title: In vitro expansion of antigen-specific CD8(+) T cells distorts the T-cell repertoire.
abstract: Short-term in vitro expansion of antigen-specific T cells is an appreciated assay for the analysis of small memory T-cell populations. However, how well short-term expanded T cells represent the direct ex vivo situation remains to be elucidated. In this study we compared the clonality of Epstein-Barr virus (EBV) and cytomegalovirus (CMV)-specific CD8(+) T cells directly ex vivo and after in vitro stimulation with antigen. Our data show that the antigen-specific T cell repertoire significantly alters after in vitro culture. Clear shifts in clonotype hierarchy were observed, with the most dominant ex vivo clonotype decreasing after stimulation at the expense of several previously subdominant clonotypes. Notably, these alterations were more pronounced in polyclonal T-cell populations compared to mono- or oligoclonal repertoires. Furthermore, TCR diversity significantly increased after culture with antigen. These results suggest that the T-cell repertoire is highly subjective to variation after in vitro stimulation with antigen. Hence, although short-term expansion of T cells provides a simple and efficient tool to examine antigen-specific immune responses, caution is required if T-cell populations are expanded prior to detailed, clonotypic analyses or other repertoire-based investigations.
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: Dan
authors: D
authors: Koning
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: Ana I
authors: AI
authors: Costa
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: Raiza
authors: R
authors: Hasrat
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands; Cluster of Infectious Diseases, Public Health Service, Amsterdam, The Netherlands.
authors: Bart P X
authors: BP
authors: Grady
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: Sanne
authors: S
authors: Spijkers
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands.
authors: Nening
authors: N
authors: Nanlohy
authors: Department of Theoretical Biology, Utrecht University, Utrecht, The Netherlands.
authors: Can
authors: C
authors: Keşmir
authors: Department of Immunology, University Medical Center Utrecht, Utrecht, The Netherlands; Department of Internal Medicine and Infectious Diseases, University Medical Center Utrecht, Utrecht, The Netherlands. Electronic address: debbie.van.baarle@rivm.nl.
authors: Debbie
authors: D
authors: van Baarle
mesh_terms: D000595:Amino Acid Sequence; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D049109:Cell Proliferation; D002478:Cells, Cultured; D002999:Clone Cells / Q000276:immunology / Q000378:metabolism; D003584:Cytological Techniques / Q000379:methods; D003587:Cytomegalovirus / Q000276:immunology*; D004854:Herpesvirus 4, Human / Q000276:immunology*; D006801:Humans; D016133:Polymerase Chain Reaction; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015203:Reproducibility of Results; D013997:Time Factors
chemical_list: D011948:Receptors, Antigen, T-Cell; D016693:Receptors, Antigen, T-Cell, alpha-beta
keywords: Antigen-specific T cells; CD8(+) T cells; T-cell repertoire; TCR