A Cellular Assay for Measuring the Inhibition of Glycogen Synthase Kinase-3 via the Accumulation of β -Catenin in Chinese Hamster Ovary Clone K1 Cells
AbstractGlycogen synthase kinase-3 (GSK3) is a serine-threonine protein kinase that exists as two isozymes, GSK3 α and GSK3 β . It plays important roles in regulating cell structure, function, and survival, and dysregulation of its function is linked to disorders such as Alzheimer's disease and type II diabetes. In resting cells, GSK3 is active and regulates the function of many downstream targets, including β -catenin. We describe the development of a cell-based assay designed to measure the activity of GSK3 by directly measuring the accumulation of β -catenin in Chinese hamster ovary clone K1 (CHOK1) cells. β -Catenin levels were assessed using an antibody-based staining protocol with a luminometric readout. The assay is set up in a 96-well format. The use of GSK3 inhibitors demonstrated that this assay could be used to compare the effects of various small molecules on GSK3 inhibition in CHOK1 cells.