Use of interferon-gamma ELISPOT in monitoring immune responses in humans

The interferon (IFN)-gamma enzyme-linked immunospot (ELISPOT) assay has become a useful tool for immunologists seeking to quantify immune responses on a per-cell basis. The assay is sensitive and allows for the enumeration of low-frequency T-cells. Many have applied this assay to clinical trials as a way to measure biological activity in a patient cohort. It is critical that each laboratory attempting to use the assay in their facility perform rigorous development and qualification work to establish an assay that suits their particular needs. This chapter serves as a demonstration of two practical and slightly different approaches to using the ELISPOT assay to monitor immune activity in the human periphery: (1) assays using whole samples of peripheral blood mononuclear cells with and without the use of additional antigen presenting cells and (2) assays using enriched T-cell populations. Detailed protocols and procedures will be covered, as well as a demonstration of results obtained from three separate applications.