Immunohistochemical Analysis of EHMT1 in Lung Cancer

An EnVision⁺ kit/HRP kit (Dako, Carpinteria, CA, USA) was used. Paraffin‐embedded sections of lung tumor specimens were processed in a microwave (90 °C) with antigen retrieval solution (pH 9) (S2367; Dako), treated with a peroxidase‐blocking reagent, and then treated with a protein‐blocking reagent (K130, X0909; Dako). Tissue sections were incubated first with a rabbit anti‐EHMT1 antibody (BIO MATRIX RESEARCH, Chiba, Japan) and then with an HRP‐conjugated secondary antibody (Dako). Immunoreactivity was visualized with a chromogenic substrate (Liquid DAB Chromogen; Dako). Finally, tissue specimens were stained with Mayer’s hematoxylin solution (Hematoxylin QS; Vector Laboratories, Burlingame, CA, USA) for 20 s to distinguish nuclei from the cytoplasm. Human lung cancer tissues were purchased from BioChain [11 (link)].