The recombinant expression vectors are designed to express human engineered T cell receptor (TCR) activator and mouse PD-L1 (GenBank Accession #NM_021893) in mammalian cells. The transfected cells can be used in conjunction with mouse PD-1/NFAT Reporter/Jurkat T cells (BPS Bioscience #79762) to study the interactions of PD-1 with PD-L1 ligand in a cellular context and to screen for modulators of this signaling pathway. Functional Validation and Assay Performance: In this assay, mouse PD-1/NFAT Reporter/Jurkat T cells are used as effector cells; HEK293 cells transiently transfected with the PD-L1 vector over-express mouse PD-L1 and an engineered T cell receptor (TCR) activator and are used as target cells. When these two cells are co-cultivated, TCR complexes on effector cells are activated by TCR activator on target cells, resulting in expression of the NFAT luciferase reporter. However, PD1 and PD-L1 ligation prevents TCR activation and suppresses the NFAT-responsive luciferase activity. This inhibition can be specifically reversed by anti-PD1 or anti-PD-L1 antibodies. Mouse PD1/PD-L1 neutralizing antibodies block PD1:PD-L1 interaction and promote T cell activation, resulting in reactivation of the NFAT responsive luciferase reporter.
