Multiplexed G-Protein–Coupled Receptor Ca2+ Flux Assays for High-Throughput Screening

An early drug discovery approach focusing on gene families can benefit fromstrategies that exploit common signalingmechanisms to more effectively identify and characterize novel chemical lead structures. Multiplexing, defined as the screening of multiple targets within the same experiment, is an example of this strategy. Here, the authors describe a technique that allows multiplexing of a common assay type used to study G-protein–coupled receptors: changes in intracellular Ca2+ levels as measured by Molecular Device's fluorometric imaging plate reader (FLIPR®). The multiplexed FLIPR assays showed the expected pharmacological properties of single assays, with good reproducibility and Z' factors. The authors used them to screen large compound libraries in 2 multiplexed assay designs. The 1st used a single-cell line expressing 2 different receptors and the 2nd amixture of 2 cell lines of the same type each expressing distinct receptors. Screening using thesemultiplexed assays produced significant savings in reagents, time, and human resources and allowed the authors to quickly identify specific and selective hits. (Journal of Biomolecular Screening 2005:780-787)