Abstract
Methodologies to generate single antigen-specific T cells are based on the T cell specificity, activation, or other subsequent functional measures. One of the most powerful tools to isolate human CD4+ T cell clones is utilization of MHC Class II tetramers. Flow cytometer-based tetramer technology mimics the recognition of the specific antigenic peptide in the context of HLA class II (tetramer) by the T cell receptor. MHC class II tetramers, which can be exogenously loaded to contain any peptide of interest that binds to them (T cell epitopes), provide a valuable tool for detection of T cells in the peripheral blood or the tissue that are specific for antigens from different viruses, tumors, or self-proteins (autoimmunity). Generation of T cell clones with a defined antigen specificity allows for a deeper characterization and functional assessment at single cell level. This is important for determination of the epitope specificity and functional phenotype of the disease associated T cells. Single cell cloning can be utilized in the direct sequencing of the T cell receptor alpha/beta pairs that are prevalent in the disease and therefore provides a platform for T cell receptor engineering, which has applications in the immunotherapy.
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References
Seay HR, Yusko E, Rothweiler SJ, Zhang L, Posgai AL, Campbell-Thompson M, Vignali M, Emerson RO, Kaddis JS, Ko D, Nakayama M, Smith MJ, Cambier JC, Pugliese A, Atkinson MA, Robins HS, Brusko TM (2016) Tissue distribution and clonal diversity of the T and B cell repertoire in type 1 diabetes. JCI Insight 1(20):e88242
Jacobsen LM, Posgai A, Seay HR, Haller MJ, Brusko TM (2017) T cell receptor profiling in Type 1 diabetes. Curr Diab Rep 17(11):118
Cerosaletti K, Barahmand-Pour-Whitman F, Yang J, DeBerg HA, Dufort MJ, Murray SA, Israelsson E, Speake C, Gersuk VH, Eddy JA, Reijonen H, Greenbaum CJ, Kwok WW, Wambre E, Prlic M, Gottardo R, Nepom GT, Linsley PS (2017) Single-cell RNA sequencing reveals expanded clones of islet antigen-reactive CD4+ T cells in peripheral blood of subjects with Type 1 diabetes. J Immunol 199(1):323–335
Reichstetter S, Ettinger RA, Liu AW, Gebe JA, Nepom GT, Kwok WW (2000) Distinct T cell interactions with HLA class II tetramers characterize a spectrum of TCR affinities in the human antigen-specific T cell response. J Immunol 165(12):6994–6998
Reijonen H, Novak EJ, Kochik S, Heninger AK, Liu AW, Kwok WW, Nepom GT (2002) Detection of GAD65 specific T-cells by MHC Class II tetramers in Type 1 diabetes patients and at-risk subjects. Diabetes 51:1375–1382
Nepom GT, Buckner JH, Novak EJ, Reichstetter S, Reijonen H, Gebe J, Wang R, Swanson E, Kwok WW (2002) HLA class II tetramers: tools for direct analysis of antigen-specific CD4+ T cells. Arthritis Rheum 46(1):5–12
Buckner JH, Holzer U, Novak EJ, Reijonen H, Kwok WW, Nepom GT (2002) Defining antigen-specific responses with human MHC class II tetramers. J Allergy Clin Immunol 110:199–208
Reijonen H, Kwok WW (2003) Use of HLA class II tetramers in tracking antigen-specific T-cells and mapping of T-cell epitopes. Methods 29(3):282–288
Reijonen H, Kwok WW, Nepom GT (2003) Detection of CD4+ autoreactive T-cells in T1D using HLA class II tetramers. Ann N Y Acad Sci 1005:82–87
Reijonen H, Mallone R, Heninger AK, Laughlin EM, Kochik SA, Falk BA, Kwok WW, Greenbaum C, Nepom GT (2004) GAD65 specific CD4+ T cells with high antigen avidity are prevalent in peripheral blood of type 1 diabetes patients. Diabetes 53:1987–1994
