Quantifying Tumor Immune Profiles

Four-micrometer-thick tumor sections were stained for PD-L1 (clone 22C3, DAKO, Agilent, CA, USA), CD4 (clone BP6028; Biolynx, Hangzhou, China), CD8 (clone: EP334, Abcam, Cambrige, UK) using BenchMark automated immunostainer (Ventana, AZ, USA). Staining was evaluated in a blinded fashion by the pathologists. Scoring was assessed based on the proportion of positive cells among nucleated cells in the ROI. The positive cells were defined as tumor cells displaying membranous staining of PD-L1. The PD-L1 expression at MPC was quantified as the proportion of PD-L1-positive tumor cells in total tumor cells within the MPC area. The tumor proportion score (TPS) is a PD-L1 measurement that has been applied in clinical trials and in clinic of lung cancer (20 (link)). The percentage of CD4+ and CD8+ T cell in the peritumor region was assessed based on the proportion of positive T cells among nucleated cells in the peritumoral area, which was described in previous studies (21 (link), 22 (link)).