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. 2020 Sep 24;10:15686. doi: 10.1038/s41598-020-72610-6

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Cellular clustering of total CD4⁺ T cells and epitope specific CD4⁺ T cells pre and post YF-Vax vaccination. (a) UMAP was applied for visualization of the high dimensional CyTOF data set which examines the surface expression of 21 different markers in CD4⁺ T cells. Marker intensities were arcsinh transformed before further data analysis. To adjust for batch differences between samples, a z-score normalization was applied to each marker relative to the total CD4 T cell expression for that marker on the same subject and visit prior to dimensionality reduction using UMAP. Clustering was performed with a total of 82,811 CD4⁺ T cells from all 9 subjects at all time points assayed (a total of 58 samples), including 17,851 YFV-specific, 1280 EBV-specific, 2913 FLU-specific, 2767 TT-specific CD4⁺ T cells and from 58,000 non-YFV, non-EBV, non-FLU, and non-TT CD4⁺ T cells (1000 cells from each subject at each time point). PhenoGraph defined a total of 19 different clusters as indicated. (b) Heatmap of the surface marker expression levels of cells within these 19 clusters with percentage of cells that are positive for each marker. Surface markers that are used to define the dominant T cell subset for each cluster are boxed. (c) Distribution of total CD4⁺ T cells, YFV-specific, EBV- specific, FLU B-specific and TT-specific T cells in UMAP.