FluoroSpot detection reagent replacement
We have improved the detection reagents used in our FluoroSpot assays. As of January 2, 2018, the new detection reagents are included in our FluoroSpot kits. The implemented changes only involve the tag/anti-tag reagents and do not affect the monoclonal antibodies or the biotin/streptavidin detection system.
Epitope tag/anti-tag reagents are used as an amplification step in FluoroSpot assays. By replacing FITC/anti-FITC-490 with BAM/anti-BAM-490, the detection of human and monkey IFN-γ is improved. As a consequence of this change, the detection reagents for IL-5, IL-17A and TNF-α in triple FluoroSpot kits have been updated from BAM/anti-BAM-640 to WASP/anti-WASP-640. The quality and performance of the WASP-tagged detection system is fully comparable to the previously used BAM-tagged system. Please note that it will not be possible to mix/combine reagents from old kits with new kits using the new tag-system.
We always aim at developing and supplying high quality FluoroSpot products with the best monoclonal antibodies, tag/anti-tag reagents and fluorophores. If you have any questions regarding the changed detection reagents or our FluoroSpot assays, we encourage you to contact us and welcome any comments you may have.
Example: Human IFN-γ/IL-22/IL-17A triple FluoroSpot assay setup. In the new system, IFN-γ is detected by the same detection mAb as in the old system, but the mAb is now conjugated to a BAM tag instead of a FITC tag. This BAM tag is detected by anti-BAM-490. IL-17A is also detected by the same detection mAb as before, but the mAb is now conjugated to a WASP tag instead of a BAM tag. This WASP tag is detected by anti-WASP-640. No changes have been made to any of the capture antibodies or to the biotinylated IL-22 detection system. Updated components are indicated in red.
Sample data: Human IFN-γ/IL-22/IL-17A triple FluoroSpot. Human PBMC (250,000 cells/well) were stimulated with PPD overnight and the number of cells secreting IFN-γ, IL-22 and IL-17A was analyzed by FluoroSpot. Note that the detection of IFN-γ is improved with the new system. There were no changes observed in spot numbers in unstimulated PBMC. Wells for each analyte are read and analyzed with the same reader settings.
Sample data: Human IFN-γ FluoroSpot. Human PBMC (250,000 cells/well) were stimulated with CEF overnight and the number of cells secreting IFN-γ was analyzed by FluoroSpot. Wells are read and analyzed with the same reader settings.