Invitrogen™ CellSensor™ LEF/TCF-blaFreeStyle™ 293F Cell Line

Description

• Wnt signaling via beta - catenin plays a central role in development and homeostasis
• This pathway is invariably disrupted in colorectal tumors and commonly affected by mutation in other cancers
• Binding of the Wnt protein to the Frizzled transmembrane receptors initiates a signaling cascade that activates the Dishevelled protein, which then inhibits the serine/threonine kinase Glycogen Synthase - 3 Beta (GSK - 3B)
• This signal leads to functional inactivation and dissociation of a multiprotein beta - catenin destruction complex made up of the tumor suppressor protein Adenomatous Polyposis Coli (APC), GSK - 3B, and a scaffold of Axin
• This results in dephosphorylation and dissociation of beta - catenin
• Unphosphorylated beta - catenin is stabilized and accumulates in the cytoplasm of the cell
• Beta - catenin then associates with the Lymphoid Enhancer Factor (LEF)/T - Cell Factor (TCF) family of transcription factors in the nucleus, leading to transcription and expression of target genes, such as c - Myc, c - jun, Fra, and cyclin D1
• The CellSensor™ LEF/TCF - bla FreeStyle™ 293 Cell Line contains a beta - lactamase reporter gene under control of the LEF/TCF stably integrated into FreeStyle™ 293 cells
• This cell line is validated for EC50 and Z' - factor under optimized conditions using mWnt3a
• This cell line has also been tested under various experimental conditions, including DMSO concentration, cell number, stimulation time, and substrate loading time
• CellSensor™ LEF/TCF - bla FreeStyle™ 293F cells were stimulated with mWnt3a over the indicated concentration range in a 384 - well format
• Cells were incubated for 5 hrs with agonist and 0.5% DMSO and then combined with LiveBLAzer™ - FRET B/G Substrate (CCF4 - AM) for 2.5 hrs
• Fluorescence emission values at 460 nm and 530 nm were obtained using a standard fluorescence plate reader
• The 460/530 ratios are shown plotted against the indicated concentrations of mWnt3a
• Academic and non-profit customers, please inquire for special pricing.