Product nameHuman Granzyme B ELISPOT Kit
See all Granzyme B kits
Sample typeCell culture supernatant, Cell culture extracts
Assay typeSandwich (qualitative)
Assay durationMultiple steps standard assay
Species reactivityReacts with: Human
This ELISPOT assay is designed to enumerate Granzyme B producing cells in a single cell suspension. This method has the advantage of requiring a minimum of in-vitro manipulations allowing Granzyme B production analysis as close as possible to in-vivo conditions in a highly specific way. This technique is designed to determine the frequency of Granzuyme B producing cells under a given stimulation, and the follow-up of such frequency during a treatment and/or a pathological state. Elispot assay constitutes an ideal tool in the TH1 / TH2 response, vaccine development, viral infection monitoring and treatment, cancerology, infectious diseases, autoimmune diseases and transplantation.
The Elispot assay is based on sandwich immuno-enzyme technology. Cell secreted cytokines or soluble molecules are captured by coated antibodies avoiding diffusion in supernatant, protease degradation or binding on soluble membrane receptors. After cell removal, the captured cytokines are revealed by tracer antibodies and appropriate conjugates.
Principle of Method
After cell stimulation, locally produced cytokines are captured by a specific monoclonal antibody. After cell lysis, trapped cytokine 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 cytokine production by individual cells.
Recognizes natural human Granzyme B
Tested applicationsSuitable for: ELISpotmore details
Storage instructionsStore at +4°C. Please refer to protocols.
Components 5 x 96 tests 10 x 96 tests 96 PVDF-bottomed-well plates. 5 units 10 units Bovine Serum Albumin 1 x 1g 2 x 1g Dry Skimmed milk 1 x 1g 2 x 1g Biotinylated detection antibody (Lyophilized) 1 vial 2 vials Granzyme B Capture Antibody 1 x 500µl 2 x 500µl Ready-to-use BCIP/NBT substrate buffer 2 x 27ml 4 x 27ml Streptavidin - Alkaline Phosphatase conjugated 1 x 50µl 2 x 50µl
RelevanceCytolytic T lymphocytes and natural killer cells share the remarkable ability to recognize, bind, and lyse specific target cells. They are thought to protect their host by lysing cells bearing on their surface 'nonself' antigens, usually peptides or proteins resulting from infection by intracellular pathogens. Granzyme B is crucial for the rapid induction of target cell apoptosis by CTL in cell-mediated immune response.
Cellular localizationCytoplasmic granule. Cytoplasmic granules of cytolytic T-lymphocytes and natural killer cells
- Cathepsin G like 1
Our Abpromise guarantee covers the use of ab46617 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ELISpot||Use at an assay dependent dilution.|
Measurement of Granzyme B release from Human NK cells using ab46617 - Granzyme B Human Elispot Kit.
NK cells were cultured with rIL-2 and with/without TKD peptide (2 µg/ml) for 4–5 days prior to stimulation with i/uRBC (1:3 or 10:1), before being isolated. Granzyme B release was determined via ELISPOT assay following the protocol, with 2000 effector cells. Experiments were repeated using a blocking antibody directed against Hsp70.
Image from Böttger E et al., PLoS One. 2012;7(3):e33774. doi: 10.1371/journal.pone.0033774. Epub 2012 Mar 15.; Fig 6.; March 15, 2012, PLoS ONE 7(3): e33774.