Growth Hormone 1 ELISA Kit

Catalog No. ABIN649033

Product Details Growth Hormone 1 ELISA Kit

Target: Growth Hormone 1 (GH1)

Reactivity: Human

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Application: ELISA

Purpose: Immunoenzymometric assay (TYPE 3): The essential reagents required for an immunoenzymometric assay include high affinity and specificity antibodies (enzyme and immobilized), with different and distinct epitope recognition, in excess, and native antigen. 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 monoclonal anti-GH antibody. Monoclonal biotinylated antibody, the enzyme-labeled antibody and a serum containing the native antigen are mixed, reaction results between the native antigen and the antibodies, without competition or steric hindrance, to form a soluble sandwich complex. After equilibrium is attained, the antibodybound fraction is separated from unbound antigen by decantation or aspiration. The enzyme activity in the antibodybound fraction is directly proportional to the native antigen concentration. By utilizing several different serum references of known antigen values, a dose response curve is generated from which the antigen concentration of an unknown is ascertained.

Analytical Method: Quantitative

Components: A. Growth Hormone Calibrators (1ml/vial). Six vials of references for hGH Antigen in human serum at levels of 0 (A), 2 (B), 10 (C), 25 (D), 50 (E) and 150 (F) (IU/ml. Store at 2-8°C. A preservative has been added. (The calibrators were referenced against International Standard WHO 2nd ISNumber 98/574). B. hGH Enzyme Reagent (13 ml/vial). One vial contains horseradish peroxidase (HRP) labeled affinity purified antibody, biotinylated monoclonal mouse IgG in buffer, dye, and preservative. Store at 2-8°C. C. Streptavidin Coated Plate (96 wells). One 96-well microplate coated with streptavidin and packaged in an aluminum bag with a drying agent. Store at 2-8°C. D. Wash Solution Concentrate (20 ml). One vial contains a surfactant in buffered saline. A preservative has been added. Store at 2-8°C. E. Substrate A (7ml/vial). One bottle contains tetramethylbenzidine (TMB) in buffer. Store at 2-8°C. F. Substrate B (7ml/vial). One bottle contains hydrogen peroxide (H2O2) in buffer. Store at 2-8°C. G. Stop Solution (8ml/vial). One bottle contains a strong acid (1N HCl). Store at 2-8°C. H. Product Instructions: Note 1: Do not use reagents beyond the kit expiration date. Note 2: Opened reagents are stable for 60 days when stored at 2-8°C. Note 3: Above reagents are for a single 96-well microplate.

Material not included: 1. Pipette capable of delivering 50µl volumes with a precision of better than 1. 5%. 2. Dispenser(s) for repetitive deliveries of 0. 100ml and 0. 350ml volumes with a precision of better than 1. 5%. 3. Microplate washers or a squeeze bottle (optional). 4. Microplate Reader with 450nm and 620nm wavelength absorbance capability. 5. Absorbent Paper for blotting the microplate wells. 6. Plastic wrap or microplate cover for incubation steps. 7. Vacuum aspirator (optional) for wash steps. 8. Timer. 9. Quality control materials.

Application Details

Application Notes: Precautions: 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 licensed reagents. 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.

Plate: Pre-coated

Reagent Preparation:

1. Wash Buffer: Dilute contents of wash concentrate to 1000ml with distilled or deionized water in a suitable storage container. Store at room temperature 20-27 °C for up to 60 days. 2. Working Substrate Solution: Pour the contents of the amber vial labeled Solution A into the clear vial labeled Solution B. Place the yellow cap on the clear vial for easy identification. Mix and label accordingly. Store at 2 - 8 °C. Note: Do not use the working substrate if it looks blue.

Sample Collection: The specimens shall be blood, serum 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. Allow the blood to clot. Centrifuge the specimen to separate the serum from the cells. Samples may be refrigerated at 2-8°C for a maximum period of five days. If the specimen(s) cannot be assayed within this time, the sample(s) may be stored at temperatures of -20 °C for up to 30 days. Avoid repetitive freezing and thawing. When assayed in duplicate, 0. 100ml of the specimen is required.

Calculation of Results:

A dose response curve is used to ascertain the concentration of growth hormone (hGH) in unknown specimens. 1. Record the absorbance obtained from the printout of the microplate reader. 2. Plot the absorbance for each duplicate serum reference versus the corresponding hGH concentration in (IU/ml on linear graph paper (do not average the duplicates of the serum references before plotting). Draw the best-fit curve through the plotted points. 4. To determine the concentration of hGH of an unknown, locate the average absorbance of the duplicates for each unknown on the vertical axis of the graph, find the intersecting point on the curve, and read the concentration (in (IU/ml) from the horizontal axis of the graph (the duplicates of the unknown may be averaged as indicated).

