Total Thyroxine (T4), 1 x 96 Assays

FOR RESEARCH USE ONLY.

Product Name: Total Thyroxine (T4)
Sample Type: Serum
No: of Samples: 1X96 Assays
Intended Use: For the quantitative determination of total thyroxine (T4) concentration in human serum.

Principles of the assay: To measure T4 by competitive immunoassay techniques, a sample of serum or plasma containing the T4 to be quantified is mixed with labeled T4 and T4 antibody.6,7 The labeled T4 contains 8-anilino-1-napthalene sulfonic acid (ANS) to inhibit binding of T4 to serum proteins, which would otherwise interfere with the assay. During incubation, a fixed amount of labeled T4 competes with the unlabeled T4 in the sample, standard, or quality control serum for a fixed number of binding sites on the specific T4 antibody.

Separation of the unbound T4 from antibody-bound T4 and the subsequent measurement of the labeled fraction of the bound phase completes the test. By comparing results of the unknown sample with those obtained from a series of T4 calibrators, an accurate measurement of the T4 concentration in the sample can be obtained.
In the GenWay Biotech T4 EIA, antibody to T4 is coated on a solid phase (microtiter well). A measured amount of serum and a constant amount of T4 labeled with horseradish peroxidase are added. During incubation, T4 in the sample and enzyme-labeled T4 compete for the limited binding sites on the T4 antibody. After a 60-minute incubation at room temperature, the solid phase is washed with water to remove unbound-labeled T4. A solution of tetramethylbenzidine (TMB) is added and incubated for 20 minutes, resulting in the development of a blue color. The color development is stopped with the addition of 1N HCI, and the resulting yellow color is measured spectrophotometrically at 450 nm. The intensity of the color formed is proportional to the amount of enzyme present and is inversely related to the amount of T4 in the sample. By reference to a series of calibrators processed in the same way, the concentration of T4 in the unknown sample is determined.

Summary and Explanation of Test:  
Storage and Stability:
1. Store the unopened kit at 2-8C upon receipt and when not in use, until the expiration shown on the kit label. Refer to the package label for the expiration date.
2. Keep microtiter plate in a sealed bag with desiccant to minimize exposure to damp air.

Storage and Stability:
1. Store the unopened kit at 2-8C upon receipt and when not in use, until the expiration shown on the kit label. Refer to the package label for the expiration date.
2. Keep microtiter plate in a sealed bag with desiccant to minimize exposure to damp air.

Limitation of the Test:
1. Reliable and reproducible results will be obtained when the assay procedure is carried out with good laboratory practice and adherence to the package insert instructions.
2. Serum samples with T4 concentrations greater than 25 ?g/dL should be reported as such. If further quantitation is desired, the sample should be diluted with the Zero Standard and re-assayed. The obtained value should then be multiplied by the dilution factor to obtain the true serum value.
3. Icteric samples with bilirubin values as high as 5 mg/dl do not affect the assay. Additionally, added hemoglobin levels of up to 100 mg/dl showed no effect on the T4 value.
4. Total serum T4 values may be influenced by a variety of factors other than thyroid malfunction. High TSH levels, pregnancy, estrogen therapy, oral contraceptives, heparin, phenytoin and propanolol may all produce invalid results.
5. The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbance readings.