TY - JOUR
T1 - Protein kinase inhibitors substantially improve the physical detection of T-cells with peptide-MHC tetramers
AU - Lissina, Anna
AU - Ladell, Kristin
AU - Skowera, Ania
AU - Clement, Matthew
AU - Edwards, Emily
AU - Seggewiss, Ruth
AU - van den Berg, Hugo A.
AU - Gostick, Emma
AU - Gallagher, Kathleen
AU - Jones, Emma
AU - Melenhorst, J. Joseph
AU - Godkin, Andrew J.
AU - Peakman, Mark
AU - Price, David A.
AU - Sewell, Andrew K.
AU - Wooldridge, Linda
PY - 2009/1/1
Y1 - 2009/1/1
N2 - 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. (c) 2008 Elsevier B.V. All rights reserved.
AB - 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. (c) 2008 Elsevier B.V. All rights reserved.
U2 - 10.1016/j.jim.2008.09.014
DO - 10.1016/j.jim.2008.09.014
M3 - Article
VL - 340
SP - 11
EP - 24
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
IS - 1
ER -