Alk clone d5f3

For IHC staining, slides were deparaffinized and steamed for either 30 minutes in 10 mmol/L sodium citrate (pH  =  6, Vector Labs) or for 45 minutes in Target Retrieval Solution (Dako). Primary antibodies and dilutions used were as follows: ALK (clone D5F3, Cell Signaling Technology, 3633T, 1:100), AR (Cell Signaling Technology, 5153T, 1:100), synaptophysin (Thermo Fisher, RM9111S, 1:80), and NKX3.1 (Thermo Fisher, 5082788, 1:50). Immunocomplexes were detected using the UltraVision Quanto Detection System with DAB as the chromogen (Thermo Fisher, TL060QHD). Tissue sections were counterstained with hematoxylin and slides were digitized on a Ventana DP 200 Slide Scanner (Roche). Immunoreactivity was scored in a blinded manner by two pathologists (M. Roudier and M.C. Haffner) whereby the optical density level (“0” for no brown color, “1” for faint and fine brown chromogen deposition, and “2” for prominent chromogen deposition) was multiplied by the percentage of cells at each staining level, resulting in a total score range of 0 to 200. The final score for each sample was the average of two duplicated tissue cores (20 (link)).