Mouse IL-2 ELISPOT Kit

Interleukin-2 (IL-2) is a pleiotropic cytokine that drives T-cell growth, augments NK cytolytic activity, induces the differentiation of regulatory T cells, and mediates activation-induced cell death. IL-2 does not specify the type of Th differentiation that occurs; instead, IL-2 modulates expression of receptors for other cytokines and transcription factors, thereby either promoting or inhibiting cytokine cascades that correlate with each Th differentiation state. The Mouse IL-2 ELISpot assay is designed for the detection of mouse IL-2 secreting cells at the single cell level and can be used to quantitate the frequency of mouse IL-2 secreting cells. ELISpot assays are highly reproducible and sensitive, and can be used to measure responses with frequencies well below 1 in 100,000. ELISpot assays do not require prior in vitro expansion of T cells and they are suitable for high-throughput analysis using only small volumes of primary cells. As such, ELISpot assays are useful tools for research in vaccine development, cytokine secretion and the monitoring of various clinical trials.

This kit is used to detect IL-2 at the cellular level.

It is for research use only.

Please see Certificate of Analysis for details of reconstitution instruction and specific concentration.

1. Unopened kit should be stored at 2℃-8℃ upon receiving.

2. Find the expiration date on the outside packaging and do not use reagents past their expiration date.

3. The opened kit should be stored per components table. The shelf life is 30 days from the date of opening.

ELISpot assays employ the quantitative sandwich enzyme-linked immunosorbent assay (ELISA) technique. The Kit consists of Pre-coated Anti-IL-2 Antibody Microplate, Positive Stimulus, Biotin-Anti-IL-2 Antibody, Streptavidin-HRP, AEC and buffers.

Your experiment will include 6 simple steps:

a) A monoclonal antibody specific for mouse IL-2 has been pre-coated onto a polyvinylidene difluoride (PVDF)-backed microplate.

b) Appropriately stimulated cells are pipetted into the wells and the microplate is placed into a humidified 37 °C CO2 incubator for a specified period of time.During this incubation period, the immobilized antibody in the immediate vicinity of the secreting cells binds secreted IL-2.

c) After washing away any cells and unbound substances, a biotinylated antibody specific for mouse IL-2 is added to the wells. Following a wash to remove any unbound biotinylated antibody, HRP conjugated to streptavidin is added.

d) Wash the plate and add the Streptavidin-HRP diluted by Dilution Buffer to the plate.

e) Wash the plate and add AEC. A reddish-brown colored precipitate forms and appears as spots at the sites of cytokine localization, with each individual spot representing an individual IL-2 secreting cell.

f) The spots can be counted with an automated ELISpot reader system or manually using a stereomicroscope.