IHC

Paraffin sections of tissue samples obtained from reduction mammaplasties, breast cancer biopsies, breast tumor resections, or patient-derived xenografts (PDX; see “PDX implantation”) were deparaffinized first and then antigen retrieval was performed in 0.01M citrate buffer (pH 6.0) or EDTA buffer (pH 8.0). After 3% hydrogen peroxide treatment and blocking, a primary antibody for MUC1 (Abcam, #ab45167, 1:500), HER2 (Abcam, #ab134182, 1:300), ERα (Abcam, #ab32063, 1:200), or PR (Abcam, #ab32085, 1:100) was incubated overnight at 4°C and the signaling was amplified via incubation for 1 hour at room temperature with HRP-conjugated secondary antibodies from an anti-mouse/rabbit IHC Secondary Antibody Kit (Dako, # GK500710). After washing, DAB peroxidase substrate (Dako, # GK500710) was dropped on top of the slides and visualized under a microscope (Olympus, BX-63) for brown staining. The reaction was stopped by dipping slides into a large amount of PBS. Images were taken with Olympus BX-63 microscope.