T3, T4, TSH CLIA Kit

Produktnummer ABIN504750

Das Human T3, T4, TSH ELISA Kit (ABIN504750) ist ein Chemiluminescent ELISA Kit zur Detektion von Human T3, T4, TSH.

Kurzübersicht für T3, T4, TSH CLIA Kit (ABIN504750)

Target: T3, T4, TSH

Reaktivität: Human

Nachweismethode: Chemiluminescent

Methodentyp: Competition ELISA

Applikation: ELISA

Produktdetails

Verwendungszweck: Competitive Enzyme Immunoassay (tT3 and tT4) Type 7 The essential reagents required for a chemiluminescence immunoassay include antibody, enzyme-antigen conjugate, native antigen and a substrate that emits light.. Upon mixing biotinylated antibody, enzyme-antigen conjugate and a serum containing the native antigen, a competition reaction results between the native antigen and the enzyme-antigen conjugate for a limited number of antibody binding sites.

Analytische Methode: Quantitative

Produktmerkmale: The Quantitative Determination of Total Thyroxine, Total Triiodothyronine, Thyroid Stimulating Hormone Concentration for a comprehensive thyroid status of a Human Serum or Plasma sample by a Microplate Chemiluminescence Immunoassay

Anwendungsinformationen

Applikationshinweise: All products that contain human serum have been found to be non-reactive for Hepatitis B Surface Antigen, HIV 1&2 and HCV Antibodies by FDA required tests. Since no known test can offer complete assurance that infectious agents are absent, all human serum products should be handled as potentially hazardous and capable of transmitting disease. Good laboratory procedures for handling blood products can be found in the Center for Disease Control / National Institute of Health, "Biosafety in Microbiological and Biomedical Laboratories," 2nd Edition, 1988, HHS Publication No. (CDC) 88-8395.

Kommentare:

Sample Volume: T3/TSH: 50 µL, T4: 25 µL One Step (Equilibrium)

Plattentyp: Pre-coated

Protokoll:

Specimien Collection and Preparation:

The specimens shall be blood, serum or plasma in type and the usual precautions in the collection of venipuncture samples should be observed. For accurate comparison to established normal values, a fasting morning serum sample should be obtained. The blood should be collected in a plain redtop venipuncture tube without additives or anti-coagulants (for serum) or evacuated tube(s) containing EDTA or heparin.. Allow the blood to clot for serum samples. Centrifuge the specimen to separate the serum or plasma from the cells. Samples may be refrigerated at 2_x001E_8oC for a maximum period of five (5) days. If the specimen(s) cannot be assayed within this time, the sample(s) may be stored at temperatures of _x001E_20oC for up to 30 days. Avoid repetitive freezing and thawing. When assayed in duplicate, 0.05ml of the specimen is required for tT4 and 0.10ml for tT3 and TSH. MATERIALS Required But Not Provided: 1. Pipette capable of delivering 25 and 50l volumes with a precision of better than 1.5%. 2. Dispenser(s) for repetitive deliveries of 0.100ml and 0.300ml volumes with a precision of better than 1.5%. 3. Adjustable volume (20-200l) and (200-1000l) dispenser(s) for conjugate and substrate dilutions. 4. Microplate washer or a squeeze bottle (optional). 5. Microplate Reader with 450nm and 620nm wavelength absorbance capability. 6. Test tubes for dilution of enzyme conjugate and substrate A and B. 7. Absorbent Paper for blotting the microplate wells. 8. Plastic wrap or microplate cover for incubation steps. 9. Vacuum aspirator (optional) for wash steps. 10. Timer. 11. Quality control materials.

Reagent Preparation:

1. Working Reagent A = tT4 or (tT3) - Tracer Solution Dilute the T4-Tracer (or T3 Tracer) 1:11 with s-T3/T4 buffer in a suitable container. For example, dilute 80l of conjugate with 0.8ml of buffer for 16 wells (A slight excess of solution is made). This reagent should be used within twenty-four hours for maximum performance of the assay. Store at 2-8(C. General Formula: Amount of s-T3/T4 Buffer required = Number of wells ( 0.05 Quantity of tT4 Tracer Reagent necessary = # of wells ( 0.005 i.e. = 16 x 0.05 = 0.8ml for s-T3/T4 Buffer 16 x 0.005 = 0.08ml (80l) for tT4 or (tT3) Tracer Reagent _x000C_2. Wash Buffer Dilute contents of Wash Concentrate to 1000ml with distilled or deionized water in a suitable storage container. Store at room temperature until expiration date printed on concentrate label. 3. Working Signal Reagent Solution - Store at 2 - 8(C. Determine the amount of reagent needed and prepare by mixing equal portions of Signal Reagent A and Signal Reagent B in a clean container. For example, add 1 ml of A and 1ml of B per two (2) eight well strips (A slight excess of solution is made). Discard the unused portion if not used within 36 hours after mixing. If complete utilization of the reagents is anticipated, within the above time constraint, pour the contents of Signal Reagent B into Signal Reagent A and label accordingly.

