Immunohistochemical staining (IHC)

IHC was used to evaluate the expression of ZNF384 in HCC tissues and paired noncancerous tissues. Paraffin-embedded sections were provided by the department of pathology. First, these sections were deparaffinized and rehydrated. For antigen retrieval, the sections were immersed in 10 mM citrate buffer (pH 6.0) and boiled for 10 min in a microwave oven. Then, endogenous peroxidase activity was blocked with 3% hydrogen peroxide for 10 min. Non-specific binding sites were blocked with 5% normal goat serum for 30 min. The sections were incubated with an antibody against ZNF384 (1:100, Atlas Antibodies, cat#HPA004051) overnight at 4 °C. The sections were then incubated with the secondary antibody, and the expression of ZNF384 in the tissues was observed via microscopy after DAB staining and haematoxylin staining. The score was evaluated by two pathologists blinded to the clinicopathological data using a German immunoreactivity score²⁹.