Free Triiodothyronine(FT3) Elisa Kit for Diagnostic Use
Immunoassay for the in vitro quantitative determination of free triiodothyronine in human serum.
Triiodothyronine is one of the thyroid hormones present in serum which regulates metabolism. Determination of this hormone concentration is important for the diagnostic differentiation of euthyroid, hyperthyroid, and hypothyroid states. The major fraction of total triiodothyronine is bound to the transport proteins (TBG, prealbumin, albumin). Free triiodothyronine (fT3) is the physiologically active form of the thyroid hormone triiodothyronine (T3). The determination of free T3 has the advantage of being independentof changes in the concentrations and binding properties of thebinding proteins; additional determination of a binding parameter(T-uptake, TBG) is therefore unnecessary. (1, 2, 3)
In normal thyroid function, as the concentrations of the carrier proteins alter, the total T3 level changes so that the FT3 concentration remains constant. (4) Thus, measurements of FT3 concentrations correlate more reliably with clinical status than total T3 levels. For example, the increase in total T3 levels associated with pregnancy, oral contraceptives and estrogen therapy result in higher total T3 levels while the FT3 con- centration remains basically unchanged. (5) In addition, it has been found that the mean FT3 value has a gradient decreasing from young to older. (6)
Competition principle. Total duration of assay:80 minutes.
• Sample, T3 derivant coated microwells and enzyme labeled Anti-T3 are combined.
• During the incubation, T3 derivant coated on microwells and FT3 present in the sample compete for binding to the enzyme labeled antibodies.
• After washing, a complex is generated between the solid phase and enzyme-linked antibodies by immunological reactions.
• Substrate solutionis then added and catalyzed by this complex, resulting in a chromogenic reaction. The resulting chromogenic reaction is measured as absorbance.
• The color intensity is inversely proportional to the amount of FT3 in the sample.
• Coated Microplate, 8 x 12 strips, 96 wells, pre-coated with T3 derivant.
• Calibrators, 6 vials, 1 ml each, ready to use; Concentrations: 0(A), 2(B), 5(C), 10(D), 20(E) and 50(F) pmol/L.
• Enzyme Conjugate, 1 vial, 6.0 ml of HRP(horseradish peroxidase) labeled sheep monoclonal Anti-T3 in Tris-NaCl buffer containing BSA (bovine serum albumin). Contains 0.2% ProClin300 preservative.
• Substrate, 1 vial, 11ml, ready to use, (tetramethylbenzidine) TMB.
• Stop Solution, 1 vial, 6.0 ml of 1 mol/l sulfuric acid.
• Wash Solution Concentrate, 1 vial, 25 ml (40X concentrated), PBS-Tween wash solution.
• IFU, 1 copy.
• Plate Lid: 1 piece.
Materials required (but not provided)
• Microplate reader with 450nm and 620nm wavelength absorbent capability.
• Microplate washer.
• Plate shaker.
• Micropipettes and multichannel micropipettes delivering 50μl with a precision of better than 1.5%.
• Absorbent paper.
• Distilled water
Precautions and warnings
• For in vitro diagnostic use only. For professional use only.
• All products that contain human serum or plasma have been found to be non-reactive for HBsAg, HCV and HIVI/II. But all products should be reared as potential biohazards in use and for disposal.
• Mix the sample in the wells thoroughly by shaking and eliminate the bubbles.
• Conduct the assay away from bad ambient conditions. e.g. ambient air containing high concentration corrosive gas such as sodium hypochlorite acid, alkaline, acetaldehyde and so on, or containing dust.
• Wash the wells completely. Each well must be fully injected with wash solution. The strength of injection, however, is not supposed to be too intense to avoid overflow. In each wash cycle, dry the liquids in each well. Strike the microplate onto absorbent paper to remove residual water droplets. It is recommended to wash the microplate with an automated microplate strip washer.
• Failure to remove adhering solution adequately in the aspiration or decantation wash step(s) may result in poor replication and spurious results.
• Do not use reagents beyond the labeled expiry date.
• Do not mix or use components from kits with different batch codes.
• If more than one plate is used, it is recommended to repeat the calibration curve.
• It is important that the time of reaction in each well is held constant to achieve reproducible results.
• Ensure that the bottom of the plate is clean and dry.
• Ensure that no bubbles are present on the surface of the liquid before reading the plate.
• Thesubstrate and stop solution should be added in the same sequence to eliminate any time deviation during reaction.
• Store at 2-8℃.
• Place unused wells in the zip-lock aluminum foiled pouch and return to 2-8 °C, under which conditions the wells will remain stable for 2 months, or until the labeled expiry date, whichever is earlier.
• Seal and return unused calibrators to 2-8 °C, under which conditions the stability will be retained for 1 month, for longer use, store opened calibrators in aliquots and freeze at -20 °C. Avoid multiple freeze-thaw cycles.
• Seal and return all the other unused reagents to 2-8 °C, under which conditions the stability will be retained for 2 months, or until the labeled expiry date, whichever is earlier.