The abrogation of TCR-independent interactions with human serum ensures a selective capture of therapeutic virus-specific CD8+T-cells by Multimer Technology in Adoptive Immunotherapy

  • Beloki L
  • Ciáurriz M
  • Mansilla C
 et al. 
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Abstract

Multimers are complexes of recombinant MHC-class I molecules conjugated with antigenic immunodominant peptides and labeled with fluorescent molecules or magnetic microbeads that allow the quantification and selection of virus-specific cytotoxic T-cell subpopulations. Specific T-cell receptors recognize the immunodominant peptides and bind to the multimers. Although these complexes are only recognized by CD8+T cells with specific T-cell receptors for the particular antigen, it has been observed that multimers can also bind non-specifically to CD8- cells, such as B-cells and monocytes.Using PBMCs from CMV-seropositive healthy donors, we analyze the tendency of Pentamer and Streptamer multimers towards non-specific interactions and describe a method to avoid this unwanted event. We find that a notable proportion of multimer-positive cells are likely to represent cross-contamination by cells lacking a TCR specific for pp65. In addition, we demonstrate that this unspecific interaction can be overcome by the pre-incubation of multimer-stained PBMCs with human AB serum, without altering their capacity to bind specifically to the CD8+T cell population of interest. In conclusion, in this study we characterize a novel method to abrogate TCR-independent interactions of multimers to ensure a pure and safe therapeutic product for Adoptive Immunotherapy. © 2013 Elsevier B.V.

Author-supplied keywords

  • CMV-specific CTL
  • Fcgamma receptor I (FcγRI)
  • Pentamers
  • Streptamers

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Authors

  • Lorea Beloki

  • Miriam Ciáurriz

  • Cristina Mansilla

  • Eva Bandrés

  • Mercedes Rodríguez-Calvillo

  • Natalia Ramírez

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