C-Reactive Protein (CRP) ELISA Kit

Antigen: C-Reactive Protein (CRP)

Synonyme: PTX1, MGC88244, MGC149895, AI255847, Aa1249, Ac1262, Ab1-341, Ab2-196, Ac1-114, Ac2-069, Ba2-693, CRP, crp

Reaktivität: Rind (Kuh)

Applikation: ELISA

Zertifikate: ISO 9001:2008

Produktnummer: ABIN364618

Produktbeschreibung

Produktmerkmale: This immunoassay allows for the in vitro quantitative determination of bovine CRP concentrations in serum, plasma and other biological fluids.

Proben: Serum, Plasma

Beschreibung: C-reactive protein (CRP) is a protein found in the blood in response to inflammation (an acute-phase protein). CRP is synthesized by the liver in response to factors released by fat cells (adipocytes). It is a member of the pentraxin family of proteins. CRP is a member of the class of acute-phase reactants as its levels rise dramatically during inflammatory processes occurring in the body. This increment is due to a rise in the plasma concentration of IL-6, which is produced predominantly by macrophages as well as adipocytes. CRP binds to phosphocholine on microbes. It is thought to assist in complement binding to foreign and damaged cells and enhances phagocytosis by macrophages, which express a receptor for CRP. It is also believed to play another important role in innate immunity, as an early defense system against infections. CRP rises up to 50,000-fold in acute inflammation, such as infection. It rises above normal limits within 6 hours, and peaks at 48 hours. Its half-life is constant, and therefore its level is mainly determined by the rate of production (and hence the severity of the precipitating cause). Serum amyloid A is a related acute-phase marker that responds rapidly in similar circumstances. 3

Spezifität: This assay recognizes recombinant and natural bovine CRP. No significant cross-reactivity or interference was observed.

Anwendungen

Prinzip: The microtiter plate provided in this kit has been pre-coated with an antibody specific to CRP. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific for CRP and Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Then a TMB (3,3',5,5' tetramethyl-benzidine) substrate solution is added to each well. Only those wells that contain CRP, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The concentration of CRP in the samples is then determined by comparing the O.D. of the samples to the standard curve.

Protokoll: Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate. All the reagents should be added directly to the liquid level in the well. The pipette should avoid contacting the inner wall of the well. 1. Add 100μl of Standard, Blank, or Sample per well. Cover with the adhesive strip. Incubate for 2 hours at 37°C. 2. Remove the liquid of each well, don’t wash. 3. Add 100μl of Biotin-antibody working solution to each well. Incubate for 1 hour at 37°C. Biotin-antibody working solution may appear cloudy. Warm up to room temperature and mix gently until solution appears uniform. 4. Aspirate each well and wash, repeating the process three times for a total of three washes. Wash: Fill each well with Wash Buffer (200μl) a nd let it stand for 2 minutes, then remove the liquid by flicking the plate over a sink. The remaining drops are removed by patting the plate on a paper towel. Complete removal of liquid at each step is essential to good performance. 9 5. Add 100μl of HRP-avidin working solution to each well. Cover the microtiter plate with a new adhesive strip. Incubate for 1 hour at 37°C. 6. Repeat the aspiration and wash five times as step 4. 7. Add 90μl of TMB Substrate to each well. Incubate for 10-30 minutes at 37°C. Keeping the plate away from drafts and other temperature fluctuations in the dark. 8. Add 50μl of Stop Solution to each well when the first four wells containing the highest concentration of standards develop obvious blue color. If color change does not appear uniform, gently tap the plate to ensure thorough mixing. 9. Determine the optical density of each well within 30 minutes, using a microplate reader set to 450 nm.

Testdurchführung: Assay Time: 1-3h
Sample Volume: 50-100ul

Applikationshinweise: Detection Wavelength: 450 nm

Lagerung: 1. Unopened test kits should be stored at 2-8°C upon receipt and the microtiter plate should be kept in a sealed bag. The test kit may be used throughout the expiration date of the kit, provided it is stored as prescribed above. Refer to the package label for the expiration date. 2. Opened test plate should be stored at 2-8°C in the aluminum foil bag with desiccants to minimize exposure to damp air. The kits will remain stable until the expiring date shown, provided it is stored as prescribed above. 3. A microtiter plate reader with a bandwidth of 10 nm or less and an optical density range of 0-3 OD or greater at 450nm wavelength is acceptable for use in absorbance measurement.

Forschungsgebiet: Serum/Plasma Proteine, Inflammation

Beschränkungen: Nur für Forschungszwecke einsetzbar