Screening self-peptides for recognition by mouse alloreactive CD8+ T cells using direct ex vivo multimer staining

• Published in: STAR Protocols
• Main Authors: Eric T. Son, Moumita Paul-Heng, Mario Leong, Chuanmin Wang, Alexandra E. Hill, Martina Denkova, Anthony W. Purcell, Nicole A. Mifsud, Alexandra F. Sharland
• Format: Article
• Language: English
• Published: Elsevier 2023-03-01
• Subjects: Cell Biology; Cell isolation; Immunology; Model Organisms; Molecular Biology; Antibody
• Online Access: http://www.sciencedirect.com/science/article/pii/S2666166722008231
• ISSN: 2666-1667

Summary: Here, we present a protocol to identify immunogenic self-peptide/allogeneic major histocompatibility complex (MHC) epitopes. We describe the generation of enriched alloreactive CD8+ T cells by priming mice with a skin graft expressing the allogeneic MHC class I molecule of interest, followed by boosting with a liver-specific AAV vector encoding the heavy chain of that donor MHC allomorph. We then use a peptide-exchange approach to assemble a range of peptide-MHC (pMHC) multimers for measuring recognition of the various epitopes by these alloreactive T cells.For complete details on the use and execution of this protocol, please refer to Son et al. (2021).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.