Figure 4. Effects of EGF on the induction of GPR30 by immunohistochemistry.

C57 wild type, αERKO and βERKO mice were either injected with 25 ng EGF or vehicle subcutaneously daily for 5 days. The glands were dissected, fixed in formalin and 5 micron sections were processed for immunohistochemical staining. The sections were incubated with blocking reagent followed by GPR30 primary antibody and biotinylated secondary antibody. Antigen specific staining was visualized using DAB (brown) as a chromogen and nuclear counterstaining was done using hematoxylin (blue). Negative controls (sections not incubated with GPR30 antibodies) are shown as an inset for each condition. Results show that EGF induces GPR30 (brown) in the mammary glands of αERKO mice. A. Vehicle, Control; B. Vehicle, αERKO; C Vehicle, βERKO; D. EGF, Control; E. EGF, αERKO; F. EGF, βERKO.