Granzyme/Perforin pathway
One mechanism used by cytotoxicT lymphocytes and NK cells to kill tumor cells and virus-infected cells is the release of granzyme and perforin (a membrane pore-protein) proteins from cytotoxic granules. Following adhesion of the cytotoxic cell to the target, granzyme B is directly exocytosed and enters the target cell, assisted by perforin. Perforin polymerizes into transmembrane pores facilitating the intracellular delivery of granzymes. Granzyme B is a serine protease and induces cleavage of procaspase-8 which activates a caspase cascade and cleavage of factors like ICAD that results in the apoptotic death of the cell.
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 number of peptides.
The granzyme B and perforin ELISPOT assays offer the advantages of being rapid and technically straightforward, requiring little sophisticated equipment, and they permit measurement of low-frequency T cells from a relatively small number of cells.
Principle
After cell stimulation, locally produced Granzyme B or Perforin is captured by a specific monoclonal antibody. After cell lysis, trapped Granzyme B or Perforin 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 or Perforin production by individual cells.
Applications
• Vaccine development
• Viral infection monitoring and treatment
• Cancerology
• Autoimmune diseases
ELISPOT Kit contents:
Available in 1, 5, 10, 15 or 20 x 96 tests:
• Granzyme B or Perforin Capture and detection antibodies.
• Streptavidin Alkaline Phosphatase Conjugate.
• Buffers.
• Blocking reagent.
• Ready-to-use substrate buffer.
• With or without PVDF plates.
Perforin Products Available
Human Perforin ELISPOT Kit, with sterile PVDF plates, 1 x 96 tests
Human Perforin ELISPOT Kit, with sterile PVDF plates, 5 x 96 tests
Human Perforin ELISPOT Kit, with sterile PVDF plates, 10 x 96 tests
Human Perforin ELISPOT Kit, with sterile PVDF plates, 15 x 96 tests
Human Perforin ELISPOT Kit, with sterile PVDF plates, 20 x 96 tests
Human Perforin ELISPOT Kit, 5 x 96 tests
Human Perforin ELISPOT Kit, 10 x 96 tests
Human Perforin ELISPOT Kit, 15 x 96 tests
Human Perforin ELISPOT Kit, 20 x 96 tests
Related Perforin products also available:
Human Perforin ELISA Kit, 1 x 96 tests
Human Perforin ELISA Kit, 2 x 96 tests
Anti-Human Perforin, Azide Free, Clone B-D48, mAb, 200µg
Anti-Human Perforin, PE conjugated, Clone B-D48, mAb, 100 tests
Anti-Human Perforin, uconjugated, Clone B-D48, mAb, 200 tests
Granzyme B Products Available
Human Granzyme B ELISPOT, 5x96 tests
Human Granzyme B ELISPOT, 10x96 tests
Human Granzyme B ELISPOT, 15x96 tests
Human Granzyme B ELISPOT, 20x96 tests
Human Granzyme B ELISPOT with PVDF plates, 5x96 tests
Human Granzyme B ELISPOT with PVDF plates, 10x96 tests
Human Granzyme B ELISPOT with PVDF plates, 15x96 tests
Human Granzyme B ELISPOT with PVDF plates, 20x96 tests
Related products also available:
Human Granzyme B ELISA Kit, 1x96 tests
Human Granzyme B ELISA Kit, 2x96 tests
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