Restrictions: For Research Use only

Handling

Handling Advice: Before proceeding with the assay, bring all reagents, serum references and controls to room temperature (20-27°C). 1. Format the microplate 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 (50µl) of the appropriate serum reference, control or specimen into the assigned well. Add 0. 100 ml (100µl) of hGH Enzyme Reagent solution to all wells. Swirl the microplate gently for 20-30 seconds to mix and cover. 5. Incubate 60 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 350µl of wash buffer (see Reagent Preparation Section), decant (tap and blot) or aspirate. Repeat two additional times for a total of three washes. An automatic or manual plate washer can be used. Follow the manufacturer's 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 additional times. 8. Add 0. 100 ml (100µl) of working substrate solution to all wells (see Reagent Preparation Section). Always add reagents in the same order to minimize reaction time differences between wells. DO NOT SHAKE THE PLATE AFTER SUBSTRATE ADDITION. 9. Incubate at room temperature for 15 minutes. 10. Add 0. 050ml (50µl) of stop solution to each well and gently mix for 15-20 seconds). Always add reagents in the same order to minimize reaction time differences between wells. 11. Read the absorbance in each well at 450nm (using a reference wavelength of 620-630nm to minimize well imperfections) in a microplate reader. The results should be read within 30 minutes of adding the stop solution.

Storage: 4 °C/-20 °C

Target Details for Growth Hormone 1

Target: Growth Hormone 1 (GH1)

Alternative Name: Growth hormone (Somatotropin) (GH1 Products)

Synonyms: GH ELISA Kit, GH-N ELISA Kit, GHN ELISA Kit, IGHD1B ELISA Kit, hGH-N ELISA Kit, Gh1 ELISA Kit, GH1 ELISA Kit, gh ELISA Kit, Gh ELISA Kit, GH2 ELISA Kit, RNGHGP ELISA Kit, ghl ELISA Kit, gh-n ELISA Kit, ghn ELISA Kit, ighd1b ELISA Kit, gh1 ELISA Kit, ghb-A ELISA Kit, GHI ELISA Kit, GHB3 ELISA Kit, growth hormone 1 ELISA Kit, growth hormone ELISA Kit, somatotropin-like ELISA Kit, somatotropin ELISA Kit, growth hormone prepeptide ELISA Kit, growth hormone 1 L homeolog ELISA Kit, growth hormone 1 S homeolog ELISA Kit, Somatotropin-1 ELISA Kit, GH1 ELISA Kit, Gh ELISA Kit, GH ELISA Kit, LOC100305005 ELISA Kit, Gh1 ELISA Kit, LOC100534452 ELISA Kit, LOC100232594 ELISA Kit, LOC100303681 ELISA Kit, gh1 ELISA Kit, LOC100356068 ELISA Kit, LOC100136588 ELISA Kit, gh1.L ELISA Kit, gh1.S ELISA Kit, LOC109081196 ELISA Kit

Background: Summary and Explanation of the test: Growth hormone (hGH, somatotropin), secreted from the anterior pituitary, is a polypeptide with two intra-chain disulfide bridges, which circulates free or bound to number of different GH-binding proteins. Several forms of growth hormone have been identified (1) with the major being of molecular weight 22,000 daltons containing 191 amino acid residues. A 20,000-dalton variant, which posses all known biological functions of GH, has also been demonstrated to be important. The primary biological actions of the hormone are in direct growth promoting. GH exerts its effect directly on target organs such as bones and muscles and indirectly through the release of somatomedins, a family of insulin-like growth factor (IGF) hormones, produced in the liver (2). In particular, somatotropin C (IGF-1) is essential for bone growth during childhood. The clinical usefulness of the measurement of growth hormone (GH) in children has been well established in ascertaining linear bone growth along the epiphyseal plate. Abnormal elevated levels lead to gigantism while complete absence slows the rate of growth to one-third to one-half of normal. In adults, the epiphyseal growth plates have fused, GH excess gradually produces acromegaly, a coarse thickening of the bones of the skull, hands and feet. In this method, GH calibrator, patient specimen or control is first added to a streptavidin coated well. Biotinylated monoclonal and enzyme labeled antibodies (directed against distinct and different epitopes of GH) are added and the reactants mixed. Reaction between the various GH antibodies and native GH forms a sandwich complex that binds with the streptavidin coated to the well. After the completion of the required incubation period, the enzyme-growth hormone antibody bound conjugate is separated from the unbound enzyme-growth hormone 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 color. The employment of several serum references of known growth hormone levels permits the construction of a dose response curve of activity and concentration. From comparison to the dose response curve, an unknown specimen's activity can be correlated with growth hormone concentration. Intended Use: The Quantitative Determination of Growth Hormone Concentration in Human Serum by a Microplate Immunoenzymometric assay. Q. C. Parameters: In order for the assay results to be considered valid the following criteria should be met: 1. The absorbance (OD) of calibrator A should be lower than 0. 10. 2. The absorbance (OD) of calibrators F should be greater than 1. 3. 3. Four out of six quality control pools should be within the established ranges.

Pathways: NF-kappaB Signaling, JAK-STAT Signaling, Intracellular Steroid Hormone Receptor Signaling Pathway, Peptide Hormone Metabolism, Regulation of Intracellular Steroid Hormone Receptor Signaling, Regulation of Hormone Metabolic Process, Response to Growth Hormone Stimulus, Regulation of Hormone Biosynthetic Process