Phospho- and total β-Catenin detection as a readout of the canonical Wnt pathway
Cisbio's cell-based homogeneous HTRF® assays, phospho-β-Catenin (Thr41/Ser37/Ser33) and total β-Catenin, enable a canonical Wnt pathway readout. The phospho-β-Catenin assay provides the quantitative detection of GSK3 β-induced phosphorylation of β-Catenin on Thr41/Ser37/Ser33, which represents the inactive form of the protein degraded by the proteasome. Cytosolic total β-Catenin is an important co-transcriptional activator of many significant genes involved in embryonic development, adult homeostasis and cell proliferation. The total β-Catenin assay serves as a normalization control of the phosphorylated form and facilitates analysis of the steady state level between the 2 pools of β-Catenin – phosphorylated inactive and the non-phosphorylated forms – with their different cellular functions. Membrane-associated β –Catenin involved in cadherin-based adherens junctions cannot be discriminated by the total assay. The phospho- β-Catenin assay is thus suitable for cell-based screening of GSK3β inhibitors or other modulators of the Wnt pathway. The simple add-and-read protocol eliminates all wash steps for a faster analysis and easy miniaturization while maintaining a high-quality specific output, and offers enhanced convenience over ELISA or WB.