T Cell Depletion by Cytotoxic Elimination

Karen S. Hathcock1

1 National Cancer Institute, Bethesda, Maryland
Publication Name:  Current Protocols in Immunology
Unit Number:  Unit 3.4
DOI:  10.1002/0471142735.im0304s00
Online Posting Date:  May, 2001
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This unit describes the complete removal of T cells from lymphocyte preparations based on the presence of the glycoprotein Thy‐1 on the cell surface of T lymphocytes. As in UNIT, cytotoxic elimination is employed; however, Thy‐1‐specific antibodies are used rather than MHC class II‐specific antibodies so that T cells are eliminated rather than B cells and accessory cells. Murine T cells can be further fractionated into subpopulations based on their expression of cell differentiation markers CD4 (Leu3, L3T4, T4) and CD8 (Leu2, Lyt2,3). A protocol in this unit describes removal of T cell subsets by cytotoxic elimination using CD‐specific antibodies. An alternate but technically more demanding method of enriching T cell subsets is the panning procedure described in UNIT.

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Table of Contents

  • Basic Protocol 1: Complete T Cell Removal Using Thy‐1‐Specific Monoclonal Antibodies
  • Basic Protocol 2: Removal of T Cell Subsets Using CD4/CD8‐Specific Monoclonal Antibodies
  • Commentary
  • Tables
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Literature Cited

Literature Cited
   Ceredig, R., Lowenthal, J.N., Nabholz, M., and MacDonald, H.R. 1985. Expression of interleukin‐2 receptors as a differentiation marker on intrathymic stem cells. Nature (Lond.) 314:98‐100.
   Dialynas, D.P., Wilde, D.B., Marrack, P., Pierres, A., Wall, K.A., Havran, W., Otten, G., Loken, M.R., Pierres, M., Kappler, J., and Fitch, F.W. 1983. Characterization of the murine antigenic determinant designated L3T4a, recognized by monoclonal antibody GK1.5: Expression of L3T4a by functional T cell clones appears to correlate primarily with class II MHC antigen‐reactivity. Immunol. Rev. 74:29‐58.
   Lake, P., Clark, E.A., Khorshidi, M., and Sunshine, G.H. 1979. Production and characterization of cytotoxic Thy‐1 antibody‐secreting hybrid cell lines: Detection of T cell subsets. Eur. J. Immunol. 9:875‐886.
   Ledbetter, J.A. and Herzenberg, L.A. 1979. Xenogeneic monoclonal antibodies to mouse lymphoid differentiation antigens. Immunol. Rev. 47:63‐90.
   Logberg, L., Wassmer, P., and Shevach, E.M. 1985. Role of the L3T4 antigen in T cell activation. I. Description of a monoclonal IgM antibody to a distinct epitope (L3T4b) of the L3T4 antigen and its effect on interleukin‐2 induced thymocyte proliferation. Cell. Immunol. 94:299‐311.
   Marshak‐Rothstein, A., Fink, P., Gridley, T., Raulet, D.H., Beven, M.J., and Gefter, M.L. 1979. Properties and applications of monoclonal antibodies directed against determinants of the Thy‐1 locus. J. Immunol. 122:2491‐2499.
   Sarmiento, M., Glasebrook, A.L., and Fitch, F. 1980. IgG or IgM monoclonal antibodies reactive to different determinant on the molecular complex bearing Lyt2 antigen block T cell mediated cytolysis in the absence of complement. J. Immunol. 125:2665‐2672.
   Shen, F.W. 1981. Monoclonal antibodies and T cell hybridomas. Res. Monogr. Immunol. 3:25‐37.
Key References
   Dialynas et al., 1983. See above.
  These articles describe the concepts and methods of fractionating T cell subsets on the basis of CD4 (L3T4) and CD8 (Lyt2) expression.
   Pierres, A., Naquet, P., Agthoven, A.V., Bekkahoucha, F., Denzot, D., Mishal, Z., Schmitt‐Verhulst, A.M., and Pierres, M. 1984. A rat anti‐mouse monoclonal antibody (H1 29‐19) inhibits the proliferation of Ia‐reactive T cell clones and delineates two phenotypically distinct (T4+Lyt2,3− and T4− Lyt2,3+) subsets among anti‐Ia cytolytic T cell clones. J. Immunol. 131:2775‐2783.
   Swain, S.L. 1981. Significance of Lyt phenotypes: Lyt2 antibodies block activation of T cells that recognize class I MHC antigens regardless of their function. Proc. Natl. Acad. Sci. U.S.A. 78:7101‐7105.
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