Abstract
IL-17 plays a key role to defend infections, but also in chronic inflammatory disorders. We developed tools for the enrichment and detection of IL-17-producing cells.
After T cell activation IL-17 was detected on the surface of IL-17-producing cells after first labeling the cells with a bispecific CD45-IL-17 antibody affinity matrix. Cells were again cultured for 45 min to allow IL-17 secretion. Secreted IL-17 was specifically caught onto the affinity matrix of IL-17-producing cells. Subsequently IL-17-secreting cells were fluorescently stained with a second IL-17 antibody or additionally labeled with magnetic beads.
Stimulation of mouse spleen cells with PMA/ionomycin showed a maximal IL-17 secretion after 4 h in <0.5% of CD4 and CD8 T cells, but in around 20% g/d T cells and 7% NKT cells. In contrast in human PBMC the IL-17 secretion was mainly restricted to CD4 T cells, with a maximum of 0.1-1% of them producing IL-17 after 4 h activation with the artificial superantigen CytoStim. Magnetic enrichment yields in IL-17+ cell populations with a purity of >90%. Nearly all of the enriched IL-17+ CD4 T cells co-expressed CD154, confirming the specificity of the detection and enrichment method.
The IL-17 secretion assays allow detection of IL-17-secreting cells and provides highly enriched populations of viable IL-17-producing cells to investigate its function and the role of IL-17 in host defense and disease development.
