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Visualizing Antigen Specific CD4+ T Cells using MHC Class II Tetramers
15:42

Visualizing Antigen Specific CD4+ T Cells using MHC Class II Tetramers

Published on: March 6, 2009

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

Diahann T S L Jansen1, Nishta Ramnoruth1, Khai L Loh2

  • 1University of Queensland Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, Brisbane, QLD, Australia.

Frontiers in Immunology
|February 7, 2018
PubMed

Insights

Antigen-specific CD4+ T cell monitoring is crucial for new autoimmune therapies. This study presents a robust flow cytometry assay using peptide-MHC class II (pMHCII) multimers for standardized immune monitoring of CD4+ T cells ex vivo.

Area of Science:

  • Immunology
  • Cellular Immunology
  • Clinical Trials

Background:

  • Novel immunotherapies aim to induce tolerance in autoimmune diseases.
  • Monitoring antigen-specific CD4+ T cell responses is vital for assessing treatment efficacy in early trials.
  • Peptide-MHC class II (pMHCII) multimers offer a method for ex vivo analysis of CD4+ T cell populations.

Purpose of the Study:

  • To compare existing protocols and reagents for pMHCII multimer assays.
  • To identify sources of variability in tetramer binding assays.
  • To establish a standardized, robust flow cytometry assay for quantifying and phenotyping antigen-specific CD4+ T cells.

Main Methods:

  • Methodological analysis comparing published protocols for pMHCII multimer assays.
  • Testing of available reagents to identify limitations.
  • Development and description of a robust pMHCII flow cytometry-based assay.
  • Utilized frozen peripheral blood mononuclear cell (PBMC) samples for ex vivo analysis.

Main Results:

  • Identified reagent and assay limitations hindering global application of pMHCII multimer assays.
  • Characterized sources of variability in tetramer binding assays.
  • Developed a robust flow cytometry assay for ex vivo quantification and phenotyping of antigen-specific CD4+ T cells.

Conclusions:

  • A standardized pMHCII flow cytometry assay is needed for reliable immune monitoring in clinical trials.
  • The described assay provides a robust method for analyzing antigen-specific CD4+ T cells ex vivo.
  • Further cross-laboratory testing is recommended to standardize immune-monitoring results.

Keywords:
antigen-specific T cellsflow cytometryprotocol standardizationtetramerizationtetramers

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