[ Coordinator's note: We try very hard to discourage commercial advertisements on the ] [ mailing list. On the other hand, we also try very hard to help facilitate the spread ] [ of information relevant to cytometry. In the spirit of 'information is good', we are ] [ sending this note on. If you think it crosses the line into junk-mail advertisement ] [ please let me know personally at kelley@flowcyt.cyto.purdue.edu. ] [ The politeness and restraint of our friends in the vendor community are very much ] [ appreciated by all subscribers and have been a big part of our success as a useful ] [ mailing list, and we really want to continue as an ad-free, but information-rich ] ] [ information medium. ] [ Steve ] Cell Sciences Offers 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 for the scientific community. The chromium release assay or the use of MHC: peptide tetramer staining constitute conventional methods to measure antigen/peptide-specific CD8+ T cell immune response. However, because of their low sensitivity and specific experimental conditions requirements, these assays are not suitable for screening large numbers of peptides. The granzyme B Elispot assay offers a rapid, technically straightforward test requiring little sophisticated equipment and permits measurement of low-frequency T cells from a relatively small number of cells. Principle: 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 recognised 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: * Vaccine development * Viral infection monitoring and treatment * Cancer research * Autoimmune disease research Irwin Libeskind Cell Sciences, Inc. 1504 Providence Highway, No. 11B Norwood, MA 02062 USA Toll Free: 888 769-1246 Tel: 781-769-0610 Fax: 781-769-0542 E-mail: irwin@cellsciences.com www.cellsciences.com
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