Regulatory T-cells in antitumor therapy: isolation and functional testing of CD4+CD25+ regulatory T-cells.
Naturally occurring CD4+CD25+ T regulatory cells originate from the thymus and play a central role regarding the maintenance of peripheral tolerance by suppression of autoreactive T-cell populations. However, T regulatory cells can have beneficial as well as harmful effects. On the one hand, they prevent a variety of autoimmune and inflammatory diseases; but on the other hand, they concomitantly inhibit antitumor immune reactions by suppressing tumor-specific T-cell responses. Therefore, these ambivalent properties of T regulatory cells require detailed investigation especially with respect to a potential therapeutic exploitation of these cells. A prerequisite for such analyses is the isolation of pure T regulatory cells and the establishment of functional tests for the analysis of their suppressive properties, since no specific markers for CD4+CD25+ T regulatory cells are known so far. In this chapter, techniques applying immunomagnetic beads have been used to establish an efficient method for isolation of human as well as murine regulatory T-cells. A combination of positive and negative selection steps using immunomagnetic beads of different sizes yields preparations of functional, active CD4+CD25+ regulatory T-cells with high purity. Additionally, co-culture assays for functional characterization of isolated CD4+CD25+ regulatory T-cells are described that are able to detect their suppressive properties for conventional CD4+ T-cells with high sensitivity.
Jonuleit
Helmut
H
Department of Dermatology, Universitätsklinik Mainz, Mainz, Germany.
Schmitt
Edgar
E
D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology / Q000276:immunology*; D002469:Cell Separation; D006801:Humans; D007167:Immunotherapy / Q000379:methods*; D051379:Mice; D009369:Neoplasms / Q000276:immunology* / Q000628:therapy*; D010641:Phenotype; D015375:Receptors, Interleukin-2 / Q000276:immunology*
D015375:Receptors, Interleukin-2