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Analyzing the phenotypic and functional complexity of lymphocytes using CyTOF (cytometry by time-of-flight)

Cellular & Molecular Immunology volume 9, pages 322–323 (2012)Cite this article

The human immune system is equipped with vast numbers of lymphocytes possessing distinct antigen specificities. The enormous repertoire and heterogeneity of lymphocytes ensures their ability to effectively fight against external microbial invaders, as well as internal aberrant cells. However, the understanding of the specific phenotypes and functions of lymphocytes is hindered by the lack of tools that can be used to visualize this complexity. Fluorescent dye-based flow cytometry, which can simultaneously analyze 8–10 parameters of a single cell, is commonly used in immunological research, but further expansion of the number of parameters (up to 10–18) is limited by the overlapping excitation and emission spectra of different fluorescent dyes. Recently, Bendall et al.1 used transition element isotopes as chelated antibody tags for the atomic mass spectrometric analysis of single cells, called cytometry by time-of-flight (CyTOF), which expanded the analysis to 34 or more parameters simultaneously. In addition to greatly increasing the number of the parameters that can be evaluated, these isotopes have very little crosstalk and no need for background adjustment, thus eliminating two other common problems associated with fluorescent dyes.

Using this newly developed CyTOF technology, Newell et al.2 analyzed the phenotypic and functional characteristics of human CD8+ T cells by measuring 17 cell surface markers, six cytokines, two cytotoxic granule components and three viral antigen peptide-MHC tetramers on human CD8+ T cells. These parameters covered nine functional attributes and distinguished all reported CD8+ T-cell subsets, such as naive, central memory, effector memory, terminal effector, long-lived memory precursor effector cells and short-lived effector cells. The authors used principal component analysis (PCA) to analyze the large CyTOF datasets and presented their results either in two-dimensional PCA or in combination with a protein structure program (PyMOL) as three-dimensional PCA.

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References

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  1. Laboratory of Molecular Biology and Immunology, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA

    Guobing Chen & Nan-ping Weng

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  1. Guobing Chen
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  2. Nan-ping Weng
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Correspondence to Nan-ping Weng.

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Chen, G., Weng, Np. Analyzing the phenotypic and functional complexity of lymphocytes using CyTOF (cytometry by time-of-flight). Cell Mol Immunol 9, 322–323 (2012). https://doi.org/10.1038/cmi.2012.16

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