Interleukin -17A (IL-17A) is a secreted, homodimeric glycoprotein linked by disulfide link with a molecular mass of 35kD. The cytokine is the proto-type of a newly discovered pro-inflammatory cytokine family which consists of IL-17A, IL-17B, IL-17C, IL- 17D, IL-17E (IL-25) and IL-17F. All the IL-17s have similar structure with four highly conserved cysteine residues and same 3 dimensional structure, which distinguish this family from other cytokines.IL-17 is produced by a group of CD4 T help cells termed Th17 cells. In addition to previous discovered Th1 and Th2 cells, the Th17 cells consist in the third group of T help cells. IL-23, a growth and stabilization factor was the first cytokine that was found to involve with the development of Th 17 cells. Later on, other cytokines including TGF-beta, IL-6, IL-21 were also found to contribute to the procedure. The transcription factors implicated in the procedure were identified as STAT3, RORgt, and RORa. IL-17, like interferon-g, is a potent mediator of delayed-type reaction. IL-17 induces the secretion of many other pro-inflammatory cytokines and chemokines such as IL-6, IL-8, GM-CSF, G- CSF, TGF-beta, TNF-alpha, GRO-alpha, MCP-1, and stimulates the expression of NF-kappaB, mitogen activated protein kinase, and prostaglandings from many cell types. By stimulating cytokine production and recruiting monocytes neutrophils to site of inflammation, the cytokine exerts its effect in response to tissue damage and invasion.Excessive expression of IL-17 was found to associate with autoimmune diseases and chronic inflammatory diseases such as multiple sclerosis, rheumatoid arthritis, psoriasis, inflammatory bowel diseases, tissue allograft rejection, allergic asthma and infectious diseases at the mucosal tissue.Sample Type:Serum, plasma, cell culture supernatant, and other biological fluidsIntended Use:This IL17A, Human, BioAssay ELISA Kit is to be used for the in vitro quantitative determination of human interleukin-17A concentrations in serum, plasma, cell culture supernatant, and other biological fluids. This kit is intended for LABORATORY RESEARCH ONLY and is not to be used for diagnostic or therapeutic procedures.Sensitivity:≤5pg/mlAssay Range:5-1000pg/mlSpecificity:Recognizes both natural and recombinant human IL-17A. No significant cross-reactivity with following human cytokines and growth factors: IL-1a, IL-1b, IL-2, IL-4, IL-5, Il-6, IL-7, IL-8, IL-10, IL-11, Il-12, Il-13Ra2, FGF basic, GM-CSF, IFN-g, M-CSF, MCP-1(MCAF), MCP3, EGF, TNF-a, TNF-b.Test Principle:This IL-17A enzyme linked immunosorbent assay (ELISA) applies a technique called a quantitative sandwich immunoassay. The microtiter plate provided in this kit has been pre-coated with a monoclonal antibody specific to IL-17A. Standards or samples are then added to the appropriate microtiter plate wells. A biotin-conjugated antibody preparation specific for IL-17A was added and incubated. IL-17A, if present, will bind and become immobilized by the antibody pre-coated on the wells. The microtiter plate wells are thoroughly washed to remove unbound IL-17A and other components of the sample. Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Avidin is a tetramer containing four identical subunits that each has a high affinity-binding site for biotin. The wells are thoroughly washed to remove all unbound HRP-conjugated Avidin, and a TMB (3,3'5, 5' tetramethyl-benzidine) substrate solution is added to each well. The enzyme (HRP) and substrate are allowed to react over a short incubation period. Only those wells that contain IL-17A, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in colour. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the colour change is measured spectrophotometrically at a wavelength of 450nm ± 2nm.In order to measure the concentration of IL-17A in the samples this kit includes two calibration diluents (Calibrator Diluent I for serum/plasma testing and Calibrator Diluent II for cell culture supernatant testing.) According to the testing system, the provided standard is diluted (2-fold) with the appropriate Calibrator Diluent and assayed at the same time as the samples. This allows the operator to produce a standard curve of Optical Density (O.D) versus IL-17A concentration (pg/mL). The concentration of IL-17A in the samples is then determined by comparing the O.D. of the samples to the standard curve.Kit Components:IL-17A microtiter plate: 1x96 wellsBiotin conjugate (Anti-human IL-17A polyclonal antibody conjugated to Biotin): 1x7mlAvidin conjugate (Avidin conjugated to HRP): 1x12mlIL-17A standard (Recombinant human IL-17A) (2000pg/vial): 1x2 vialsCalibrator diluent 1: 1x25ml Calibrator diluent 2: 1x25mlWasher buffer (20X): 1x60mlSubstarte A (Buffered solution with H2O2): 1x10mlSubstrate B (Buffered solution with TMB): 1x10mlStop Solution (H2SO4): 1x14mlStorage and Stability:Store components at 4°C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
