Multiplexed combinatorial tetramer staining in a mouse model of virus infection

Tania Cukalac, Sophie Alessandra Valkenburg, Nicole L. La Gruta, Stephen J. Turner, Peter C Doherty, Katherine Kedzierska

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8 Citations (Scopus)


Use of fluorescently labelled multimers, particularly tetramers of peptide and MHC class I glycoprotein (pMHC-I) complexes, is essential for the analysis of CD8+ T cell immunity in basic research and clinical settings. A recently described combinatorial approach using pMHC-I multimers coupled to a unique combination of distinct fluorochromes has facilitated the simultaneous screening of multiple T cell specificities within a single human blood sample. The present analysis establishes that this multiplexed tetramer staining protocol can also be applied in mouse models of a disease to detect multiple subdominant CD8+ T cell specificities in the presence of prominent immunodominant T cell sets at different stages of infection. We have established a modified protocol that concurrently identified influenza-specific CD8+ T cells at the acute and long-term memory phases of influenza virus infection in B6 mice. Highly dominant (DbNP366 +CD8+ and DbPA224 +CD8+) and subdominant (KbPB1703 +CD8+, DbPB1-F262 +CD8+ and KbNS2114 +CD8+) T cell responses can be detected simultaneously at levels comparable to the conventional tetramer staining with this combinatorial approach. The technique proved particularly useful with aged mice, where we used 5-fold fewer animals, making the detection of multiple T cell specificities more cost-effective and less time-consuming. Overall, our study establishes that this comprehensive concurrent analysis of multiple T cell specificities is of value for analysing mouse models of disease, especially in situations where sample size and/or response magnitude is limiting.

Original languageEnglish
Pages (from-to)157-161
Number of pages5
JournalJournal of Immunological Methods
Issue number1-2
Publication statusPublished - Aug 2010
Externally publishedYes


  • Cytotoxic T cells
  • Mouse B6 model
  • PMHC tetramers
  • T cells
  • Viral infection

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