Evaluation of electrochemiluminescent technology for inhibitors of granulocyte colony-stimulating factor receptor binding

An electrochemiluminescent (ECL) assay was developed to identify compounds that inhibit the interaction of granulocyte colony-stimulating factor (GCSF ) with its recombinant human receptor. The ECL technology uses a tris(bipyr idine) chelate of ruthenium, which, in the presence of excess tripropylamin e, undergoes a redox reaction cycle to produce light, Paramagnetic beads wi th primary antibody were coated with secondary anti-GCSF receptor antibody, which were then bound with GCSF receptor. These samples were incubated wit h ruthenylated GCSF in the presence and absence of test compounds. The bead density, receptor and ligand concentrations, and incubation time were opti mized in the assay. A set of mixed compound plates was screened to examine the feasibility of using this technology in high throughput screening. The results from this format were found to be comparable to the assay performed using a time-resolved fluorescence format.