Automated IHC Profiling of PD-1/PD-L1 and MMR

An automated staining platform (Ventana Medical Systems, Inc.) was used to detect PD-1/PD-L1 protein expression by IHC: the SP142 Ab (laboratory developed test) was used to stain for PD-L1 expression on TCs of prostate, colorectal, ovarian, and pancreatic cancers, with a positive threshold of ≥2+ intensity and ≥5% cell stained; the SP142 Ab (Ventana) was used to stain for PD-L1 on immune cells of breast cancer, with a positive threshold of ≥1% of cells stained; the 22c3 Ab was used in NSCLC and ovarian cancers, with a positive threshold of tumor proportion score (TPS) ≥1 and combined positive score (CPS) ≥1, respectively; the 28-8 Ab was used to stain for PD-L1 expression on TCs for NSCLC, with a positive threshold of ≥1+ intensity and ≥1% cell stained; and the PD-1 Ab was used for ovarian cancer, with a positive threshold of ≥1% cells stained. Benign tonsil samples served as a positive control. MMR protein expression was tested by IHC (Ventana Medical Systems) using Ab clones for the four mismatch repair proteins [MLH1, M1 Ab; MSH2, G2191129 Ab; MSH6, 44 Ab; and PMS2 (Abcam, catalog no. ab203457, RRID:AB 2889230), EPR3947 Ab (Ventana Medical Systems, Inc.)]. The complete absence of protein expression of any of the four proteins tested (0+ in 100% of cells) was considered deficient MMR as described previously (21 (link)).