Anti-Thyroid-Globulin Antibody (TGAB) CLIA Kit

Produktnummer ABIN504741

Produktdetails

Target: Anti-Thyroid-Globulin Antibody (TGAB)

Reaktivität: Human

Nachweismethode: Chemiluminescent

Methodentyp: Sandwich ELISA

Applikation: ELISA

Verwendungszweck: A Sequential CLIA Method The reagents required for the sequential CLIA assay include immobilized antigen, circulating auto-antibody and enzyme-linked species specific antibody. In this procedure, the immobilization takes place during the assay at the surface of a microplate well through the interaction of streptavidin coated on the well and exogenously added biotinylated thyroglobulin antigen. Upon mixing biotinylated antigen and a serum containing the auto-antibody, reaction results between the antigen and the antibody to form an immune-complex.

Analytische Methode: Quantitative

Produktmerkmale: The Quantitative Determination of Thyroglobulin (Tg) Autoantibodies in Human Serum or Plasma by a Microplate Chemiluminescence Immunoassay (CLIA). Measurements of Tg autoantibodies may aid in the diagnosis of certain thyroid diseases such as Hashimotos and Graves as well as nontoxic goiter.

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.

Probenmenge: 50 μL

Plattentyp: Pre-coated

Protokoll:

Specimien Collection and Preparation:

Collect sample(s) by venipuncture in ten (10) ml silicone evacuated tube(s) or evacuated tube(s) containing EDTA or heparin. The usual precautions in the collection of venipuncture samples should be observed. Separate the red blood cells by centrifugation use serum or plasma for the anti-Tg procedure. Specimen(s) may be refrigerated at 2_x001E_8(C for a maximum period of 48 hours. If the specimen(s) can not be assayed within 48 hours, the sample(s) may be stored at temperatures of _x001E_20(C for up to 30 days. When assayed in duplicate, 0.100ml of the diluted specimen is required.

Reagent Preparation:

1. Serum Diluent Dilute the serum diluent concentrate to 200ml in a suitable container with distilled or deionized water. Store at 2-8(C. 2. Wash Buffer Dilute contents of Wash Concentrate to 1000ml with distilled or deionized water in a suitable storage container. Store diluted buffer at room temperature 20-27(C. 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. 4. Patient Sample Dilution (1/100) Dispense 0.010ml (10l) of each patient specimen into 1ml of serum diluent. Cover and vortex or mix thoroughly by inversion. Store at 2-8(C for up to forty-eight (48) hours

Test Procedure:

Before proceeding with the assay, bring all reagents, serum references and controls to room temperature (20-27C). 1. Format the microplates wells for each serum reference, 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 0.050 ml (50l) of the appropriate serum reference, control or diluted patient specimen into the assigned well. 3. Add 0.100 ml (100l) of Tg Biotinylated Conjugate Solution. 4. Swirl the microplate gently for 20-30 seconds to mix and cover. 5. Incubate 30 minutes at room temperature. 6. Discard the contents of the microplate by decantation or aspiration. If decanting, blot the plate dry with absorbent paper. 7. Add 350l 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 four (4) additional times. 8. Add 0.100 ml (100l) of anti-Tg Tracer Reagent to all wells. Always add reagents in the same order to minimize reaction time differences between wells. 9. Incubate for thirty (30) minutes at room temperature. 10. Discard the contents of the microplate by decantation or aspiration. If decanting, blot the plate dry with absorbent paper. 11. Add 350l 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 four (4) additional times.. Add 0.100 ml (100l) of working signal reagent to all wells (see Reagent Preparation Section). Always add reagents in the same order to minimize reaction time differences between wells. 13. Incubate at room temperature for five (5) minutes in the dark. 14. 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 re-assaying specimens with concentrations greater than 2000 IU/ml, dilute the sample an additional 1:5 or 1:10 using the original diluted material. Multiply by the dilution factor to obtain the concentration of the specimen.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Antigendetails

Target: Anti-Thyroid-Globulin Antibody (TGAB)

Andere Bezeichnung: Antibodies to thyroglobulin (Tg) (TGAB Produkte)

Synonyme: AITD3 CLIA Kit, TGN CLIA Kit, thyroglobulin CLIA Kit, TG CLIA Kit

Substanzklasse: Antibody

Hintergrund: Antibodies to thyroglobulin have been shown to be characteristically present from patients with thyroiditis and primary thyrotoxicosis1, 2. This has lead to the clinical measurement becoming a valuable tool in the diagnosis of thyroid dysfunction. Passive Hemaglutination (PHA) methods have been employed in the past for measurements of antibodies to Tg. PHA tests do not have the sensitivity of enzyme immunoassay and are limited by subjective interpretation. This procedure, with the enhanced sensitivity of chemiluminescence, permits the detectability of subclinical levels of antibodies to Tg. In addition, the results are quantitated by a luminometer, which eliminates subjective interpretation. Autoantibodies to Tg are often present in patients with autoimmune thyroid disease. Approximately 10 percent of healthy individuals have autoantibodies to Tg at low levels, higher concentrations are found in 30% and 85% of patients with Graves disease and Hashimotos Thyroiditis respectively. Antibodies to thyroid peroxidase (TPO) occur more frequently than autoantibodies to Tg in these conditions thus rendering anti-Tg assays without any practical use. However, anti-Tg assays are useful while determining Tg levels in patients with thyroid conditions. The presense of Tg autoantibodies produces false results in determination of Tg levels both by competitive assays and by sandwitch immunoassays. Monobind's microplate chemiluminescence immunoassay methodology provides the technician with optimum sensitivity while requiring few technical manipulations. In this method, serum reference, diluted patient specimen, or control is first added to a microplate well. Biotinylated thyroglobulin (Tg) is added, then the reactants are mixed. Reaction results between the auto-antibodies to Tg and the biotinylated Tg to form an immune complex, which is deposited to the surface of streptavidin coated wells through the high affinity reaction of biotin and streptavidin. After the completion of the required incubation period, aspiration or decantation separates the reactants that are not attached to the wells. An anti-human IgG- enzyme conjugate is then added to permit quantitation of reaction through interacting with human IgG of the immune complex. After washing, the enzyme activity is determined by reaction with substrate to produce light (luminescence). The employment of several serum references of known antibody activity permits construction of a dose response curve (graph) of enzyme and antibody activities. From comparison to the dose response curve, an unknown specimen's enzyme activity can be correlated with auto-immune antibody level. The intensity of light is directly proportional to the concentration of anti-thyroglobulin antibody in the specimen.