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R&D Systems ELISpot Kits are complete microplate-based immunoassays designed for the detection and enumeration of single cells secreting cytokines or other antigens. These assays are highly sensitive and can quantitate actively secreting cells even when cell frequencies fall below 1 in 100,000. Unlike other methods that assess antigen-specific immune cell responses, this assay does not require prior in vitro expansion of cells and is suitable for high-throughput analysis. ELISpot Kits are complete ready-to-run assays and require no further development or refinement.
R&D Systems ELISpot Kits employ antibody-coated, membrane-backed microplates to identify secreting cells. The immobilized capture antibodies bind to cytokines that have been secreted in their vicinity, and enzyme conjugates are used to visualize the secreted proteins. Colored precipitates form at the sites of cytokine localization and appear as spots, with each individual spot representing an individual cytokine-secreting cell.
|Mouse splenocytes enriched for CD4+ cells were assessed for Granzyme B secretion using the Mouse CD4+/Granzyme B ELISpot Kit (Catalog # EL6024). CD4+ cells were first enriched by a short incubation in ELISpot wells coated with anti-CD4 antibodies. Following a wash, CD4+ enriched cells were then stimulated overnight in culture media with PMA/Ca2+ ionomycin. During the incubation, anti-mouse Granzyme B antibodies capture the secreted enzyme, which is then visualized as blue spots.|