FluoroSpot kit for enumeration of cells secreting TNF-α/GM-CSF

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 TNF-α/GM-CSF FluoroSpot kit enables convenient enumeration of cells secreting either one or both analytes. The kit contains low fluorescent plates, matched quantities of monoclonal capture and detection antibodies, secondary reagents conjugated to fluorophores and a fluorescence enhancer. LPS is included.

The assay is recommended for the analysis of macrophages, monocytes and dendritic cells. These kits have not been evaluated for T-cells.

For analysis, we recommend the use of an automated FluoroSpot reader equipped with filters for FITC (excitation 490 nm/emission 510 nm) and Cy3 (excitation 550 nm/emission 570 nm). Filters should have high specificity to avoid bleed-through artifacts. Double secreting cells are identified by a computerized overlay.

Analyte Information

GM-CSF
Granulocyte macrophage-colony stimulating factor (GM-CSF) can be produced by several cell types among others activated T cells, macrophages, endothelial cells and stromal cells. GM-CSF stimulates the survival and functional activities of neutrophils, eosinophils, and monocyte-macrophages. It also stimulates the differentiation and proliferation of hematological progenitors.

TNF-α
Tumor necrosis factor-α (TNF-α, also known as TNF-alpha, TNF-a, TNFa, and Tumor necrosis factor-alpha) is produced by many different cell types e.g. monocytes, macrophages, T cells and B cells. Among the many effects of TNF-α, are protection against bacterial infection, cell growth modulation, immune system regulation and involvement in septic shock.

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