Analysis of gag-specific cytotoxic T lymphocytes in simian immunodeficiency virus-infected rhesus monkeys by cell staining with a tetrameric major histocompatibility complex class I-peptide complex

Marcelo J. Kuroda, Jörn E. Schmitz, Dan H. Barouch, Abie Craiu, Todd M. Allen, Alessandro Sette, David Watkins, Meryl A. Forman, Norman L. Letvin

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Abstract

A tetrameric recombinant major histocompatibility complex (MHC) class I- peptide complex was used as a staining reagent in flow cytometric analyses to quantirate and define the phenotype of Gag-specific cytotoxic T lymphocytes (CTLs) in the peripheral blood of simian immunodeficiency virus macaque (SIVmac)-infected rhesus monkeys. The heavy chain of the rhesus monkey MHC class I molecule Manre-A*01 and β2-microglobulin were refolded in the presence of an SIVmac Gag synthetic peptide (p11C, C-M) representing the optimal nine-amino acid peptide of Mamu-A*01-restricted predominant CTL epitope to create a tetrameric Mamu-A*01/p11C, C-M complex. Tetrameric Mamu- A*01/p11C, C-M complex bound to T cells of SIVmac-infected, Mamu-A*01+, but not uninfected, Mamu-A*01+, or infected, Mamu-A*01- rhesus monkeys. Specific staining of peripheral blood mononuclear cells (PBMC) from SIVmac- infected, Mamu-A*01+ rhesus monkeys was only found in the cluster of differentiation (CD)8α/β+ T lymphocyte subset and the percentage of CD8α/β+ T cells in the peripheral blood of four SIVmac-infected, Mamu- A*01+ rhesus monkeys staining with this complex ranged from 0.7 to 10.3%. Importantly, functional SIVmac Gag p11C-specific CTL activity was seen in sorted and expanded tetrameric Mamu-A*01/p11 C, C-M complex-binding, but not nonbinding, CD8α/β+ T cells. Furthermore, the percentage of CD8α/β+ T cells binding this tetrameric Mamu-A*01/p11C, C-M complex correlated well with p11C-specific cytotoxic activity as measured in both bulk and limiting dilution effector frequency assays. Finally, phenotypic characterization of the cells binding this tetrameric complex indicated that this lymphocyte population is heterogeneous. These studies indicate the power of this approach for examining virus-specific CTLs in in vivo settings.

Original languageEnglish
Pages (from-to)1373-1381
Number of pages9
JournalJournal of Experimental Medicine
Volume187
Issue number9
DOIs
StatePublished - May 4 1998
Externally publishedYes

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Simian Immunodeficiency Virus
Macaca
Cytotoxic T-Lymphocytes
Major Histocompatibility Complex
Macaca mulatta
Staining and Labeling
Peptides
T-Lymphocytes
T-Lymphocyte Epitopes
T-Lymphocyte Subsets
Blood Cells
Lymphocytes
Viruses
Phenotype
Amino Acids
Population

ASJC Scopus subject areas

  • Immunology

Cite this

Analysis of gag-specific cytotoxic T lymphocytes in simian immunodeficiency virus-infected rhesus monkeys by cell staining with a tetrameric major histocompatibility complex class I-peptide complex. / Kuroda, Marcelo J.; Schmitz, Jörn E.; Barouch, Dan H.; Craiu, Abie; Allen, Todd M.; Sette, Alessandro; Watkins, David; Forman, Meryl A.; Letvin, Norman L.

In: Journal of Experimental Medicine, Vol. 187, No. 9, 04.05.1998, p. 1373-1381.

Research output: Contribution to journalArticle

Kuroda, Marcelo J. ; Schmitz, Jörn E. ; Barouch, Dan H. ; Craiu, Abie ; Allen, Todd M. ; Sette, Alessandro ; Watkins, David ; Forman, Meryl A. ; Letvin, Norman L. / Analysis of gag-specific cytotoxic T lymphocytes in simian immunodeficiency virus-infected rhesus monkeys by cell staining with a tetrameric major histocompatibility complex class I-peptide complex. In: Journal of Experimental Medicine. 1998 ; Vol. 187, No. 9. pp. 1373-1381.
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abstract = "A tetrameric recombinant major histocompatibility complex (MHC) class I- peptide complex was used as a staining reagent in flow cytometric analyses to quantirate and define the phenotype of Gag-specific cytotoxic T lymphocytes (CTLs) in the peripheral blood of simian immunodeficiency virus macaque (SIVmac)-infected rhesus monkeys. The heavy chain of the rhesus monkey MHC class I molecule Manre-A*01 and β2-microglobulin were refolded in the presence of an SIVmac Gag synthetic peptide (p11C, C-M) representing the optimal nine-amino acid peptide of Mamu-A*01-restricted predominant CTL epitope to create a tetrameric Mamu-A*01/p11C, C-M complex. Tetrameric Mamu- A*01/p11C, C-M complex bound to T cells of SIVmac-infected, Mamu-A*01+, but not uninfected, Mamu-A*01+, or infected, Mamu-A*01- rhesus monkeys. Specific staining of peripheral blood mononuclear cells (PBMC) from SIVmac- infected, Mamu-A*01+ rhesus monkeys was only found in the cluster of differentiation (CD)8α/β+ T lymphocyte subset and the percentage of CD8α/β+ T cells in the peripheral blood of four SIVmac-infected, Mamu- A*01+ rhesus monkeys staining with this complex ranged from 0.7 to 10.3{\%}. Importantly, functional SIVmac Gag p11C-specific CTL activity was seen in sorted and expanded tetrameric Mamu-A*01/p11 C, C-M complex-binding, but not nonbinding, CD8α/β+ T cells. Furthermore, the percentage of CD8α/β+ T cells binding this tetrameric Mamu-A*01/p11C, C-M complex correlated well with p11C-specific cytotoxic activity as measured in both bulk and limiting dilution effector frequency assays. Finally, phenotypic characterization of the cells binding this tetrameric complex indicated that this lymphocyte population is heterogeneous. These studies indicate the power of this approach for examining virus-specific CTLs in in vivo settings.",
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AU - Barouch, Dan H.

AU - Craiu, Abie

AU - Allen, Todd M.

AU - Sette, Alessandro

AU - Watkins, David

AU - Forman, Meryl A.

AU - Letvin, Norman L.

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