Human IL-5 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.
PRODUCT SUMMARY
ELISpot kits are highly sensitive, microplate-based assays for the detection of cytokine secreting cells. This kit is designed for the detection and enumeration of human IL-5. Complete ELISpot kits are ready-to-run and require no assay development or refinement.
This ELISpot assay employs a capture antibody specific for human IL-5, pre-coated onto a PVDF-backed microplate. Appropriately stimulated cells are pipetted directly into the wells and the immobilized antibody in the immediate vicinity of the secreting cells binds secreted human IL-5. Following wash steps and incubation with a biotinylated detection antibody, alkaline-phosphatase conjugated to streptavidin is added. Unbound enzyme is subsequently removed by washing and a substrate solution (BCIP/NBT) is added. A blue-black colored precipitate forms at the sites of cytokine localization and appears as spots, with each individual spot representing an individual human IL-5 secreting cell. The spots can be counted with an automated ELISpot reader system or manually using a stereomicroscope.
PRODUCT FEATURES
- Detect and quantitate individual cells secreting human IL-5
- 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
KIT CONTENTS
- Human IL-5 Microplate
- Biotinylated Detection Antibody
- Streptavidin conjugated to Alkaline Phosphatase
- Dilution Buffers
- Wash Buffer Concentrate
- BCIP/NBT Chromogen
- Human IL-5 Positive Control
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
Product Datasheets
Preparation and Storage
Background: IL-5
IL-5 (Interleukin-5) is a secreted cytokine that is expressed as a covalent dimer. It is produced by activated Th2 cells, eosinophils, mast cells, and iNKT cells and promotes eosinophilic and basophilic differentiation and activation. IL-5 signals through a receptor complex containing IL-5 R alpha and the Common beta Chain/CD131 on eosinophils and basophils. Naturally occurring soluble forms of IL-5 R alpha retain the ability to bind IL-5 and limit its bioavailability.
Citations for Human IL-5 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.
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Helminth antigens modulate immune responses in cells from multiple sclerosis patients through TLR2-dependent mechanisms.
Authors: Correale J, Farez M
J. Immunol., 2009-10-07;183(9):5999-6012.
Species: Human
Sample Types: Whole Cells
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Interferon-gamma is induced in human peripheral blood immune cells in vitro by sodium stibogluconate/interleukin-2 and mediates its antitumor activity in vivo.
Authors: Fan K, Borden E, Yi T
J. Interferon Cytokine Res., 2009-08-01;29(8):451-60.
Species: Human
Sample Types: Whole Cells
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