Cell Sciences, Inc.
Neponset Valley Tech Park
480 Neponset Str., Bldg. 12A
Canton, MA, 02021
Website: http://www.cellsciences.com
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Human Granzyme B ELISPOT Assay
Granzyme B is a serine protease found in the lymphocyte granules of activated cytotoxic T cells and NK cells. Lymphocyte granule-mediated cytotoxicity appears to protect the host from invasion by pathogens and tumor cells. Granzyme B is critically involved in the rapid apoptotic signal delivered by CTL and NK cells. Following adhesion of the cytotoxic cell to the target, granzyme B is directly exocytosed and enters the target cell, assisted by perforin, a membrane pore-forming protein also present in lymphocyte granules. Perforin polymerizes into transmembrane pores facilitating the intra-cellular delivery of the granzymes. Since granzymes and perforin constitute the primary effectors of the granule exocytose pathway, identification of this phenomena by development of assays to assess the involvement of activated CTL or NK in pathological processes is of high interest.
The granzyme B Elispot assay offers a rapid, technically straightforward test and permits measurement of low-frequency T cells from a relatively small number of cells. After cell stimulation, locally produced granzyme B. is captured by a specific monoclonal antibody. After cell lysis, trapped granzyme B molecules are revealed by a secondary biotinylated detection antibody, which is in turn recognized by streptavidin conjugated to alkaline phosphatase. PVDF-bottomed-well plates are then incubated with BCIP/NBT substrate. Colored "purple" spots indicate Granzyme B production by individual cells. Applications include vaccine development, viral infection monitoring and treatment, cancer research, autoimmune disease research.
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