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Updated: Jun 21, 2026

Isolation of CD4+ T-cells and Analysis of Circulating T-follicular Helper (cTfh) Cell Subsets from Peripheral Blood Using 6-color Flow Cytometry
Published on: January 7, 2019
TCR repertoire and Foxp3 expression define functionally distinct subsets of CD4+ regulatory T cells
Michal Kuczma1, Iwona Pawlikowska, Magdalena Kopij
1Center for Biotechnology and Genomic Medicine, Medical College of Georgia, Augusta, GA 30912, USA.
Abstract:
Despite extensive research efforts to characterize peripheral regulatory T (T(reg)) cells expressing transcription factor Foxp3, their subset complexity, phenotypic characteristics, TCR repertoire and Ag specificities remain ambiguous. In this study, we identify and define two subsets of peripheral T(reg) cells differing in Foxp3 expression level and TCR repertoires. T(reg) cells expressing a high level of Foxp3 and TCRs not used by naive CD4(+) T cells present a stable suppressor phenotype and dominate the peripheral T(reg) population in unmanipulated mice. The second T(reg) subset, expressing a lower level of Foxp3 and using TCRs shared with naive CD4(+) T cells constitutes a small fraction of all T(reg) cells in unmanipulated mice and enriches T(reg) population with the same Ag specificities as expressed by activated/effector T cells. This T(reg) subset undergoes extensive expansion during response to Ag when it becomes a major population of Ag-specific T(reg) cells. Thus, T(reg) cells expressing TCRs shared with naive CD4(+) T cells have a flexible phenotype and may down-regulate Foxp3 expression which may restore immune balance at the conclusion of immune response or convert these cells to effector T cells producing inflammatory cytokines.
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