title: ELISpot and DC-ELISpot Assay to Measure Frequency of Antigen-Specific IFNγ-Secreting Cells.
abstract: ELISpot is a highly sensitive method in immunology to enumerate cells producing a given cytokine. Cells are stimulated in a microtiter plate pre-coated with a specific anti-analyte antibody. In response to the stimulation, cells release cytokines that are bound to the anti-analyte antibody. After a washing step, which removes the cells from the wells, the location of the cytokine-releasing cell is visualized by an enzyme-labeled detection antibody and its corresponding chromogenic substrate. The end result is a set of colored spots, each of which represents an area where a cell secreting the cytokine had been located. Here we describe the standard ELISpot protocol and a variation denominated dendritic cell (DC)-ELISpot for the detection of IFNγ-secreting cells upon stimulation with oligopeptides and protein antigens, respectively.
authors.affiliation: Department of Hematology, Leiden University Medical Center, 9600, Leiden, 2300 RC, The Netherlands, m.a.navarrete@lumc.nl.
authors.forename: Marcelo A
authors.initials: MA
authors.lastname: Navarrete
mesh_terms: D000469:Alkaline Phosphatase / Q000737:chemistry; D055910:Antibodies, Immobilized / Q000737:chemistry; D000941:Antigens / Q000494:pharmacology; D002452:Cell Count; D003713:Dendritic Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D058501:Enzyme-Linked Immunospot Assay / Q000379:methods*; D006801:Humans; D007371:Interferon-gamma / Q000032:analysis* / Q000096:biosynthesis / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009842:Oligopeptides / Q000494:pharmacology; D019809:Streptavidin / Q000737:chemistry
chemical_list: D055910:Antibodies, Immobilized; D000941:Antigens; D009842:Oligopeptides; D007371:Interferon-gamma; D019809:Streptavidin; D000469:Alkaline Phosphatase