSH is 23.0
miU/ml (N=0.2-3.5), and the serum cholesterol is 255 mg/dl.
3. Case 2
• History:
• A 50 year old housewife complains of progressive weight gain
of 20 pounds in 1 year, fatigue, postural dizziness, loss of
memory, slow speech, deepening of her voice, dry skin,
constipation, and cold intolerance.
• Physical examination:
• Vital signs include a temperature 96.8oF, pulse 58/minute and
regular, BP 110/60. She is moderately obese and speaks
slowly and has a puffy face, with pale, cool, dry, and thick
skin. The thyroid gland is not palpable. The deep tendon
reflex time is delayed.
• Laboratory studies:
• CBC and differential WBC are normal. The serum T4
concentration is 3.8 ug/dl (N=4.5-12.5), the serum TSH is 1
miU/ml (N=0.2-3.5), and the serum cholesterol is 255 mg/dl
(N<200).
4. Hypothyroidism
A deficiency in thyroid hormone production or secretion producing a
variety of clinical signs and symptoms of hypometabolism.
Often overlooked may present with serious signs and symptoms.
Treatable with a good prognosis.
Prevalence - undiagnosed hypothyroidism 5%, previously diagnosed
hypothyroidism 3%
Incidence – 226 per 100,000 per year
5. Hypothyroidism
Worldwide, the most common cause of
hypothyroidism is still iodine deficiency.
In developed countries, the most common cause of
primary hypothyroidism is Hashimoto thyroiditis.
Hypothyroidism
• is more common in women than men,
• increases with age
• higher in whites than in blacks or Hispanics
6. Classification
Age of onset
• Congenital
• Acquired
HPT level
• Primary (defect in the thyroid)
• Secondary (defect in the
hypothalamus or pituitary gland, also
called central hypothyroidism)
Severity
• Overt (clinical)
• Mild [subclinical)
Duration
• Permanent
• Transient
7. Primary Hypothyroidism
Thyroid dysgenesis
Destruction of thyroid tissue
Chronic autoimmune thyroiditis: atrophic and goitrous forms
Radiation: 131I therapy for thyrotoxicosis, external radiotherapy to
the head and neck for non-thyroid malignant disease
Subtotal and total thyroidectomy
8. Primary Hypothyroidism
Infiltrative diseases of the thyroid (amyloidosis, sarcoid, lymphoma,
hemochromatosis, scleroderma)
Defective thyroid hormone biosynthesis
Congenital defects in thyroid hormonal biosynthesis
Iodine deficiency
Drugs with antithyroid actions: lithium, iodine and iodine containing
drugs, radiographic contrast agents
9. Central hypothyroidism
Insufficient stimulation by TSH of an
otherwise normal thyroid gland.
Prevalence -
• 1 : 20,000 - 80,000
• 1 in 1000 hypothyroid patients.
• 1 in 160,000 congenital hypothyroidism of
central origin.
10. Central Hypothyroidism
• pituitary macroadenomas, craniopharyngiomas,
meningiomas, gliomas, Rathke cleft cysts,
metastases, carotid aneurysms
Invasive
lesion
• cranial surgery or irradiation, drugsIatrogenic
• head traumas, traumatic deliveryInjury
• postpartum necrosis (Sheehan syndrome),
pituitary apoplexyInfarction
17. Pre-analytical Variables
Age
Pregnancy
TSH/FT4 relationship
Biological differences
Age
• Both TSH and FT4
concentrations are higher in
children, especially in the 1st
week of life and throughout
the 1st year.
• TSH increases with age
• Increased Mean variability
Pregnancy
• ↑ Estrogen levels ↑ 2-3x mean TBG
concentration than pre-pregnancy level by 20
weeks of gestation shift in the TT4 and
TT3 reference range to approximately 1.5
times the nonpregnant level by 16 weeks of
gestation
• ↑ HCG levels cross-react partly with TSH
receptor mildly suppressed levels.
• The peak rise in HCG and nadir in serum TSH
level occurs together at about 10-12 weeks of
gestation.
18. Analytical methods
Gold Standard - Equilibrium dialysis.
