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J Biol Chem. 2010 Oct 22;285(43):33144-53. doi: 10.1074/jbc.M110.141051. Epub 2010 Aug 20.

Bi-specific MHC heterodimers for characterization of cross-reactive T cells.

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Department of Pathology, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA.


T cell cross-reactivity describes the phenomenon whereby a single T cell can recognize two or more different peptide antigens presented in complex with MHC proteins. Cross-reactive T cells have previously been characterized at the population level by cytokine secretion and MHC tetramer staining assays, but single-cell analysis is difficult or impossible using these methods. In this study, we describe development of a novel peptide-MHC heterodimer specific for cross-reactive T cells. MHC-peptide monomers were independently conjugated to hydrazide or aldehyde-containing cross-linkers using thiol-maleimide coupling at cysteine residues introduced into recombinant MHC heavy chain proteins. Hydrazone formation provided bi-specific MHC heterodimers carrying two different peptides. Using this approach we prepared heterodimers of the murine class I MHC protein H-2K(b) carrying peptides from lymphocytic choriomeningitis virus and vaccinia virus, and used these to identify cross-reactive CD8+ T cells recognizing both lymphocytic choriomeningitis virus and vaccinia virus antigens. A similar strategy could be used to develop reagents to analyze cross-reactive T cell responses in humans.

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