Insulin Kit ELISA

N° du produit ABIN2014348

Kit ELISA Humain Insulin, test Colorimetric pour la quantification de Humain Insulin.

Aperçu rapide pour Insulin Kit ELISA (ABIN2014348)

Antigène: Insulin (INS)

Reactivité: Humain

Méthode de détection: Colorimetric

Type de méthode: Sandwich ELISA

Gamme de detection: 0.1817-138 mU/L

Application: ELISA

Type d'échantillon: Serum, Plasma

Détail du produit Insulin Kit ELISA

Seuil minimal de détection: 0.1817 mU/L

Fonction: This test kit is intended for use in the quantitative determination of human Insulin in serum or plasma.

Analytical Method: Quantitative

Specificité: This assay measures human Insulin without any cross-reaction to C-peptide.

Réactivité croisée (Details): This assay measures human Insulin without any cross-reaction to C-peptide.

Ingrédients: 1. Anti-human Insulin Antibody Coated Microplate
One bottle containing 12 mL ready-to-use buffer. It should be used only for tracer antibody dilution according to the assay procedures. This reagent should be stored at 2-8 °C and is stable until the expiration date on the kit box.

Matériel non inclus: 1. Precision single channel pipettes capable of delivering 100 µL.
2. Disposable pipette tips suitable for above volume dispensing.
3. Aluminum foil.
4. Deionized or distilled water.
5. Plastic microtiter well cover or polyethylene film.
6. ELISA multichannel wash bottle or automatic (semi-automatic) washing system.
7. Spectrophotometric microplate reader capable of reading absorbance at 450/650 or 450/620 nm.

Information d'application

Volume d'échantillon: 50 μL

Durée du test: 4 h

Plaque: Pre-coated

Protocole: This ELISA kit is designed, developed and produced for the quantitative measurement of human Insulin in serum and /or EDTA-plasma samples. The assay utilizes the sandwich technique with selected antibodies that bind to various epitopes of Insulin.Assay standards, controls and patient samples are added directly to wells of a microplate that is coated with an anti-human Insulin specific antibody. Simultaneously, a horseradish peroxidase-conjugated monoclonal Insulin specific antibody is added to each well. After the first incubation period, the antibody on the wall of microtiter well captures human Insulin in the sample and unbound proteins in each microtiter well are washed away. A sandwich of anti-Insulin antibody --- human Insulin --- HRP conjugated tracer antibody is formed. The unbound tracer antibody is removed in the subsequent washing step. For the detection of this immunocomplex, the well is then incubated with a substrate solution in a timed reaction and then measured in a spectrophotometric microplate reader. The enzymatic activity of the immunocomplex bound to human Insulin on the wall of the microtiter well is directly proportional to the amount of Insulin in the sample. A standard curve is generated by plotting the absorbance versus the respective human Insulin concentration for each standard on point-to-point or 4 parameter curve fit. The concentration of human Insuloin in test samples is determined directly from this standard curve.

Préparation des réactifs:

(1) Prior to use allow all reagents to come to room temperature. Reagents from different kit lot numbers should not be combined or interchanged.
(2) ELISA Wash Concentrate must be diluted to working solution prior to use. Please see REAGENTS section for details.
(3) Reconstitute assay standards and controls by adding 0.5 mL of deminerialized water to each standard and control bottle. Allow the standards and controls to sit undisturbed for 5 minutes, then mix well by inversions or gentle vortexing. Make sure that all solid is dissolved completely prior to use. These reconstituted standards and controls may be stored at 2- 8 C for up to 3 days or below ?20 C for long-term storage. Do not exceed 3 freeze-thaw cycles.
(4) Prepare Tracer Antibody working solution by 1:21 fold dilution of the Insulin Tracer Antibody by adding the tracer antibody into the Tracer Antibody Diluent . Following is a table that outlines the relationship of strips used and antibody mixture prepared. NOTE: the tracer antibody should be prepared just prior to the end of the first incubation cycle.
(5) Test Configuration
(6) Place a sufficient number of Anti-human Insulin antibody-coated microwell strips in a holder to run human Insulun standards, controls and unknown samples in duplicates.

