| Interleukin-25 (IL-25), also known as Interleukin-17E (IL-17E), is a protein that in humans is encoded by the IL25 gene. It is a cytokine that belongs to the IL-17 cytokine family and mapped to 14q11.2. IL-25 is secreted by type 2 helper T cells (Th2) and mast cells. It has been found that IL-25 can induce Th2-type cytokine expression and production by non-T, non-B accessory cells that express high levels of major histocompatibility complex class II and low levels of Cd11c. This cytokine also can induce NF-kB activation, and stimulate the production of IL8. IL-25 is an important molecule controlling immunity of the gut and has been implicated in chronic inflammation associated with the gastrointestinal tract. The IL-17E/IL-25, BioAssay(TM) Kit is a sandwich ELISA for quantitative detection of human IL-17E/IL-25 in cell culture supernates, cell lysates, serum, plasma (heparin, EDTA). Assay Range: 62.5-4000pg/ml Sensitivity: <10pg/ml Specificity: Natural and recombinant human IL-17E. Test Principle: The IL-17E/IL-25, BioAssay(TM) ELISA Kit is a solid-phase immunoassay specially designed to measure Human IL25 with a 96-well strip plate that is pre-coated with antibody specific for IL25. The detection antibody is a biotinylated antibody specific for IL25. The capture antibody is monoclonal antibody from mouse and the detection antibody is polyclonal antibody from goat. The kit contains recombinant human IL-17E/IL-25 with immunogen: Expression system for standard: E. coli, Immunogen sequence: Y33-G177 To measure human IL25, add standards and samples to the wells, then add the biotinylated detection antibody. Wash the wells with PBS or TBS buffer, and add Avidin-Biotin-Peroxidase Complex (ABC-HRP). Wash away the unbounded ABC-HRP with PBS or TBS buffer and add TMB. TMB is an HRP substrate and will be catalyzed to produce a blue color product, which changes into yellow after adding the acidic stop solution. The absorbance of the yellow product at 450nm is linearly proportional to Human IL25 in the sample. Read the absorbance of the yellow product in each well using a plate reader, and benchmark the sample wells readings against the standard curve to determine the concentration of Human IL25 in the sample. Kit Components: Microtiter Strip, 1x96-well Human IL25 Standard, 2x1vial Anti-human IL25 (Biotin), 1x100ul (1:100 dilution) Avidin-Biotin-Peroxidase Complex (ABC), 1x100ul (1:100 dilution) Sample diluent buffer, 1x30ml Antibody diluent buffer, 1x12ml ABC diluent buffer, 1x12ml TMB color developing agent, 1x10ml Stop solution, 1x10ml Wash Buffer, 25X, 1x20ml Storage and Stability: Store human IL25 standard vials at 4C. Once reconstituted store at 4C for up to 12 hours or at -20C for up to 48 hours. Store other components at 4C. Stable for 6 months For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Assay Summary: 1. Add 100ul of samples and standards and incubate the plate at 37C for 90 minutes or at RT for 2 hours. Do not wash. 2. Add 100ul biotinylated antibody and incubate the plate at 37C for 60 minutes or at RT for 90 minutes. Wash plate 3 times with Working Wash Buffer. 3. Add 100ul of ABC working solution and incubate the plate at 37C for 30 minutes or at RT for 40 minutes. Wash plate 5 times with Working Wash Buffer. 4. Add 90ul of TMB color developing agent and incubate the plate at 37C in dark for 15-25 minutes or at RT for 30 minutes. 5. Add 100ul TMB Stop Solution and read. |
