Chromogranin A ELISA Kit

Produktnummer ABIN1305164

Produktdetails Chromogranin A ELISA Kit

Target: Chromogranin A (CHGA)

Reaktivität: Human

Nachweismethode: Colorimetric

Methodentyp: Sandwich ELISA

Detektionsbereich: 5.0-830 ng/mL

Untere Nachweisgrenze: 5 ng/mL

Applikation: ELISA

Verwendungszweck: This ELISA (enzyme-linked immunosorbent assay) kit is intended for the quantitative determination of human chromogranin A levels in EDTA-plasma samples. This assay exclusively measures human chromogranin A without the high dose hook effect up to 1,000,000 ng/ml. This test may be used as an aid for detecting patients with pheochromocytoma and neuroendocrine tumors (carcinoids).

Marke: ED™

Proben: Plasma

Analytische Methode: Quantitative

Bestandteile: 1. Anti-CgA Antibody Coated Microplate
Two vials each containing human chromogranin A in a lyophilized bovine serum-based matrix with a non-azide, non-mercury preservative. Refer to vials for exact concentration range for each control.
Both controls should be stored at 2-8 °C and are stable until the expiration date on the kit box.

Benötigtes Material: 1. Precision single channel pipettes capable of delivering 25 µL, 50 µL, 100 µL, and 1000 µL etc.
2. Repeating dispenser suitable for delivering 100 µL.
3. Disposable pipette tips suitable for above volume dispensing.
4. Disposable 12 x 75 mm or 13 x 100 glass or plastic tubes.
5. Disposable plastic 100 mL and 1000 mL bottle with caps.
6. Aluminum foil.
7. Deionized or distilled water.
8. Plastic microtiter well cover or polyethylene film.
9. ELISA multichannel wash bottle or automatic (semi-automatic) washing system.
10. Spectrophotometric microplate reader capable of reading absorbance at 450 nm.

Anwendungsinformationen

Probenmenge: 50 μL

Testdauer: 4 h

Plattentyp: Pre-coated

Protokoll: Assay standards, controls and patient samples are added directly to wells of microplate that is coated with a polyclonal chromogranin A antibody. After the first incubation period, the antibody on the wall of microtiter well captures human chromogranin A in the sample and unbound protein in each microtiter well is washed away. Then a horseradish peroxidase (HRP)-labeled monoclonal anti-human chromogranin A antibody is added to each microtiter well and a sandwich of monoclonal antibody human chromogranin A polyclonal antibody is formed. The unbound monoclonal antibody is removed in the subsequent washing step. For the detection of this immunocomplex, the well is 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 the chromogranin A on the wall of the microtiter well is directly proportional to the amount of chromogranin A in the sample. A standard curve is generated by plotting the absorbance versus the respective human chromogranin A concentration for each standard on point-to-point or cubical scales or 4 parameter curve fits. The concentration of human chromogranin A in test samples is determined directly from this standard curve.

Aufbereitung der Reagenzien:

(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 use. Please see REAGENTS section for details.
(3) Reconstitute all assay standards and controls by adding 0.5 mL of deminerialized water to each vial. Allow the standards and controls to sit undisturbed for 10 minutes, and then mix well by inversions or gentle vortexing. Make sure that all solids are dissolved completely prior to use. These reconstituted standards and controls must be stored at - 10 C or below. Do not exceed 3 freeze-thaw cycles.

Probennahme: Only 50 µL of human EDTA-plasma is required for human chromogranin A measurement in duplicate. No special preparation of individual is necessary prior to specimen collection. Whole blood should be collected with lavender-top Vaccutainer. Separate the plasma from cells by centrifugation (850 ? 1500xg for 10 minutes). The plasma should be separated from the cells within one hour of blood collection and transferred to a clean test tube. Plasma samples should be stored at -20 °C if the assay is not to be performed within 72 hours. Otherwise, the plasma samples should be stored at room temperature for up to 72 hours. It is important that the plasma samples must not be stored at 2 ? 8 C in any circumstance. Avoid more than three freeze-thaw cycles of specimen. Serum sample can also be used for chromogranin A measurement. However, intact chromogranin A molecule seems much more unstable in serum than in EDTA-plasma. Therefore, we strongly recommend using EDTA-plasma.

