Interferon gamma (IFN-γ) is produced by cells that are involved in inflammatory immune responses. IFN-γ is predominantly released by memory and effector CD4+
T cells as well as by NK cells upon activation. Quantitative analysis of IFN-γ–producing cells can provide important information on the course of immune responses. Anti-IFN‑γ antibodies are designed for intracellular staining of IFN‑γ–producing cells. Cells can be stimulated for IFN-γ–production, e.g. by polyclonal stimulation with mitogens. For induction of IFN‑γ–production by antigen-specific T cells, cells are restimulated with respective antigen. IFN‑γ can be accumulated in the cells by addition of secretion inhibitors like brefeldin A. After fixation and permeabilization of the cell sample, IFN‑γ–producing cells can be stained intracellularly with Anti-IFN‑γ antibodies. Staining of surface markers allows simultaneous flow cytometric analysis of subsets and activation status of the IFN-γ–producing cells.