Mouse IFN-gamma/IL-5 Dual-Color ELISpot Kit

  • Assay Type
    Quantitative Sandwich ELISA
  • Assay Format
    96-well PVDF-backed microplate
  • Assay Length
    3 hours 35 mins to 4 hours 50 mins*
  • Sample Type
    Whole Cells
  • Specificity
    Please see the product datasheet
* 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 IgG/IgM. 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 IgG/IgM. 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 IgG/IgM.

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 IgG/IgM 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 IgG/IgM
  • 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


  • Microplate
  • Biotin-conjugated Detection Antibody
  • HRP-conjugated Detection Antibody
  • Streptavidin-conjugated to Alkaline Phosphatase
  • Dilution Buffers
  • Wash Buffer Concentrate
  • BCIP/NBT Chromogen
  • AEC Chromogen
  • Positive Controls


  • 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

Background IFN-gamma/IL-5


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