Testdurchführung:

(1) Place a sufficient number of antibody coated microwell strips in a holder to run human chromogranin A standards, controls and unknown samples in duplicate.
(2) Test Configuration
(3) Add 25 µL of standards, controls and patient samples into the designated microwells.
(4) Add 100 µL of assay buffer to each well.
(5) Cover the plate with one plate sealer and also with aluminum foil, and incubate plate with orbital shaking 170 rpm or 450 rpm on an ELISA plate shaker at room temperature for 2 hours.
(6) Prepare Chromogranin A Tracer antibody working solution by 1:21 fold dilution of the antibody (30153) with the Tracer Antibody Diluent (30017). For each strip, mix 1 mL of the Tracer Antibody Diluent with 50 µL of the antibody in a clean test tube.
(7) Remove aluminum foil and plate sealer. Aspirate the contents of each well. 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.
(8) Add 100 µL of above diluted Chromogrnin A Tracer Antibody working solution to each of the wells.
(9) Cover the plate with the plate sealer and also with aluminum foil, and incubate plate with orbital shaking 170 rpm or 450 rpm on an ELISA plate shaker at room temperature for 1 hour.
(10) Remove aluminum foil and plate sealer. Aspirate the contents of each well. 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.
(11) Add 100 µL of ELISA HRP Substrate into each of the wells.
(12) Cover the plate with one plate sealer and also with aluminum foil to avoid exposure to light. (13) Incubate plate at room temperature for 20 minutes. (14) Remove the aluminum foil and plate sealer. Add 100 µL of ELISA Stop Solution into each of the wells. Mix gently. (15) Read the absorbance at 450 nm within 10 minutes in a microplate reader. NOTE: to reduce the background, one can set the instrument to dual wavelength measurement at 450 nm with background wavelength correction set at 595 nm, 620 nm or 630 nm.

Ergebnisberechnung:

1. Calculate the average absorbance for each pair of duplicate test results.
   2. Subtract the average absorbance of the STD 1 (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 chromogranin A concentrations for the controls and patient samples are read directly from the standard curve using their respective corrected absorbance

Testpräzision: The intra-assay precision is validated by measuring two controls samples in a single assay with 20 replicate determinations. The inter-assay precision is validated by measuring two control samples in duplicate in 12 individual assays.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Vorsichtsmaßnahmen: The reagents must be used in a professional laboratory environment and are for in vitro diagnostic use. Source material which contains reagents of bovine serum albumin 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 were 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.

Lagerung: 4 °C

Referenzen für Chromogranin A ELISA Kit (ABIN1305164)

Lackermair, Schüttler, Kellnar, Schuhmann, Weckbach, Brunner: "Combined effect of acute altitude exposure and vigorous exercise on platelet activation." in: Physiological research, Vol. 71, Issue 1, pp. 171-175, (2022) (PubMed).

Details zu Chromogranin A

Target: Chromogranin A (CHGA)

Andere Bezeichnung: Chromogranin A (CHGA Produkte)

Synonyme: CGA ELISA Kit, CG-ALPHA ELISA Kit, FSHA ELISA Kit, GPHA1 ELISA Kit, GPHa ELISA Kit, HCG ELISA Kit, LHA ELISA Kit, TSHA ELISA Kit, ChrA ELISA Kit, fj05f09 ELISA Kit, si:dkey-177p2.2 ELISA Kit, wu:fj05f09 ELISA Kit, zgc:101749 ELISA Kit, zgc:56075 ELISA Kit, CHGA ELISA Kit, cgA ELISA Kit, chga ELISA Kit, chromogranin A ELISA Kit, glycoprotein hormones, alpha polypeptide ELISA Kit, uncharacterized CHGA ELISA Kit, chromogranin A S homeolog ELISA Kit, CHGA ELISA Kit, CGA ELISA Kit, Chga ELISA Kit, chga ELISA Kit, chga.S ELISA Kit

Hintergrund: Chromogranin A is a 49 kDa acidic protein that consists of 439 amino acids encoded on chromosome 14. Chromogranin A has been identified in a number of normal and neopastic endocrine tissues. It is demonstrated that an elevated level of circulating chromogranin A is a marker for tumors of neuroendocrine origin. However, the most significant clinical use of chromogranin A is related to the diagnostic procedure in patients with pheochromocytoma. The following is a short summary of the potential usages of chromogranin A. 1. A very sensitive (83 % ) and highly specific (96 %) marker in the evaluation of actual or suspected pheochromocytoma. Drugs commonly employed in the diagnosis or treatment of pheochromocytoma have little effect on the plasma chromogranin A level, which is a great advantage of measuring chromogranin A over catecholamines. 2. To ascertain the source of a tumor. A high chromogranin A level indicates that the tumor arises from neuroendocrine tissues.3. Endocrine tumors that do not produce their specific hormones, for example, calcitonin negative but chromogranin A positive C-cell carcinoma, zero-cell carcinoma, beta-cell carcinoma, parathyroid carcinoma.

Molekulargewicht: 49 kDa

Gen-ID: 1113

NCBI Accession: NP_001266

UniProt: P10645

Pathways: Negative Regulation of Hormone Secretion, cAMP Metabolic Process, Regulation of G-Protein Coupled Receptor Protein Signaling