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Enzyme-linked immunosorbent spot (ELISPOT) is an affinity-based method, usually carried out in multi-well plates, for visualizing an immune response to an antigen by detecting immune cell secretions like cytokines. In ELISPOT, antibodies specific to the desired secreted product, often a cytokine, are immobilized in plates with membrane-backed wells. Immune cells, such as T cells, are cultured in the plate with the antigen to be tested. The plate is washed to remove everything but what is bound to the immobilized antibodies. Each secreting cell leaves behind a “spot” of bound product. The bound product is then detected either directly, when the antigen is bound by a labeled primary antibody, or indirectly, when the bound unlabeled primary antibody is then bound by a second labeled antibody (secondary antibody). The antibody label usually creates a color change, weak radioactivity, fluorescence, or chemiluminescence so the number of spots can be counted. ELISPOT is commonly used in immunology research, as a diagnostic for infections and disease such as tuberculosis, and in vaccine development. (Credit: Brooke Anderson-White)
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