FluoroSpot Plus: Human IFN‑γ/IL‑4/IL‑5
FluoroSpot Plus: Human IFN‑γ/IL‑4/IL‑5
Components
| Plate | Pre-coated FluoroSpot plate (mAbs 1-D1K, IL4-I, and TRFK5) |
| Detection mAbs | Anti-IFN-γ mAb (7-B6-1), BAM |
| Anti-IL-4 mAb (IL4-II), biotin | |
| Anti-IL-5 mAb (5A10), WASP | |
| Fluorophore conjugates | Anti-BAM mAb, 490 |
| SA-550 | |
| Anti-WASP mAb, 640 | |
| Buffer/Solution | FluoroSpot enhancer |
| Positive control | Anti-CD3 mAb (CD3-2) |
| Co-stimulus | Anti-CD28 mAb (CD28-A) |
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Performance
PBMC (250,000 cells/well) were incubated overnight in the absence or presence of mixture of CEF and CEFTA peptide pools (available from Mabtech, code 3616-1, 3617-1) and anti-CD28 mAb. The number of cells secreting IFN-γ, IL-4 and IL-5 was analyzed by FluoroSpot. Single cytokine secreting cells as well as double- and triple-secreting cells were determined. Spot center was used to identify spots from double- and triple-secreting cells, visualized in an image overlay of LED490, FITC (IFN-γ) and LED550, Cy3 (IL-4) and LED640, Cy5 (IL-5). Spot analysis was made with Mabtech IRIS.
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Analyte information
IFN-γ
| Analyte description | Interferon-γ (IFN-γ) is the only type II interferon. This proinflammatory cytokine is secreted by activated T cells and NK cells. It activates macrophages and endothelial cells and regulates immune responses by affecting APCs, T cells, and B cells. Production of IFN-γ by helper T cells and cytotoxic T cells is a hallmark of the Th1-type phenotype. Thus, high-level production of IFN-γ is typically associated with effective host defense against intracellular pathogens. |
| Alternative names | Interferon-γ, Interferon-gamma, IFN-γ, IFN-gamma, IFN-g, IFNg, IFG, IFI, IMD69 |
| Cell type | T cell, Tc, Th1, NK cell |
| Gene ID | 3458 |
IL-4
| Analyte description | Interleukin 4 (IL-4) is produced primarily by Th2 cells, mast cells, eosinophils, and basophils. IL-4 was first identified to costimulate B cell growth; the cytokine is essential for B cell secretion of IgE and augments the production of IgG1 in vivo. In addition to regulating B cell growth and immunoglobulin secretion, IL-4 also affects T cells. In vitro, IL-4 promotes T cell growth and can induce cytolytic T cell activity. IL-4 also induces the differentiation of naive CD4 T cells into Th2 cells, which are characterized by their capacity to secrete the cytokines IL-4, IL-5, and IL-10 upon activation, while simultaneously inhibiting the generation of Th1 cells. |
| Alternative names | Interleukin 4, IL-4, IL4, BCGF-1, BSF-1, BCGF1, BSF1 |
| Cell type | T cell, Th2, Treg |
| Gene ID | 3565 |
IL-5
| Analyte description | Interleukin 5 (IL-5) is a pleiotropic cytokine primarily produced by T and NK cells. IL-5 specifically controls the survival, differentiation, and chemotaxis of eosinophils. But initially, IL-5 was characterized by its ability to support the growth and terminal differentiation of B cells. |
| Alternative names | Interleukin-5, IL-5, IL5, BCGF-2, EDF, TRF |
| Cell type | T cell, Th2, NK cell |
| Gene ID | 3567 |