Differential impact of cryopreservation on CD4+ and CD8+ T cell repertoires and clonotypic expansion

ABSTRACT: PBMC cryopreservation is a fundamental technique in immunology research and clinical applications, which is used to assess T cell responses to antigens across disease settings. However, its differential impact on CD4+ and CD8+ T cell repertoire dynamics has not been studied and is of high relevance owing to the increasing number of T cell assays using clonotypic expansions as a functional readout. In this study, we systematically compared the effects of cryopreservation on CD4+ and CD8+ T cell clonotypic expansions using a functional T cell receptor (TCR) sequencing assay. We found that cryopreservation significantly reduces the reproducibility of antigen-specific CD4+ T cell clonotypic expansions compared to CD8+ T cells. The clonality of uncultured CD4+ T cells at baseline was lower than CD8+ T cells, but this alone did not account for the observed differences post-cryopreservation. Our results suggest that freeze-thaw cycles stochastically affect CD4+ T cell clone survival and expansion capacity, leading to inconsistent repertoire changes after antigenic stimulation. These findings have important implications for experimental design and data interpretation in T cell studies, particularly those relying on cryopreserved samples for CD4+ T cell functional assays. We recommend caution when interpreting TCR repertoire data from cryopreserved CD4+ T cell data and suggest using fresh samples where possible for CD4+ T cell studies.

ORGANISM(S): Homo sapiens

PROVIDER: GSE303712 | GEO | 2025/08/31

REPOSITORIES: GEO