• Complex and not widely available.
Commonly used methods - one-step or
two-step immunoassay method.
• Two-step method more reliable than one-step
method.
Chemiluminescence
19. Analytical methods
Analytical interferants - Heterophile
antibodies
• Falsely high or low TFTs.
• Antibodies induced by external antigens
(heterophile antigens) that cross-react with self-
antigens.
• Human anti-mouse antibodies (HAMA) Reacts
with the mouse monoclonal antibodies (used in
many immunometric assays like TSH estimation)
erroneously high or low TSH values
20. Analytical methods
Analytical interferants - Macro-TSH
• Rare condition
• Serum contains antibodies against TSH (anti-TSH
Ig) binds to TSH and neutralizes its activity, but
leaves open epitope to interact with assay
antibodies leading to spuriously high value.
• Detected by:
• Linearity test
• PEG precipitation
• TSH sequestration test
• Gel filtration chromatography
21. Analytical methods
Specimen
• Serum is preferred specimen and ideally whole
blood samples should be allowed to clot for more
than 30 min and then centrifuged and separated.
Storage
• 4-8°C for up to 7 days or −20°C (Long-term).
Collection
• Barrier gel tubes does not affect the results of
TFTs.
22. Analytical methods
Stability
• Quite stable.
• TSH and T4 in dried whole blood spots used to
screen for neonatal hypothyroidism are also
stable for months when stored with a desiccant.
Interferants
• Hemolysis, hyperlipidemia, and
hyperbilirubinemia do not produce interference
in hormone estimation by different assays
23. Measurement of TSH
Immunometric assays with chemiluminescent probes and
solid phase capture Antibodies analytical sensitivity.
Sensitivity is a major issue it is necessary to measure
well below the population reference interval to
differentiate primary hyperthyroidism from other causes
of low serum TSH concentration.
The previously used “generational” concept for TSH
assays is now largely redundant because clinical
guidelines now specify the appropriate sensitivity
required for TSH assays.
24. Measurement of TSH
First-generation assays sensitive enough only to
discriminate normal from hypothyroid subjects
Second-generation assays detect TSH below the
reference interval but not well enough to reliably
discriminate primary hyperthyroidism from other causes
of low TSH.
All assays in clinical practice should be “third generation,”
that is, they should have a coefficient of variation (CV) of
less than 20% (functional sensitivity) at a concentration
of 0.01 mIU/L.
25. Measurement of T3 and T4
Assay Hierarchy
Direct
Equilibrium dialysis
Ultrafiltration
Indirect
Immunoassay
One step
Immunoassay
Two step
Immunoassay
Free Hormone hypothesis
26. Measurement of T3 and T4
fT4
pbT4
Dialysis/Ultrafiltration
Competitive
immunoassay
Mass
Spectrometry
27. Measurement of T3 and T4
Indirect immunoassay methods
• make the assumption that the fT4:tT4 equilibrium is
maintained during immunoassay to an extent sufficient to
return a clinically relevant estimation of fT4.
One-step methods
• incubate the assay antibody and tracer in the presence of all
serum constituents.
Two-step or “back-titration” methods
• allow T4 to equilibrate with the assay antibody in the presence
of all serum components but wash away uncaptured
components before back titrating with tracer.
28. Measurement of T3 and T4
Estimation of total T3 and T4
• Mass spectrometric measurements are now
the method of choice straightforward with
high sensitivity and selectivity.
• Competitive immunoassay
• These methods include a displacing agent
such as 8-anilino-1-napthalene-sulfonic acid to
release thyroid hormone from high-affinity
serum binding sites
31. Inference
• Hypothyroidism is the commonest disorder of thyroid function.
• It is more common in women, and the risk of developing
hypothyroidism increases with age.
• Hypothyroidism is a known risk factor for cardiovascular disease.
• Excluding the newborn period and iodine deficiency, AITD is the
most common cause of primary hypothyroidism.
• Central hypothyroidism (TSH deficiency) is a rare cause of
hypothyroidism.
