Flow Cytometric Clinical Immunomonitoring Using Peptide-MHC Class II Tetramers: Optimization of Methods and Protocol Development.

Autor:	Jansen DTSL; University of Queensland Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, Brisbane, QLD, Australia., Ramnoruth N; University of Queensland Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, Brisbane, QLD, Australia., Loh KL; Infection and Immunity Program and The Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia., Rossjohn J; Infection and Immunity Program and The Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia.; ARC Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, VIC, Australia.; School of Medicine, Institute of Infection and Immunity, Cardiff University, Cardiff, United Kingdom., Reid HH; Infection and Immunity Program and The Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia.; ARC Centre of Excellence in Advanced Molecular Imaging, Monash University, Clayton, VIC, Australia., Nel HJ; University of Queensland Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, Brisbane, QLD, Australia., Thomas R; University of Queensland Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, Brisbane, QLD, Australia.
Jazyk:	angličtina
Zdroj:	Frontiers in immunology [Front Immunol] 2018 Jan 22; Vol. 9, pp. 8. Date of Electronic Publication: 2018 Jan 22 (Print Publication: 2018).
DOI:	10.3389/fimmu.2018.00008
Abstrakt:	With the advent of novel strategies to induce tolerance in autoimmune and autoimmune-like conditions, clinical trials of antigen-specific tolerizing immunotherapy have become a reality. Besides safety, it will be essential to gather mechanistic data on responding CD4 + T cells to assess the effects of various immunomodulatory approaches in early-phase trials. Peptide-MHC class II (pMHCII) multimers are an ideal tool for monitoring antigen-specific CD4 + T cell responses in unmanipulated cells directly ex vivo . Various protocols have been published but there are reagent and assay limitations across laboratories that could hinder their global application to immune monitoring. In this methodological analysis, we compare protocols and test available reagents to identify sources of variability and to determine the limitations of the tetramer binding assay. We describe a robust pMHCII flow cytometry-based assay to quantify and phenotype antigen-specific CD4 + T cells directly ex vivo from frozen peripheral blood mononuclear cell samples, which we suggest should be tested across various laboratories to standardize immune-monitoring results.
Databáze:	MEDLINE
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