Principle:This IL-4 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 polyclonal antibody specific to IL-4. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated monoclonal antibody preparation specific for IL-4 and incubated. IL-4 if present, will bind and become immobilized by the antibody pre-coated on the wells and then be "sandwiched" by biotin conjugate. The microtiter plate wells are thoroughly washed to remove unbound IL-4 and other components of the sample. In order to quantitate the amount of IL-4 present in 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 have 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-4, 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 450nm ± 2nm.
Kit Components:I8426-01M: Microtiter Plate 1x96wellsI8426-01N: Mab (Biotin) 1x7mlI8426-01P: Avidin (HRP) 1x14ml*I8426-01Q: Standard 2x1vialI8426-01R: Calibrator Diluent I 1x22ml For serum/plasma testingI8426-01S: Calibrator Diluent II 1x22ml For culture supernatantI8426-01T: Wash Buffer, 20X 1x60mlI8426-01U: Substrate A 1x10mlI8426-01V: Substrate B 1x10mlI8426-01W: Stop Solution 1x14ml
Specificity:This sandwich ELISA recognizes both natural and recombinant human IL-4, exhibiting no detectable crossreactivity with human; SAA, EGF, IL-8, IL-16, MCAF, MCP-3, TNF-g, M-CSF, GM-CSF, TGF-b1, RANTES, FGF, MIP-1a, CRP, EPO, IFN-g.
Storage and Stability:Store *I8426-01Q: Standard powder at 4°C, liquid stock at -20°C. Store other components at 4°C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vial prior to removing the cap.
