Mouse IFN-gamma/IL-2 Dual-Color ELISpot Kit Summary
* Provided that the recommended microplates, buffers, diluents, substrates and solutions are used, and the assay is run as summarized in the Assay Procedure provided.
* Following cell stimulation.
ELISpot kits are highly sensitive, microplate-based assays for the detection of cytokine secreting cells. This kit is designed for the simultaneous detection and enumeration of IFN-γ/IL-2. Complete ELISpot kits are ready-to-run and require no assay development or refinement.
Appropriately stimulated cells are pipetted directly into the wells of a PVDF-backed microplate. The PVDF membrane has been pre-coated with capture antibodies specific for IFN-γ/IL-2. The microplate is then placed in a humidified 37 °C CO2 incubator for the desired period of time. During this incubation period, the immobilized capture antibodies in the immediate vicinity of the secreting cells bind secreted IFN-γ/IL-2.
At the desired time point, one analyte is detected using a biotinylated detection antibody and alkaline-phosphatase (AP)-conjugated to streptavidin. The second analyte is detected using a horseradish peroxidase (HRP)-conjugated detection antibody. Following incubation with the chromogens BCIP/NBT (substrate for AP, blue-black precipitate) and AEC (substrate for HRP, red precipitate), spots form to indicate the presence of IFN-γ/IL-2 secreting cells. Each spot represents an individual secreting cell. The spots can be counted with an ELISpot reader system or using a stereomicroscope.
- Detect and quantitate individual cells secreting IFN-γ/IL-2
- High sensitivity - ELISpot assays can measure responses with frequencies well below 1 in 100,000 cells
- No in vitro expansion of cells required
- High-throughput - ELISpot assays use only a small number of primary cells
- Biotin-conjugated Detection Antibody
- HRP-conjugated Detection Antibody
- Streptavidin-conjugated to Alkaline Phosphatase
- Dilution Buffers
- Wash Buffer Concentrate
- BCIP/NBT Chromogen
- AEC Chromogen
OTHER REAGENTS REQUIRED
- Pipettes and pipette tips
- Deionized or distilled water
- Squirt bottle, manifold dispenser, or automated microplate washer
- 500 mL graduated cylinder
- 37 °C CO2 incubator
- Sterile culture media
- Dissection microscope or an automated ELISpot reader
Preparation and Storage
Citation for Mouse IFN-gamma/IL-2 Dual-Color ELISpot Kit
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
1 Citation: Showing 1 - 1
The critical role of kinase activity of interleukin-1 receptor-associated kinase 4 in animal models of joint inflammation.
Authors: Koziczak-Holbro M, Littlewood-Evans A, Pollinger B, Kovarik J, Dawson J, Zenke G, Burkhart C, Muller M, Gram H
Arthritis Rheum., 2009;60(6):1661-71.
Sample Types: Whole Cells
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