DiaMetra FT3 Elisa Kit

PRINCIPLE
The free T3 (FT3, antigen) in the sample competes with the antigenic T3 conjugated with horseradish peroxidase (HRP) for binding to the limited number of antibodies anti T3 coated on the microplate (solid phase) (the enzyme conjugate should have no measurable binding to serum proteins especially TBG and albumin).

CLINICAL SIGNIFICANCE 
The thyroid hormone, triiodothyronine (T3), is produced by the thyroid gland. An important component in the synthesis is iodine. Thyroxine is converted to the active T3 (three to four times more potent than T4) within cells by deiodinases (5’- iodinase).
Thyroxine-binding globulin (TGB) is the major carrier protein for circulating thyroid hormone.
Only a very small fraction of the circulating hormone is free (unbound) 0.3%; this fraction is biologically active. Thus, measurements of free triiodothyronine concentrations correlate more reliably with clinical status than total triiodothyronine levels. For example, the increase in total triiodothyronine levels associated with pregnancy, oral contraceptives and oestrogen therapy result in higher total T3 levels while the free T3 (FT3) concentration remains basically unchanged. The concentrations of the carrier proteins are altered in many clinical conditions, such as pregnancy. In normal thyroid function as the concentrations of the carrier proteins alters, the total triiodothyronine level changes so that the free triiodothyronine concentration remains constant. The binding of T3 plays a key role in the feedback control of the thyroid, with FT3 acting on the pituitary to inhibit thyroid hormone secretion. The thyronines act on the body to increase the basal metabolic rate, affect protein synthesis and increase the body’s sensitivity to catecholamine (such as adrenaline) by permissiveness. The thyroid hormones are essential to proper development and differentiation of all cells of the human body. These hormones also regulate protein, fat, and carbohydrate metabolism, affecting how human cells use energetic compounds. Numerous physiological and pathological stimuli influence thyroid hormone synthesis.
Thyrotoxicosis or hyperthyroidism is the clinical syndrome caused by an excess of circulating free thyroxine, free triiodothyronine, or both.
Both T3 and T4 are used to treat thyroid hormone deficiency (hypothyroidism).

PRECAUTIONS
Please adhere strictly to the sequence of pipetting steps provided in this protocol. The performance data represented here were obtained using specific reagents listed in this Instruction For Use.
All reagents should be stored refrigerated at 2-8°C in their original container. Any exceptions are clearly indicated. The reagents are stable until the expiry date when stored and handled as indicated.
Allow all kit components and specimens to reach room temperature (22-28°C) and mix well prior to use.
Do not interchange kit components from different lots. The expiry date printed on box and vials labels must be observed.. Do not use any kit component beyond their expiry date.
If you use automated equipment, the user has the responsibility to make sure that the kit has been appropriately tested.
The incomplete or inaccurate liquid removal from the wells could influence the assay precision and/or increase the background. To improve the performance of the kit on automatic systems is recommended to increase the number of washes.
It is important that the time of reaction in each well is held constant for reproducible results. Pipetting of samples should not extend beyond ten minutes to avoid assay drift. If more than 10 minutes are needed, follow the same order of dispensation. If more than one plate is used, it is recommended to repeat the dose response curve in each plate.
Addition of the TMB Substrate solution initiates a kinetic reaction, which is terminated by the addition of the Stop Solution. Therefore, the TMB Substrate and the Stop Solution should be added in the same sequence to eliminate any time deviation during the reaction.
Observe the guidelines for performing quality control in medical laboratories by assaying controls and/or pooled sera.

QUALITY CONTROL
Each laboratory should assay controls at levels in the hypothyroid, euthyroid and hyperthyroid range for monitoring assay performance. These controls should be treated as unknowns and values determined in every test procedure performed. Quality control charts should be maintained to follow the performance of the supplied reagents. Pertinent statistical methods should be employed to ascertain trends. The individual laboratory should set acceptable assay performance limits. Other parameters that should be monitored include the 80, 50 and 20% intercepts of the calibration curve for run-to-run reproducibility. In addition, maximum absorbance should be consistent with past experience. Significant deviation from established performance can indicate unnoticed change in experimental conditions or degradation of kit reagents. Fresh reagents should be used to determine the reason for the variations.

Preparation of the Sample
The determination of FT3 should be performed in human serum or plasma.
Specimens may be refrigerated at 2-8°C (for a maximum period of 48 hours). If the specimens cannot be assayed within 48 hours, the samples may be stored at temperatures of –20°C for up to 30 days. Avoid repetitive freezing and thawing of samples. When assayed in duplicate, 0.10 mL of the specimen is required.