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Isolation of CD4+ T-cells and Analysis of Circulating T-follicular Helper (cTfh) Cell Subsets from Peripheral Blood Using 6-color Flow Cytometry
Published on: January 7, 2019
Novel human CD4+ T lymphocyte subpopulations defined by CD300a/c molecule expression
Georgina J Clark1, Min Rao, Xinsheng Ju
1DC Program, Mater Medical Research Institute, Aubigny Place, Raymond Tce, South Brisbane, Queensland, Australia. gclark@mmri.mater.org.au
Journal of Leukocyte Biology
|August 19, 2007
Summary
The CD300a and CD300c molecules on T lymphocytes influence their function. The CMRF-35 antibody identifies distinct CD4+ T cell subsets with differing proliferation and cytokine production capabilities, including Th1 memory cells.
Area of Science:
- Immunology
- Molecular Biology
Background:
- CD300a and CD300c are leukocyte surface proteins involved in immune regulation, encoded on human chromosome 17.
- The CMRF-35 monoclonal antibody recognizes a common epitope on CD300a and CD300c, present on most leukocytes except B cells and some T cells.
Purpose of the Study:
- To characterize CD300a and CD300c expression on CD4+ T lymphocytes.
- To investigate the functional differences between CMRF-35 positive (CMRF-35(pos)) and negative (CMRF-35(neg)) CD4+ T cell subpopulations.
Main Methods:
- Analysis of CD300a and CD300c gene expression (mRNA) in resting and activated CD4+ T cells.
- Assessment of T cell proliferation in response to mitogens and allogeneic antigens.
- Evaluation of apoptosis rates in different T cell subpopulations.
- Measurement of IFN-gamma production following in vitro activation.
Main Results:
- Resting CD4+ T cells express CD300a mRNA and low CD300c. Activation increases both CD300a and CD300c expression.
- CMRF-35(neg) CD4+ T cells show greater proliferation than CMRF-35(pos) cells, which exhibit increased apoptosis.
- The CMRF-35(++)CD4+CD45RO+ subpopulation responds to recall antigens and produces IFN-gamma early upon activation, identifying them as Th1 memory cells.
Conclusions:
- CD300 molecule expression and CMRF-35 binding delineate distinct CD4+ T lymphocyte subsets with differential functional capacities.
- The CMRF-35(++)CD4+ T cell subset represents Th1 memory effector cells with unique activation and cytokine profiles.
