Technical brief: Innate Immunity Testing

Our basic test program for innate immunity testing quantifies the extent to which ligands specific for the nine known human Toll-like receptors (TLR) stimulate peripheral blood mononuclear cells (PBMC) isolated from individuals. The degree of stimulation is quantified via the amount of Tumor Necrosis Factor-alpha (TNF-alpha) secreted into the culture supernatants.

An example for a typical experiment is given here (click on the image to enlarge view):

PBMC were isolated from the blood of two human volunteers and cultured in the presence of distinct ligands for toll-like receptors (TLR-1 to -9). After a given interval of time the culture supernatants were harvested and tested for the amount of secreted TNF-alpha by enzyme-linked immunosorbent assay (ELISA).

The ligands and the corresponding TLRs are shown on the X-axis. The Y-axis provides the read-out parameter, i.e. amount of TNF-alpha in the culture supernatants. The left-most sample in both diagrams represents TNF-alpha secretion the unstimulated control culture.

Strongest responses were found for the TLR-4 ligand lipopolysaccharide (LPS) and the TLR 8 ligand single-stranded RNA (ssRNA40). The weak-responding TLR 3, 7 and 9 stimulations are again highlighted in the right hand diagram with a differently-scaled Y-axis in order to demonstrate the above-background response.

Please note, that this basic ex vivo assay design can be adapted to the specific requirements of a given project – be it for immune screening purposes in clinical trials or be it for mode-of-action or PD measurements in compound development.