Interferon alpha (IFNA) ELISA Kit

Antigen: Interferon alpha (IFNA)

Reaktivität: Schwein

Applikation: ELISA

Zertifikate: ISO 9001:2008

Produktnummer: ABIN365525

Produktbeschreibung

Produktmerkmale: This immunoassay allows for the in vitro quantitative determination of porcine IFN-α concentrations in serum, plasma and other biological fluids.

Weitere Bezeichnung: Interferon alpha (IFN-alpha)

Proben: Serum, Plasma

Beschreibung: Interferons (IFNs) are natural cell-signaling proteins produced by the cells of the immune system of most vertebrates in response to challenges such as viruses, parasites and tumor cells. Interferons belong to the large class of glycoproteins known as cytokines. Interferons are produced by a wide variety of cells in response to the presence of double-stranded RNA, a key indicator of viral infection. Interferons assist the immune response by inhibiting viral replication within host cells, activating natural killer cells and macrophages, increasing antigen presentation to lymphocytes, and inducing the resistance of host cells to viral infection. There are three major classes of interferons that have been described for humans according to the type of receptor through which they signal: type I, II and III IFNs.

Spezifität: This assay recognizes recombinant and natural porcine IFN-α. 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 IFN-α. Standards or samples are then added to the appropriate microtiter plate wells with a 3 biotin-conjugated antibody preparation specific for IFN-αand 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 IFN-α, 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 IFN-α 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) and 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 9 good performance. 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: Zytokine

Beschränkungen: Nur für Forschungszwecke einsetzbar

Publikationen

Publikationen: Xiong, Lin, Yuan et al.: "Expression of exogenous IFN-alpha by bypassing the translation block protects cells against FMDV infection." in: Antiviral research, Vol. 84, Issue 1, pp. 60-6, 2009 (PubMed).