Interferon Gamma + Granzyme B Human Elispot Kit (ab48464)
- Product nameInterferon Gamma + Granzyme B Human Elispot KitSee all Interferon Gamma + Granzyme B kits ...
- Detection methodColorimetric
- Tests5 x 96 well plate
- Sample typeCell culture supernatant, Cell culture extracts
- Assay typeSandwich (qualitative)
- Assay durationMultiple steps standard assay
- Species reactivityReacts with: Human
- Product overview
The ELISPOT assay is designed to enumerate cytokine producing cells in a single cell suspension. This method has the advantage of requiring a minimum of in-vitro manipulations allowing cytokine production analysis as close as possible to in-vivo conditions in a highly specific way. This technique is designed to determine the frequency of cytokine 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, oncology, infectious diseases, autoimmune diseases and transplantation.
This 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.
The dual colour Elispot allows you to monitor the production of two cytokines simultaneously in the same well.
Principle of the Method
After cell stimulation, locally produced cytokines are captured by IFN gamma and Granzyme B specific monoclonal antibodies. After cell lysis, trapped cytokine molecules are revealed by a secondary anti-IFN gamma FITC conjugated antibody and a biotinylated anti-Granzyme B antibody. Those are in turn recognised by anti-FITC HRP and streptavidin-AP conjugates. PVDF-bottomed-well plates are then incubated first with AEC substrate buffer, washed and subsequently incubated with BCIP/NBT. Coloured red/brownish spots indicate IFN gamma production while Granzyme B is revealed by blue/purple spots.
- Tested applicationsMeasuring secreted proteins from cells more details
- Storage instructionsStore at +4°C. Please refer to protocols.
Components 5 x 96 tests 10 x concentrate buffer for the preparation of AEC buffer 1 x 5ml 50 x concentrate AEC substrate buffer 1 x 1ml 96 PVDF-bottomed-well plates. 5 units Anti-FITC antibody HRP conjugate 1 x 100µl Bovine Serum albumin 1 x 1g Dry skimmed milk 1 unit Granzyme B Biotinylated detection antibody 1 x 0.55ml Granzyme B Capture Antibody 1 x 0.5ml Human IFNγ Capture antibody 1 x 0.5ml IFNγ FITC conjugated detection antibody 1 x 0.55ml Ready-to-use BCIP/NBT substrate buffer 1 x 50ml Streptavidin - Alkaline Phosphatase conjugated 1 x 50µl
- Research Areas
- RelevanceInterferon Gamma is produced by lymphocytes activated by specific antigens or mitogens. IFN-gamma, in addition to having antiviral activity, has important immunoregulatory functions. It is a potent activator of macrophages, it has antiproliferative effects on transformed cells and it can potentiate the antiviral and antitumor effects of the type I interferons. Granzyme B is necessary for target cell lysis in cell-mediated immune responses. It cleaves after Asp. Seems to be linked to an activation cascade of caspases (aspartate-specific cysteine proteases) responsible for apoptosis execution. Cleaves caspase-3, -7, -9 and 10 to give rise to active enzymes mediating apoptosis.
- Cytotoxic T lymphocyte proteinase 2
- Granzyme 2
- Granzyme B
- IFN gamma
- Immune interferon
Our Abpromise guarantee covers the use of ab48464 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
References for Interferon Gamma + Granzyme B Human Elispot Kit (ab48464)
ab48464 has not yet been referenced specifically in any publications.