预期应用
ELISA法定量测定大鼠血清、血浆或其它相关液体中FT4含量。
实验原理
本试剂盒应用双抗体夹心酶标免疫分析法测定标本中FT4水平。用纯化的抗体包被微孔板，制成固相抗体，往包被单抗的微孔中依次加入FT4抗原、生物素化的抗大鼠FT4抗体、HRP标记的亲和素，经过彻底洗涤后用底物TMB显色。TMB在过氧化物酶的催化下转化成蓝色，并在酸的作用下转化成最终的黄色。颜色的深浅和样品中的FT4呈正相关。用酶标仪在450nm波长下测定吸光度（OD值），计算样品浓度。
Intended use
This immunoassay kit allows for the specific measurement of rat FT4 concentrations in serum and plasma.
Introduction
L-Thyroxine (T4) or 3,5,3’,5’-tetraiodothyronine is the most commonly measured thyroid hormone for the diagnosis of thyroid function. T4 has its primary infuence on protein synthesis and oxygen consumption in virtually all tissues but it is also important for growth, development, and sexual maturation. T4 is synthesized by the thyroid gland and is secreted into the bloodstream. Here the T4 becomes bound to serum proteins for transport to the cells. The major transport protein is Thyroxine Binding Globulin (TBG) which normally accounts for 80% of the bound T4. Other thyroid hormone binding proteins are Thyroxine Binding Prealbumin and Albumin. Most of the serum T4 is bound to these transport proteins leaving only about 0.03% free to exert its effect on cells. It is the free T4 (fT4) that represents the metabolically active fraction; for this reason the measurement of fT4 concentration is considered to be an indicator of patient thyroid status. Primary hypothyroidism results in underproduction of T4 by the thyroid gland and consequently an abnormally low circulating fT4 concentration in the blood. Primary hyperthyroidism leads to excessive thyroid production on T4 and resulting elevated fT4 concentration.
Total serum T4 concentrations are dependent on the level of circulating TBG as well as the patient’s thyroid status. The concentration of TBG can be affected by certain drugs, steriod hormones, pregnancy, and by various nonthyroid illnesses. In an earlier generation of thyroid function tests, the effect of variable TBG concentration was dealt with by calculating a Free Thyroxine Index (FTI). This FTI is the product of Total T4 concentration and Thyroid Uptake (TU), which assesses the number of available binding sites on the TBG. This approach requires carrying out two separate assay determinations (total T4 and TU), but does provide a better indicator of thyroid status than total T4 alone.
Test principle
The essential reagents required for a solid phase enzyme immunoassay include immobilized T4 antibody, enzyme-T4 conjugate and native T4 antigen. The enzyme-T4 conjugate should have no measurable binding to serum proteins especially TBG and albumin. The method achieves this goal. Upon mixing immobilized antibody, enzyme-T4 conjugate and a serum containing the native free T4 antigen, a competition reaction results between the native free T4 and the enzyme-T4 conjugate for a limited number of immobilized binding sites. After equilibrium is attained, the antibody-bound fraction is separated from unbound antigen by decantation or aspiration. The enzyme activity in the antibody-bound fraction is inversely proportional to the native free antigen concentration. By utilizing several different serum references of known antigen concentration, a dose response curve can be generated from which the antigen concentration of an unknown can be ascertained.
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