Comparison of peptide-major histocompatibility complex tetramers and dextramers for the identification of antigen-specific T cells

Clin Exp Immunol. 2014 Jul;177(1):47-63. doi: 10.1111/cei.12339.


Fluorochrome-conjugated peptide-major histocompatibility complex (pMHC) multimers are widely used for flow cytometric visualization of antigen-specific T cells. The most common multimers, streptavidin-biotin-based 'tetramers', can be manufactured readily in the laboratory. Unfortunately, there are large differences between the threshold of T cell receptor (TCR) affinity required to capture pMHC tetramers from solution and that which is required for T cell activation. This disparity means that tetramers sometimes fail to stain antigen-specific T cells within a sample, an issue that is particularly problematic when staining tumour-specific, autoimmune or MHC class II-restricted T cells, which often display TCRs of low affinity for pMHC. Here, we compared optimized staining with tetramers and dextramers (dextran-based multimers), with the latter carrying greater numbers of both pMHC and fluorochrome per molecule. Most notably, we find that: (i) dextramers stain more brightly than tetramers; (ii) dextramers outperform tetramers when TCR-pMHC affinity is low; (iii) dextramers outperform tetramers with pMHC class II reagents where there is an absence of co-receptor stabilization; and (iv) dextramer sensitivity is enhanced further by specific protein kinase inhibition. Dextramers are compatible with current state-of-the-art flow cytometry platforms and will probably find particular utility in the fields of autoimmunity and cancer immunology.

Keywords: T cell receptors; T cells; autoimmunity; diabetes; tumour immunology.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biotin / chemistry
  • CD4-Positive T-Lymphocytes / cytology
  • CD4-Positive T-Lymphocytes / immunology*
  • CD8-Positive T-Lymphocytes / cytology
  • CD8-Positive T-Lymphocytes / immunology*
  • Cell Line
  • Cell Separation / methods*
  • Clone Cells
  • Dextrans / chemistry
  • Flow Cytometry
  • Fluorescent Dyes / chemistry
  • HLA-A2 Antigen / chemistry
  • HLA-DR1 Antigen / chemistry
  • HLA-DR1 Antigen / metabolism
  • Hemagglutinins, Viral / metabolism
  • Humans
  • Insulin / metabolism
  • Major Histocompatibility Complex / immunology*
  • Peptide Fragments / metabolism
  • Protein Binding
  • Protein Precursors / metabolism
  • Streptavidin / chemistry
  • T-Cell Antigen Receptor Specificity / immunology
  • Telomerase / metabolism


  • Dextrans
  • Fluorescent Dyes
  • HLA-A2 Antigen
  • HLA-DR1 Antigen
  • Hemagglutinins, Viral
  • Insulin
  • Peptide Fragments
  • Protein Precursors
  • preproinsulin
  • Biotin
  • Streptavidin
  • telomerase reverse transcriptase (540-548)
  • Telomerase