Immunohistochemical Analysis of Xenograft Tissues

Harvested xenografts were embedded in paraffin before the preparation of 5 μM thick tissue sections on slides that were either stained with H&E, mouse anti-Kaiso 12H monoclonal (1:800)^{50 (link)} or p-Smad2 (CST-138D4; 1:200 for MDA-231 xenografts and 1:50 for Hs578T xenografts) primary antibodies overnight at 4 °C. Briefly, xenograft tissues were dewaxed by warming on a slide warmer at 60 °C for 20 min followed by immersion in xylenes 3 × 5 min. All other steps were performed as previously described,^{31 (link)} but we utilized PBS in place of TBS. Images were obtained using the Aperio Slide scanner (Leica Biosystems, Concord, ON, Canada).