High-throughput screening for the identification of small-molecule inhibitors of retinoblastoma protein phosphorylation in cells.

摘要: The tumor suppressor protein, pRb, regulates progression through the G1 phase of cell cycle by its ability to bind and regulate activity a variety transcription factors. This function pRb is disabled phosphorylation cyclin-dependent kinase (CDK) family serine/threonine kinases. In many human cancers, genetic alteration such as loss CDK inhibitor deregulated cyclin expression leads inappropriate hence inactivation this suppressor. Identification cell-permeable small molecules that block in these tumors could therefore lead development an effective anticancer treatment. As result, we have developed high-throughput assay detect changes level cells. Signal detection time-resolved fluorescence-based cellular immunosorbant on fixed monolayer comprises mouse monoclonal antibody recognizes phosphorylated form serine 608 known site phosphorylation, Europium-labeled secondary for signal detection. reproducible amenable automation has been used screen 2000 compounds search will phosphorylation.