493 Distinct antibody clones detect PD-1 checkpoint expression and block PD-L1 interactions on live murine melanoma cells

Monoclonal antibodies (abs) targeting the programmed cell death 1 (PD-1) immune checkpoint pathway have revolutionized tumor therapy. Because T-cell-directed PD-1 blockade boosts immunity, anti-PD-1 abs been developed for examining T-cell-PD-1 functions. More recently, expression has also reported directly on cancer cells of various etiology, including in melanoma. Nevertheless, there is a paucity studies validating ab clone utility specific assay types characterizing cell-intrinsic PD-1. Here, we demonstrate reactivity several anti-murine clones and recombinant PD-L1 with live B16-F10 melanoma cells, using multiple independent methodologies, positive negative PD-1-specific controls, PD-1-overexpressing knockout cells. Flow cytometric analyses two separate clones, 29F.1A12 RMP1-30, revealed surface protein which was corroborated by marked enrichment gene (Pdcd1) expression. Immunoblotting, immunoprecipitation, mass spectrometric sequencing confirmed Recombinant recognized cell-expressed PD-1, fully abrogated PD-1:PD-L1 binding. Together, our data provides lines evidence establishing validates systems cell-directed investigations.