Binding assay optimization

The development of maraviroc (21), much like other chemokine receptor antagonists, started with a high-throughput screen employing a competition binding assay and led to the hit compound UK-107,543. Chemical optimization of this compound led to the development candidate UK-427,857 during this optimization phase, a parallel characterization of the compounds was performed... 

Each reaction contains 1 x 10 neutrophils/mL of HEPES-buffered Hank s BSS. In performing binding assays, it is important to work with a cell density where cell clumping is not occurring during the reaction. A cell density of 1 x 10 neutrophils/mL appears to be an optimal concentration for this assay. 

SPA has been developed as a receptor binding assay using molecules prepared from a number of different sources ranging from crude tissue preparations to soluble purified proteins. The method, which is applicable to both 3H and 125I ligands, provides a number of factors and must be carefully optimized in the assay design and development. 

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