Test Procedure:

Before proceeding with the assay, bring all reagents, serum references and controls to room temperature (20 - 27(C). 1. Format the microplates wells for each serum calibrator, control and patient specimen to be assayed in duplicate. Replace any unused microwell strips back into the aluminum bag, seal and store at 2-8(C. 2. Pipette 0.025 ml (25l) of the appropriate serum reference, control or specimen into the assigned well for tT4. Pipette 0.050ml (50l) for tT3. Pipette 0.050ml (50l) for TSH. 3. Add 0.050 ml (50l) of Working Reagent A, tT4 or tT3 -tracer solution to the appropriate wells (see Reagent Preparation Section). For TSH, add 0.100 of TSH Tracer Reagent and skip steps 4 and 5). 4. Swirl the microplate gently for 20-30 seconds to mix. 5. Add 0.050 ml (50l) of x-T4 or (x-T3) Biotin Reagent solution to the appropriate wells. 6. Swirl the microplate gently for 20-30 seconds to mix and cover. 7. Incubate 45 minutes at room temperature. 8. Discard the contents of the microplate by decantation or aspiration. If decanting, blot the plate dry with absorbent paper. 9. Add 300l of wash buffer (see Reagent Preparation Section), decant (tap and blot) or aspirate. Repeat four (4) additional times for a total of five (5) washes. An automatic or manual plate washer can be used. Follow the manufacturers instruction for proper usage. If a squeeze bottle is employed, fill each well by depressing the container (avoiding air bubbles) to dispense the wash. Decant the wash and repeat two (2) additional times. 10. Add 0.100ml (100l) of working signal reagent solution to all wells (see Reagent Preparation Section). Always add reagents in the same order to minimize reaction time differences between wells. 11. Incubate for five (5) minutes at room temperature in the dark. 12. Read the RLUs (Relative Light Units) in each well in a microplate luminometer for at least 0.2 seconds/ well. The results can be read within 30 minutes of adding the substrate solution. Note: For reassaying specimens with concentrations greater than highest calibrator, dilute 12.5l (tT4) or 25l (tT3-TSH) of the specimen and 12.5l (tT4) or 25l (tT3-TSH) of the 0 serum reference into the sample well (this maintains a uniform protein con_x001F_centration). Multiply the readout value by 2 to obtain the thyroxine concentration.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Antigendetails

Target: T3, T4, TSH

Andere Bezeichnung: T3, T4 & TSH

Hintergrund: Measurements of thyroid hormones (tT3, tT4 and TSH) are generally re_x001F_garded as invaluable in-vitro diagnostic tests for assessing thyroid function. This importance has provided the impetus for the significant improvement in assay methodology that has occurred in the last three decades. This procedural evolution can be traced from the empirical protein bound iodine (PBI) test (1) to the theoretically sophisticated radioimmunoassay (2) and currently used EIA, ELISA, FIA and Chemiluminescence. The Combination Thyroid Panel (CTP) provides the convenience of combination calibrators, universal plate and flexible reagent selection allowing technicians to perform a variety of assay designs. In this method, serum reference, patient specimen, or control is first added to a microplate well. Enzyme-tT4 (tT3) conjugate and biotinylated tT4 or tT3 antibody are added, and the reactants are mixed. In the case of TSH, the biotinylated and enzyme conjugate are added in one step. .A reaction results between the enzyme conjugate, biotinylated conjugate and the native thyroid hormone (tT3, tT4 or TSH) for the antibody combining sites. Immobilization takes place through the reaction of the incorporated biotin and streptavidin coated on the well. After the completion of the required incubation period, the bound enzyme conjugate is separated from the unbound enzyme conjugate by aspiration or decantation. The activity of the enzyme present on the surface of the well is quantitated by reaction with a suitable substrate to produce light. The employment of several serum references of known thyroid hormone concentration (s) permits construction of a graph of activity and con_x001F_centration. From comparison to the dose response curve (s), an unknown specimen's activity can be correlated with hormone concentration.