Immunohistochemical and Immunofluorescence Analysis of PABPC1 and IFI27 Expression

IHC staining was performed using the Envision Labeled Peroxidase System (Dako, Carpinteria, CA, USA) as described previously [19 (link)]. PABPC1 and IFI27 expression were analyzed based on the proportion and intensity of positively-stained tumor cells. The scores for intensity and fraction of positive cells were multiplied; scores of 0–4 were defined as low expression, and scores of 6–12 were defined as high expression. Immunofluorescence was performed as previously described [19 (link)]. Briefly, cells were first fixed in 3% paraformaldehyde, and then permeabilized with 0.5% Triton X-100. The cells were blocked with 5% FBS in PBS, followed by incubation with primary antibody, fluorescent secondary antibody (Invitrogen) and counterstained with DAPI (Beyotime) were then incubated with cells to visualize the targeted proteins and nuclei. The data were then analyzed by fluorescence microscopy.