The FluoroSpot assay is an inventive development of the ELISpot assay and uses fluorescent detection instead of colorimetric. This allows for simultaneous detection of several analytes, which is a major advantage. The human IL-22/IFN-γ/IL-5/IL-17A FluoroSpot kit is perfect for users who wish to have a convenient assay with minimized assay variability. It enables enumeration of cells secreting one, two, three and four analytes and includes low fluorescent plates pre-coated with monoclonal capture antibodies, monoclonal detection antibodies, secondary reagents conjugated to fluorophores, a fluorescence enhancer and an anti-CD28 monoclonal antibody for co-stimulation. A polyclonal activator for human T cells (mAb CD3-2) is supplied as a positive control for cytokine secretion.

Find out more about 4-color FluoroSpot.

Pre-coated plates enable a more rapid assay with minimized variability.

For analysis, we recommend the use of an automated FluoroSpot reader equipped with filters for excitation 380 nm/emission 430 nm (LED380), excitation 490 nm/emission 510 nm (LED490, FITC) for 490-reagent detection, excitation 550 nm/emission 570 nm (LED550, Cy3) for 550-reagent detection and excitation 640 nm/ emission 660 nm (LED640, Cy5) for 640-reagent detection. Filters should have high specificity to avoid bleed-through artifacts.

IFN-γInterferon-γ (IFN-γ, also known as IFN-gamma, IFN-g, IFNg and Interferon-gamma) is produced mainly by activated T cells and NK cells. It is a proinflammatory cytokine that activates macrophages and endothelial cells, but it also regulates immune responses by effecting APC and T and B cells. Production of IFN-γ by helper T cells as well as 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.

IL-17AInterleukin-17A (IL-17A, IL17A) is a potent proinflammatory cytokine produced by activated Th17 (T-helper 17) cells and certain cells belonging to the innate immune system. In mice, IL-17 has been shown to also be produced by activated CD8+ T cells and γδ T cells. Th17 cells play an important role in autoimmune diseases and protection against bacteria and fungi. IL-17A acts on a broad range of cell types to induce expression of cytokines, chemokines and metalloproteinases. As a result, secretion of IL-17A promotes inflammatory responses, which leads to the recruitment of neutrophils, enhancement of antibody production and activation of T cells. Increased expression of IL-17A is seen in autoimmune diseases such as multiple sclerosis and rheumatoid arthritis. It is also associated with asthma, psoriasis, cancer and transplant rejection.

IL-22The cytokine Interleukin-22 (IL-22, IL22) is mainly produced by activated CD4+ T-helper 17 (Th17) and T-helper 22 (Th22) cells. The IL-22 receptor is expressed on non-immune cells, particularly epithelial cells and keratinocytes, and IL-22 promotes innate immune responses versus bacterial infections in these target cells. In addition, Th22 cells have been shown to play a role in the pathophysiology of many human skin diseases.

IL-5Interleukin-5 (IL-5, IL5) is produced by activated CD4+ TH2-type cells, NK cells, mast cells and B cells. It specifically controls the production, activation, and localization of eosinophils.

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