Reagent Details

Applications

For ligand binding, fluorescence imaging and high content analysis, kinetic analysis and cell sorting at adenosine A1 / A2A / A3 receptors use solutions up to 100 nM.

Pharmacological Validation

The CA200645 ligand was shown to antagonize the activity of the adenosine receptor agonist, NECA, in three separate recombinant CHO cell lines expressing the human A1, A2A or A3 receptor and a cyclic AMP-responsive secreted placental alkaline phosphatase (SPAP) reporter gene.

The cyclic AMP-induced expression of SPAP was measured under basal and forskolin-stimulated (maximal) conditions. Addition of CA200645 to the basal or forskolin-stimulated cells did not significantly alter basal and stimulated SPAP levels, demonstrating that CA200645 has no intrinsic agonist activity.

To determine the apparent KD for CA200645, cells were treated with varying concentrations of NECA alone, or in the presence of 1µM CA200645, and the cyclic AMP-induced expression of SPAP measured.

The apparent KD at A1, A2A and A3 receptors was calculated from the rightward shift of the agonist response curve in the presence of CA200645, compared to the response curve for the agonist alone, for each receptor-expressing cell line.