Me1004b

Melanoma tumor arrays (US Biomax, Inc, ME1004b and ME1004e) were immunostained with primary antibody 1:50 anti-pS209APT1 and 1:200 anti-APT1. Staining procedure was performed as described in Walter et al. (2017) (link). Images were obtained using 20X objective.

Human Malignant melanoma, metastatic malignant melanoma and nevus tissue array (ME1004a and ME1004b, US Biomax, Rockville, MD, USA) were used for immunohistochemistry staining. Antigen retrieval was achieved by microwaving arrays immersed in 1 liter Tris-EDTA buffer (10 mM Tris base, 1 mM EDTA solution, 0.05% Tween 20, pH 9.0) for 30 min. Tissue arrays were treated with 1% H₂O₂ in water for 15 min and blocked in 10% BSA in Tris-buffered saline for 2 h at room temperature. Anti-Mov10 (1:200) in 10% BSA/Tris-buffered saline or anti-FASN (1:500) in 1% BSA/Tris-buffered saline were applied and incubated at 4 °C overnight. Then, arrays were incubated with biotinylated anti-rabbit (1:500, Vector Lab.) for Mov10 or rabbit anti-mouse (1:200, Vector Lab., Burlingame, CA, USA) for FASN for 1 h at room temperature followed by being incubated with VECTASTAIN ABC kit (PK-4000, Vector Lab.) for 30 min. Mov10 and FANS were visualized by Vector Novared substrate kit (SK-4800, Vector Lab.). The arrays were counterstained with hematoxylin for 1.5 min.