Anti beta catenin

U87 and U373 glioma cell lysates were prepared in 0.1% Triton buffer containing protease and phosphatase inhibitors (Sigma-Alrich) and also were prepared cytoplasmic and nuclear extracts using cell compartment isolation kit (Millipore). Further total protein was quantitated using the BCA Protein Assay Kit (Thermo Scientific, Rockford, IL). Blots were exposed to anti-GSK3 (Millipore), anti-phosphotyrosine GSK3 (Millipore), anti-phospho-ERK1/2 (Thr202/Tyr204) (Millipore), anti-α-tubulin, and anti-beta-catenin (Millipore) and recognized by an HRP-conjugated horse anti-mouse secondary antibody (Santa Cruz Inc); antibodies to detect phospho-AKT1/2/3 (Ser473), total Akt1, hnRNPA1, and Anxa7 (Santa cruz Inc) SF2/ASF1 (Santa Cruz Inc) were recognized by an HRP-conjugated goat anti-rabbit secondary antibody (Santa Cruz Inc.). Enhanced chemiluminescent reagent (GE Healthcare) was added to the membranes according to the manufacturer's protocol and visualized by autoradiography.

Western blotting analysis was performed according to standard methods as previously described.^{12 (link)} The following primary antibodies were used: anti-Sam68 (sc-333, dilution, 1:500; Santa Cruz Biotechnology, Delaware Ave Santa Cruz, CA), anti-beta-catenin, anticyclin D1, anti-p21^cip1, anti-p27^KIP1, anti-p-Rb, antitotal-Rb, anti-p-AKT, antitotal-AKT, anti-c-Myc (1:1000, Millipore, Billerica, MA), anti-P-84, anti-LEF-1 (1:500, Abcam, Cambridge, MA), anti-α-tubulin, anti-TCF-4, anti-LEF1, and anti-GAPDH (1:1000, Sigma, Saint Louis, MO). Nuclear extracts were prepared using the Nuclear Extraction Kit (Active Motif), according to the manufacturer's instructions.