32. Recent advances
Allen Herndon Dudley syndrome
• Mutation in monocarboxylate transporter-8 gene required
for thyroid hormone transportation into various cells raised
T3, low T4, and normal or elevated TSH
Thyroid receptor-α mutation
• A similar hormonal profile with raised T3, low T4, and normal
TSH Thyroid resistance
Iodotyrosine deiodinase deficiency
• Genetic condition
• have raised T4, normal/low T3, and normal TSH levels.
• Serum and urinary measurement of monoiodothyronine and
diiodothyronine is used to detect
34. Case 1
• History:
• A 50 year old housewife complains of progressive weight gain
of 20 pounds in 1 year, fatigue, slight memory loss, slow
speech, dry skin, constipation, and cold intolerance.
• Physical examination:
• Vital signs include a temperature 96.8oF, pulse 58/minute and
regular, BP 140/100. She is moderately obese and speaks
slowly and has a puffy face, with pale, cool, dry, and thick
skin. The thyroid gland is slightly enlarged, firm, not nodular,
mobile, and not tender. The deep tendon reflex time is
delayed.
• Laboratory studies:
• CBC and differential WBC are normal. The serum T4
concentration is 3.8 ug/dl (N=4.5-12.5), the serum TSH is 23.0
miU/ml (N=0.2-3.5), and the serum cholesterol is 255 mg/dl.
35. Case 2
• History:
• A 50 year old housewife complains of progressive weight gain
of 20 pounds in 1 year, fatigue, postural dizziness, loss of
memory, slow speech, deepening of her voice, dry skin,
constipation, and cold intolerance.
• Physical examination:
• Vital signs include a temperature 96.8oF, pulse 58/minute and
regular, BP 110/60. She is moderately obese and speaks
slowly and has a puffy face, with pale, cool, dry, and thick
skin. The thyroid gland is not palpable. The deep tendon
reflex time is delayed.
• Laboratory studies:
• CBC and differential WBC are normal. The serum T4
concentration is 3.8 ug/dl (N=4.5-12.5), the serum TSH is 1
miU/ml (N=0.2-3.5), and the serum cholesterol is 255 mg/dl
(N<200).
The term myxoedema is used in severe or complicated cases but strictly refers only to the appearance of the skin as it becomes infiltrated with glycosaminoglycans.
Age - both raised and suppressed TSH in elderly have been shown to be associated with increased cardiovascular morbidity
Pregnancy - higher cut-offs for T4, T3 and lower cut-offs for TSH are suggested during pregnancy, which should be standardized in local laboratory
measurement of free thyroid hormone may vary from assay methods. It is advisable to generate normative data for free thyroid hormone in local laboratory with particular assay method
Manufacturers are currently employing various approaches to deal with the HAMA issue with varying degrees of success, including the use of chimeric antibody combinations and blocking agents to neutralize the effects of HAMA on their methods
Manufacturers are currently employing various approaches to deal with the HAMA issue with varying degrees of success, including the use of chimeric antibody combinations and blocking agents to neutralize the effects of HAMA on their methods
It is beholden to the clinical chemist to be aware of and to monitor this aspect of the assay.
It is beholden to the clinical chemist to be aware of and to monitor this aspect of the assay.
specificity of TSH assays is largely of historical concern because modern assays show little cross-reactivity with the other highly homologous pituitary glycoprotein hormones despite sharing a common α-subunit.
whether a wash step is included to remove serum constituents before the addition of the T4 immunoassay tracer. Modern immunoassay methods
are also “analog” because chemically modified T4 probes are used rather than historic radiolabeled hormones.
whether a wash step is included to remove serum constituents before the addition of the T4 immunoassay tracer. Modern immunoassay methods
are also “analog” because chemically modified T4 probes are used rather than historic radiolabeled hormones.
It is beholden to the clinical chemist to be aware of and to monitor this aspect of the assay.
specificity of TSH assays is largely of historical concern because modern assays show little cross-reactivity with the other highly homologous pituitary glycoprotein hormones despite sharing a common α-subunit.
This is less of an issue for tT3 methods owing to the weaker binding of T4 to serum thyroid hormone–binding