Human Tumor Necrosis Factor alpha ELISA Kit

Applications:Sandwich ELISA, Reactivity:Human, Cross reactivity:This assay has shown no cross-reactivity with other cytokines tested, Specificity:This sandwich ELISA recognises both natural and recombinant human TNF-?, Product range:4-2000 pg/ml, Sensitivity:< 4 pg/ml, Storage:Store at 2-8?C, Target:Tumor Necrosis Factor alpha, Sample type:Serum, plasma, cell culture supernatant, and other biological fluids, Principle assay:This TNF-? 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 TNF-?. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated monoclonal antibody preparation specific for TNF-? and incubated. TNF-?, 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 TNF-? and other components of the sample. In order to quantify the amount of TNF-? 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 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-enzidine) 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 TNF-?, 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 450 nm ? 2 nm.

In order to measure the concentration of TNF-? 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 TNF-? concentration (pg/mL). The concentration of TNF-? in the samples is then determined by comparing the O.D. of the samples to the standard curve, Assay type:Sandwich (quantitative), Detection type:Colorimetric, Platform:Microplate, Assay time:2h 30m – 3h

+25° C.