Immunohistochemical analysis of HDAC3 in human colon cancer. Samples were incubated in HDAC3 polyclonal antibody (Product # PA1-41039) using a dilution of 1:50. Bond Rx autostainer (Leica Biosystems). The assay involved 20 minutes of heat induced antigen retrieval (HIER) using 10mM sodium citrate buffer (pH 6.0) and endogenous peroxidase quenching with peroxide block. The sections were incubated with primary antibody for 30 minutes and Bond Polymer Refine Detection (Leica Biosystems) with DAB was used for signal development followed by counterstaining with hematoxylin. Whole slide scanning and capturing of representative images was performed using Aperio AT2 (Leica Biosystems). Nuclear staining of HDAC3 was observed. Staining was performed by Histowiz.