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J Immunol Methods. 2009 Jan 1;340(1):11-24. doi: 10.1016/j.jim.2008.09.014. Epub 2008 Oct 16.

Protein kinase inhibitors substantially improve the physical detection of T-cells with peptide-MHC tetramers.

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1
Cardiff University School of Medicine, Heath Park, Cardiff, CF14 4XN, UK.

Abstract

Flow cytometry with fluorochrome-conjugated peptide-major histocompatibility complex (pMHC) tetramers has transformed the study of antigen-specific T-cells by enabling their visualization, enumeration, phenotypic characterization and isolation from ex vivo samples. Here, we demonstrate that the reversible protein kinase inhibitor (PKI) dasatinib improves the staining intensity of human (CD8+ and CD4+) and murine T-cells without concomitant increases in background staining. Dasatinib enhances the capture of cognate pMHC tetramers from solution and produces higher intensity staining at lower pMHC concentrations. Furthermore, dasatinib reduces pMHC tetramer-induced cell death and substantially lowers the T-cell receptor (TCR)/pMHC interaction affinity threshold required for cell staining. Accordingly, dasatinib permits the identification of T-cells with very low affinity TCR/pMHC interactions, such as those that typically predominate in tumour-specific responses and autoimmune conditions that are not amenable to detection by current technology.

PMID:
18929568
PMCID:
PMC3052435
DOI:
10.1016/j.jim.2008.09.014
[Indexed for MEDLINE]
Free PMC Article
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