Simple SummaryThe quantification of PD-L1 expression in tumor samples through the tumor proportion score (TPS) and the combined positive score (CPS) conditions provides access to anti-PD-1/PD-L1 immunotherapy in patients with various solids cancers. Reproducibility studies have shown very heterogeneous inter- and intra-pathologist agreements, from poor to excellent ones, in TPS and CPS quantification. We studied the inter- and intra-pathologist agreements in TPS and CPS quantification comparing single PD-L1 immunohistochemistry (S-IHC) and double immunohistochemistry (D-IHC) combining PD-L1 staining and tumor nuclear markers, trying to improve the distinction between tumor cells and immune cells necessary to TPS and CPS calculations. Our study concluded in excellent (TPS) to good (CPS) inter- and intra-pathologist agreements with both S-IHC and D-IHC with slightly higher intraclass correlation coefficients using D-IHC. D-IHC could help the pathologists to quantify PD-L1 expression through TPS and CPS for subsequent therapeutic choices in patients with advanced cancers.AbstractWe studied the pathologists’ agreements in quantifying PD-L1 expression through the tumor proportion score (TPS) and the combined positive score (CPS) using single PD-L1 immunohistochemistry (S-IHC) and double immunohistochemistry (D-IHC) combining PD-L1 staining and tumor cell markers. S-IHC and D-IHC were applied to 15 cancer samples to generate 60 digital IHC slides (30 whole slides images and 30 regions of interest of 1 mm2) for PD-L1 expression quantification using both TPS and CPS, twice by four pathologists. Agreements were estimated calculating intraclass correlation coefficients (ICC). Both S-IHC and D-IHC slides analyses resulted in excellent (for TPS, ICC > 0.9) to good (for CPS, ICC > 0.75) inter- and intra-pathologist agreements with slightly higher ICC with D-IHC than with S-IHC. S-IHC resulted in higher TPS and CPS than D-IHC (+5.6 and +6.1 mean differences, respectively). High reproducibility in the quantification of PD-L1 expression is attainable using S-IHC and D-IHC.