Principle of Assay
The Human IL-17A ELISpot Kit (A334658) employs the double-antibody sandwich ELISpot technique for the detection of IL-17A-secreting cells. A capture antibody specific for IL-17A has been pre-coated onto a 96-well microplate. Test cells, such as PBMCs, are added into the wells, and upon stimulation with a cell stimulant, IL-17A secreted by individual cells is captured by the immobilized antibody. A biotinylated anti-IL-17A detection antibody, which also binds the captured cytokine, and Streptavidin-HRP, which binds the biotinylated detection antibody, are then added, and the plate is incubated. Following incubation, unbound detection antibody and Streptavidin-HRP are removed by washing, and AEC substrate solution is added. Colored spots develop at the sites of cytokine secretion in proportion to the number of IL-17A-producing cells. The intensity and number of spots can then be quantified using an ELISpot reader. This ready-to-use kit includes pre-coated plates, detection antibody, Streptavidin-HRP, AEC coloring system, dilution buffer, cell stimulant, and washing buffer.