Mallone R, Kochik SA, Laughlin EM, Gersuk VH, Reijonen H, Kwok WW, Nepom GT (2004) Differential recognition and activation thresholds in human autoreactive GAD-specific T-cells. Diabetes 53(4):971–977
Mallone R, Kochik SA, Reijonen H, Carson B, Ziegler SF, Kwok W, Nepom GT (2005) Functional avidity directs T-cell fate in autoreactive CD4+ T-cells. Blood 106:2798–2805
Yang J, Danke NA, Berger D, Reichstetter S, Reijonen H, Greenbaum C, Pihoker C, James EA, Kwok WW (2006) Islet-specific glucose-6-phosphatase catalytic subunit-related protein-reactive CD4+ T cells in human subjects. J Immunol 176(5):2781–2789
Laughlin E, Burke G, Pugliese A, Falk B, Nepom G (2008) Recurrence of autoreactive antigen-specific CD4+ T cells in autoimmune diabetes after pancreas transplantation. Clin Immunol 128:23–30
Vendrame F, Pileggi A, Laughlin E, Allende G, Martin-Pagola A, Molano RD, Diamantopoulos S, Standifer N, Geubtner K, Falk BA, Ichii H, Takahashi H, Snowhite I, Chen Z, Mendez A, Chen L, Sageshima J, Ruiz P, Ciancio G, Ricordi C, Reijonen H, Nepom GT, Burke GW III, Pugliese A (2010) Recurrence of type 1 diabetes after simultaneous pancreas-kidney transplantation, despite immunosuppression, associated with autoantibodies and pathogenic autoreactive CD4 T-cells. Diabetes 59(4):947–957
Öling V, Geubtner K, Ilonen J, Reijonen H (2010) A low antigen dose selectively promotes expansion of high avidity autoreactive T cells with distinct phenotypic characteristics: a study of human autoreactive CD4+ T cells specific for GAD65. Autoimmunity 43(8):573–582
Hänninen A, Soilu-Hänninen M, Hampe CS, Deptula A, Geubtner K, Ilonen J, Knip M, Reijonen H (2010) Characterization of CD4+ T cells specific for glutamic acid decarboxylase (GAD65) and proinsulin in a patient with stiff-person syndrome but without type 1 diabetes. Diabetes Metab Res Rev 26(4):271–279
Robins H, Desmartis C, Matthis J, Livingston R, Andriesen J, Reijonen H, Carlson C, Nepom G, Yee C, Cerosaletti K (2012) Ultra-sensitive detection of rare T cell clones. J Immunol Methods 375(1–2):14–19
Yang J, Wen X, Xu H, Torres-Chinn N, Speake C, Greenbaum CJ, Nepom GT, Kwok WW (2017) Antigen-specific T cell analysis reveals that active immune responses to β cell antigens are focused on a unique set of epitopes. J Immunol 199(1):91–96
Kimura T, Kobiyama K, Winkels H, Tse K, Miller J, Vassallo M, Wolf D, Ryden C, Orecchioni M, Dileepan T, Jenkins MK, James EA, Kwok WW, Hanna DB, Kaplan RC, Strickler HD, Durkin HG, Kassaye SG, Karim R, Tien PC, Landay AL, Gange SJ, Sidney J, Sette A, Ley K (2018) Regulatory CD4+ T cells recognize MHC-II-restricted peptide epitopes of apolipoprotein B. Circulation 138(11):1130–1143
Acknowledgments
We thank Ms. Kelly Geubtner, Elsa Laughlin, Nancy Danke, and Drs. Erik Novak, Eddie James, and William Kwok for their valuable contribution to the development of MHC class II tetramer assay and T cell cloning protocols.
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Matthis, J., King, V., Reijonen, H. (2019). Production of Antigen-Specific Human CD4+ T Cell Lines and Clones. In: van Endert, P. (eds) Antigen Processing. Methods in Molecular Biology, vol 1988. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9450-2_27
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DOI: https://doi.org/10.1007/978-1-4939-9450-2_27
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