Thyroid-stimulating hormone (TSH or thyrotropin) is a glycopolipeptide hormone synthesized and secreted by the anterior pituitary gland which regulates the endocrine function of the thyroid gland. TSH consists of two subunits, the alpha and the beta subunit. The a subunit is identical to that of human chorionic gonadotropin (HCG), luteinising hormone (LH), follicle-stimulating hormone (FSH). The β  subunit is unique to TSH, and therefore determines its function.

TSH production is controlled by a Thyrotropin Releasing Hormone, (TRH), which is manufactured in the hypothalamus and transported to the pituitary gland, where it increases TSH production and release. Somatostatin is also produced by the hypothalamus, and has an opposite effect on the pituitary production of TSH, decreasing or inhibiting its release. TSH stimulates the thyroid gland to secrete the hormones thyroxine (T4) and triiodothyronine (T3). Release of TSH is regulated by the circulating free fraction of thyroid hormones in the blood.

TSH levels are depressed when peripheral concentrations of the free fraction of thyroid hormones are high. Conversely, TSH levels are high when peripheral concentrations of thyroid hormones are low. This effect creates a regulatory negative feedback loop. TSH levels are tested in the blood of patients suspected of suffering from excess (hyperthyroidism), or deficiency (hypothyroidism) of thyroid hormone. Generally, a normal range for TSH is between 0.3 and 3.0 mIU/mL, but the interpretation depends also on what the blood levels of thyroid hormones (T3 and T4) are. Higher than normal levels of TSH combined with high levels of thyroid hormone (T3 and T4) may indicate dysfunction of the hypothalamus and pituitary gland. In these case, a high TSH is often produced by a benign tumour of the pituitary (adenoma). Conversely, low levels of TSH, while blood levels of T3 and T4 are also low, indicates abnormally low function of the pituitary, known as hypopituitarism. On the other hand, due to the negative feedback described above, abnormally high levels of Thyroid hormone, due to overproduction in the thyroid, results in low TSH levels. This occurs in diseases such as hyperthyroidism or Grave?s disease.

Conversely, an underproduction of T3 and T4 caused by diseases such as congenital hypothyroidism (cretinism), hypothyroidism or thyroid hormone resistance, gives rise to an increase in the measured TSH. Clearly both TSH and T3 and T4 should be measured to ascertain where a specific thyroid disfunction is caused by primary pituitary or by a primary thyroid disease.

May be diagnosed by:

• Determination of specific antibodies based on the ELISA technique
• Determination of specific antibodies based on the CLIA-technique

TSH ELISA:

Immunoenzymatic colorimetric method (ELISA) for quantitative determination of TSH in serum or plasma.

Principle of the Assay:

In this method, TSH calibrators, patient specimens and/or controls containing the native antigen are first added to streptavidin coated wells. Biotinylated monoclonal and horseradish peroxidase (HRP) labelled antibodies are added and the reactants are mixed. The different types of antibodies used have high affinity and specificity and are directed against distinct and different epitopes of TSH. Reaction between the various TSH antibodies and native TSH occurs in the microwells without competition or sterics hindrance forming a soluble sandwich complex.
The interaction is illustrated by the following equation:

BtnAb(m) = Byotinilated Monoclonal Antibody (Excess Quantity)
AgTSH = Native Antigen (Variable Quantity)
HRP-Ab(p) = HRP labeled Antibody (Excess Quantity)
HRP-Ab(p)-AgTSH-BtnAb(m) = Antigen-Antibodies Sandwich Complex
Ka = Rate Constant of Association
K-a = Rate Constant of Dissociation

Simultaneously, the complex is fixed to the well through the high affinity reaction of streptavidin and biotinylated antibody. This interaction is illustrated below:

Streptavidin CW = Streptavidin immobolized on well.
Immobilized Complex = Antibodies-Antigen sandwich bound.

After equilibrium is attained, the antibody-bound fraction is separated from unbound antigen by aspiration. The native antigen concentration is directly proportional to the HRP activity in the antibody-bound fraction. The activity of the conjugated HRP is quantitated by reaction with TMB substrate to produce blue colour. The reaction is terminated by adding stop solution which turns the blue colour into yellow. The absorbance is measured on a plate reader.

Specific performance characteristics: 

Sensitivity:
On the basis of results of 16 replicate determinations of standard 0, the minimum TSH concentration detectable by the present method is 0.078 mIU/l, for 1 hr incubation, and 0.027 mIU/l, for 2 hr incubation. The detection limit is defined as the value deviating by 2 SD from that of standard 0. 

Order information:

Correlation with CLIA performed on 57 samples is r = 0.993