A T‐cell reporter platform for high‐throughput and reliable investigation of TCR function and biology

OBJECTIVE: Transgenic re‐expression enables unbiased investigation of T‐cell receptor (TCR)‐intrinsic characteristics detached from its original cellular context. Recent advancements in TCR repertoire sequencing and development of protocols for direct cloning of full TCRαβ constructs now facilitate large‐scale transgenic TCR re‐expression. Together, this offers unprecedented opportunities for the screening of TCRs for basic research as well as clinical use. However, the functional characterisation of re‐expressed TCRs is still a complicated and laborious matter. Here, we propose a Jurkat‐based triple parameter TCR signalling reporter endogenous TCR knockout cellular platform (TPR(KO)) that offers an unbiased, easy read‐out of TCR functionality and facilitates high‐throughput screening approaches. METHODS: As a proof‐of‐concept, we transgenically re‐expressed 59 human cytomegalovirus‐specific TCRs and systematically investigated and compared TCR function in TPR(KO) cells versus primary human T cells. RESULTS: We demonstrate that the TPR(KO) cell line facilitates antigen‐HLA specificity screening via sensitive peptide‐MHC‐multimer staining, which was highly comparable to primary T cells. Also, TCR functional avidity in TPR(KO) cells was strongly correlating to primary T cells, especially in the absence of CD8αβ co‐receptor. CONCLUSION: Overall, our data show that the TPR(KO) cell lines can serve as a surrogate of primary human T cells for standardised and high‐throughput investigation of TCR biology.