CRISPR/Cas9 edited, singly expressed HLA class II cells for the study of HLA-DQ immunogenicity

Our overarching goal is to achieve stable transduction of either HLA-DQB1 or HLA-DRB1 into a multiple knockout CRISPR/Cas9 HLA-class II genome edited cell for the purpose of studying HLA-DQ versus HLA-DR specific immune processes including antigen processing and presentation and T cell activation pathways. To this aim we will: 1. Stably rescue either HLA-DQ or HLA-DR expression in HLA class II knockout cells 2. Demonstrate the ability of the rescued cells to restore T cell allorecognition 3. Demonstrate the ability of the rescued cells to bind either HLA-DQ or HLA-DR specific antibodies The ability to demonstrate stable, genome edited, 3D appropriate conformation expression of isolated HLA-DQ or HLA-DR molecules, with physiologic characteristics, will provide the required infrastructure to proceed with functional studies interrogating pathways activated by either of these molecules as they participate in different immune processes