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OMIP 073: Analysis of human thymocyte development with a 14‐color flow cytometry panel
Author(s) -
Bremer SarahJolan,
Glau Laura,
Gehbauer Christina,
Boxnick Annika,
Biermann Daniel,
Sachweh Jörg Siegmar,
Tolosa Eva,
Gieras Anna
Publication year - 2021
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.24326
Subject(s) - thymocyte , cd8 , biology , flow cytometry , cd28 , microbiology and biotechnology , cd34 , progenitor cell , phenotype , t cell , immunology , stem cell , immune system , genetics , gene
Abstract This panel was designed for the identification and detailed characterization of the different developmental steps of human thymocytes. We optimized the panel for fresh tissue in order to provide an unbiased analysis of T cell development. Accurate selection of antibodies and precise gating allow us to phenotype 14 major stages of human thymocyte development and illustrate the trajectories of T cell development from early thymic progenitors (ETP) to mature T cells that are ready to populate the periphery. The panel identifies ETPs, T‐lineage‐committed cells (TC), CD34‐positive immature single‐positive CD4 cells (ISP4 CD34+), CD34‐negative immature single‐positive CD4 cells (ISP4 CD34‐), CD45‐low early double‐positive cells (EDP CD45low), CD45‐high early double‐positive cells (EDP CD45high), late double‐positive cells (LDP), single‐positive CD4 cells (SP4), single‐positive CD8 cells (SP8), ready‐to‐egress single‐positive CD4 cells (rSP4), ready‐to‐egress single‐positive CD8 cells (rSP8), T γδ cells (Tγδ), T regulatory cells (Treg), and ready‐to‐egress T regulatory cells (rTreg). To highlight important checkpoints during T cell development, we added antibodies relevant for specific developmental steps to the panel. These include CD1a to define TCs, CD28 as a marker for ß‐selection and CD69 in combination with CD45RA to determine the maturation stage of thymocytes shortly before they become ready to egress the thymus and colonize the periphery. Moreover, Annexin V, as a marker for apoptosis, provides valuable extra information concerning the apoptotic death of thymocytes. Currently, we use this panel to identify aberrations in T cell development in health and disease.

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