Immunostaining Protocol for Bcl9 and β-Catenin

3-Aminopropyltriethoxysilane (APS), bovine serum albumin (BSA, essentially fatty acid- and globulin-free), 30% Brij^® L23 solution, normal goat IgG and rabbit IgG were purchased from Sigma-Aldrich (St. Louis, MO, USA). Normal mouse IgG was from Dako (Glostrup, Denmark). Mouse monoclonal anti-human Bcl9 (BMR00368; 2.0 μg/ml) was a gift from Bio Matrix Research Inc. (Nagareyama, Chiba, Japan) and rabbit polyclonal anti-human Bcl9 (ab37305; 5.0 μg/ml) was purchased from Abcam (Cambridge, MA, USA) [12 (link), 30 ]. Mouse monoclonal anti-β-catenin (CTNNB1, UM500015; 1.33 μg/ml) was purchased from OriGene (Rockville, MD, USA). Horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG (AP308P; 10.0 μg/ml) and HRP-conjugated goat anti-rabbit IgG (AP307P; 10.0 μg/ml) antibodies were from Millipore (Temecula, CA, USA). 3,3'-Diaminobenzidine-4HCl (DAB) was from Dojin Chemical Co. (Kumamoto, Japan). Permount was purchased from Thermo Fisher Scientific (Hudson, NH, USA). All other reagents used in this study were from Wako Pure Chemicals (Osaka, Japan).