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Negative and Positive Separation Techniques for the Isolation of Antigen-Specific CD8+ T Cells from Blood and Tumor Tissue

  • Margherita Gigante
  • Sharon Natasha Cox
  • Elena RanieriEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1186)

Abstract

Adoptive transfer of tumor-reactive cytotoxic T cells (CTLs) is a promising therapeutic approach for the treatment of solid tumors. To develop these strategies, it is necessary to isolate specific leukocyte subpopulations from peripheral blood or tumor tissue (referred to as tumor-infiltrating lymphocytes (TIL)) that will be reinfused into the patient after expansion in vitro. The ideal cell isolation approach should be performed rapidly, thereby maximizing the recovery and viability of the purified cells. Here, we describe the negative or the positive separation procedures to isolate CD8+ T cells from whole blood, from peripheral blood mononuclear cells (PBMCs), or from cancer tissue. Purified CD8+ cells will be used for different downstream applications such as protein and gene expression profile analysis in order to assess their intrinsic cytotoxic ability.

Key words

Tumor-infiltrating lymphocytes (TIL) CD8+ T cells Microbeads Rosettes Cancer 

Notes

Acknowledgments

This work was supported by Ministero dell’Istruzione, dell’Università e della Ricerca (MIUR) PROGRAMMA OPERATIVO NAZIONALE (PON) RICERCA E COMPETITIVITÀ 2007–2013 PONa3_00395 “BIOSCIENCES AND HEALTH” (B&H). The authors declare no conflicts of interest.

References

  1. 1.
    Kershaw MH, Trapani JA, Smyth MJ (1995) Cytotoxic lymphocytes: redirecting the cell-mediated immune response for the therapy of cancer. Ther Immunol 2:173–181PubMedGoogle Scholar
  2. 2.
    Klebanoff CA, Gattinoni L, Torabi-Parizi P et al (2005) Central memory self/tumor-reactive CD8+ T cells confer superior antitumor immunity compared with effector memory T cells. Proc Natl Acad Sci U S A 5:9571–9576CrossRefGoogle Scholar
  3. 3.
    Anikeeva N, Sykulev Y (2011) Mechanisms controlling granule-mediated cytolytic activity of cytotoxic T lymphocytes. Immunol Res. doi: 10.1007/s12026-011-8252-8 PubMedCentralPubMedGoogle Scholar
  4. 4.
    Shablak A, Hawkins RE, Rothwell DG, Elkord E (2009) T cell-based immunotherapy of metastatic renal cell carcinoma: modest success and future perspective. Clin Cancer Res. doi: 10.1158/1078-0432 PubMedGoogle Scholar
  5. 5.
    Cavalcanti E, Gigante M, Mancini V et al (2010) JAK3/STAT5/6 pathway alterations are associated with immune deviation in CD8 T cells in renal cell carcinoma patients. J Biomed Biotechnol. doi: 10.1155/2010/935764 Google Scholar
  6. 6.
    Gigante M, Mandic M, Wesa AK et al (2008) Interferon-alpha (IFN-alpha)-conditioned DC preferentially stimulate type-1 and limit Treg-type in vitro T-cell responses from RCC patients. J Immunother. doi: 10.1097/CJI.0b013e318167b023 PubMedGoogle Scholar
  7. 7.
    Groh V, Rhinehart R, Randolph-Habecker J et al (2001) Costimulation of CD8alphabeta T cells by NKG2D via engagement by MIC induced on virus-infected cells. Nat Immunol 2:255–260PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Margherita Gigante
    • 1
  • Sharon Natasha Cox
    • 1
  • Elena Ranieri
    • 2
    Email author
  1. 1.Department of Emergency and Organ TransplantationUniversity of BariBariItaly
  2. 2.Department of Surgical and Medical Sciences, School of Medicine, Ospedali RiunitiUniversity of FoggiaFoggiaItaly

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