Western blot and immunocytochemistry (ICC) results assessing HES1 expression in canine osteosarcoma cells. (A) Western blot analysis of whole cell (W), nuclear (N) and cytoplasmic (C) fractions of canine osteosarcoma Abrams and Gracie cell lines. A 30 kDa band (HES1) is present in whole cell and enriched in extracted nuclear lysates. Larger non-specific bands are enriched in the cytoplasmic fractions. Equal amounts of total protein were loaded in each lane. (B) ICC shows nuclear staining for HES1 in canine OSA cells (Gracie). Panel on the right is the secondary-only negative control. Photomicrographs were taken at 20× and 100× (oil, inset) magnification; haemotoxylin counterstain.