Prélèvement de l'échantillon: Serum, EDTA-plasma, and urine samples are suitable specimens for human Insulin measurement. Only 50 µL of human sample is required for a duplicate determination of human Insulin with this test kit. No special preparation of the individual is necessary prior to specimen collection. Samples should be collected by standard technologies of the clinical laboratory practices and recommended by the manufacturer of sample collection tube. It is extremely important to carefully separate the serum and plasma from blood cells to avoid hemolyzation, etc. Serum/EDTA-plasma should be transferred to a clean test tube right after centrifugation. Human samples should be stored at 2 ? 8 C if the assay is to be performed within 72 hours. Otherwise, patient samples should be stored at -20 °C or below until measurement. Avoid more than three times freeze-thaw cycles of specimen. Do not use hemolyzed, hyperlipermic, heat-treated or any contaminated specimens.

Procédure de l'essai:

(1) Add 25 µL of Standards, Controls and patient samples into the designated microwells.
(2) Add 100 µL of the above diluted Tracer Antibody working solution to each well.
(3) Seal the plate wells securely, cover with foil or similar material to protect from light. Incubate the plate shaking, at room temperature for 1 hr. 5 minutes.
(4) Wash each well 5 times by dispensing 350 µL of working wash solution into each well and then completely aspirating the contents. Alternatively, an automated microplate washer can be used.
(5) Add 100 µL of ELISA HRP Substrate into each of the wells.
(6) Cover the plate with aluminum foil or similar material to avoid exposure to light. Incubate the plate static, at room temperature for 20 minutes.
(7) Immediately add 100 µL of ELISA Stop Solution into each of the wells. Mix gently.
(8) Read the absorbance at 450 nm with reference filter at 620 nm or 650 nm

Calcul des résultats:

It is recommended to use a point-to-point or 4-parameter standard curve fitting.
1. Calculate the average absorbance for each pair of duplicate test results.
2. Subtract the average absorbance of the level 1 standard (0 ng/mL) from the average absorbance of all other readings to obtain corrected absorbance.
3. The standard curve is generated by the corrected absorbance of all standard levels on the ordinate against the standard concentration on the abscissa using point-to-point or log-log paper. Appropriate computer assisted data reduction programs may also be used for the calculation of results. The human Insulin concentrations for the controls and the patient samples are read directly from the standard curve using their respective corrected absorbance.

Précision du teste: The intra-assay precision was validated by measuring three control samples with 16 replicate determinations. The inter-assay precision was validated by measuring two control levels in duplicate in 9 individual assays.

Restrictions: For Research Use only

Stockage

Précaution d'utilisation: The reagents must be used in professional laboratory. Source material for reagents containing bovine serum was derived in the contiguous 48 United States. It was obtained only from healthy donor animals maintained under veterinary supervision and found free of contagious diseases. Wear gloves while performing this assay and handle these reagents as if they are potentially infectious. Avoid contact with reagents containing TMB, hydrogen peroxide, or sulfuric acid. TMB may cause irritation to skin and mucous membranes and cause an allergic skin reaction. TMB is a suspected carcinogen. Sulfuric acid may cause severe irritation on contact with skin. Do not get in eyes, on skin, or on clothing. Do not ingest or inhale fumes. On contact, flush with copious amounts of water for at least 15 minutes. Use Good Laboratory Practices.

Stock: 4 °C

Détail du antigène

Antigène: Insulin (INS)

Autre désignation: Insulin

Sujet: Insulin is a peptide hormone, produced by beta cells of the pancreas. Enzymatic cleavage of proinsulin results in the production of equimolar amounts of insulin and C-peptide in circulation. Insulin is central to regulating carbohydrate and fat metabolism in the body. Excessive amounts of insulin are associated with excess amounts of glucose in the system. High levels of insulin are known to cause weight gain, water bloating, high blood pressure, magnesium deficiency and an increase in the amount of inflammatory compounds in the blood, which causes blood clots and blood vessel damage.Insulin, when insufficiently produced, results in diabetes mellitus.In most cases, a high fasting insulin level is consistent with insulin resistance symptoms, but in some cases, it can be caused by more serious conditions such as Cushing's syndrome, acromegaly or possibly insulinoma.

Pathways: Signalisation NF-kappaB, Signalisation RTK, Positive Regulation of Peptide Hormone Secretion, Peptide Hormone Metabolism, Hormone Activity, Carbohydrate Homeostasis, ER-Nucleus Signaling, Regulation of Carbohydrate Metabolic Process, Feeding Behaviour, Autophagy, Negative Regulation of intrinsic apoptotic Signaling, Brown Fat Cell Differentiation, Positive Regulation of fat Cell Differentiation