Immunohistochemical Analysis of c-Myc and AR

Immunohistochemistry was performed as previously described [22 (link)]. Briefly, heat-induced antigen retrieval was carried out in a steam chamber followed by washing in Tris-buffered saline. Endogenous peroxidases were inactivated with Bloxall (Vector labs), and sections were blocked with Dako blocking buffer (Agilent Technologies), incubated with primary antibodies (anti-c-Myc, anti-AR), washed, and then incubated with horse radish peroxidase-conjugated secondary antibodies. Chromagen was developed with DAB solution (Vector Labs) and counterstained with Meyer’s hematoxylin. Primary antibodies used were anti-cMyc (Abcam, clone 769, 1:1000), anti-Androgen Receptor (Santa Cruz, clone N-20, 1:500), and anti-CK8 (Covance, clone 1E8, 1:1000).