Physical cognate pairing of mouseTCR alpha-beta VDJ-rearrangements with phenotype directly from FACS sorted cells

Abstract

Abstract Phenotypic characterization of single cells paired with immune receptor information can provide valuable insight into T cell function, which may be beneficial for the development of personalized therapies. In order to comprehensively analyze T cell function and specificity, it is necessary to integrate antigen receptor sequencing with phenotypic markers at the single cell level. We therefore developed a sensitive technology called iPair+™ that enables the simultaneous amplification and sequencing of paired mouse TCR alpha and beta chains with functional gene markers in a single, sorted immune cell via a combination of flow cytometry, multiplex PCR, and next generation sequencing. We utilized iPair+™ to study CD4 and CD8 T cells isolated from treated and non-treated mice from a breast cancer model. CDR3 diversity, clonality, and frequency were analyzed together with 80+ phenotypic genes. When cells isolated from both treated and non-treated mice were compared, we found gene expression levels changed significantly with treatment. Our data demonstrates that genes that were upregulated prior to treatment were associated with cancer growth, while genes upregulated after treatment were associated with T cell activation and function in cancer repression or growth inhibition. Our study indicates that iPair+ technology could reveal important insights into T cell function, clonal development, and immunotherapy response during treatment.