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. 2021 Mar 31;12:1998. doi: 10.1038/s41467-021-22303-z

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© The Author(s) 2021

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Experimental workflow. Cell suspensions: Single-cell suspensions from PDTX samples or cell lines were derived as described in “Methods” and fixed in 4% PFA (in black) in batches of up to 20 samples; barcoding: individual samples were barcoded with the combination of the 6 palladium isotopes to enable multiplexing of up to 20 samples; cell staining and cytometry with time-of-flight (CyTOF): cell suspensions were stained with the breast cancer mass cytometry (BCMC) panel and then run through the Helios CyTOF platform. Protein markers were organised in 4 subpanels: HTC (human tumour compartment), MSC (mouse stroma compartment), OSA (oncogenic signalling activation) and CCA (cell cycle and apoptosis). b MC-based intensity distribution of HTC markers in a collection of 7 breast cancer cell lines (N_cells = 20,846). Horizontal lines (grey) represent overall median values across the samples. c tSNE plots of 7 breast cancer cell lines (as per panel b). Each spot represents a cell, coloured as per cell line of origin. All BCMC markers were included in the analysis. d tSNE plots as in c. Cells are coloured by the intensity of selected HTC markers, namely EGFR, HER2 and Keratin 8/18. All marker intensities are reported in Supplementary Fig. 1b. e tSNE plot of cells from 4 distinct samples: (i) 4T09, a mouse mammary tumour cell line, (ii) mPER, murine peritoneal tissue from NSG mice, (iii) the human breast cancer cell lines MCF7 and (iv) MDA-MB-231 (N_cells = 26,154). All BCMC markers were included in the analysis. f MC-based intensity distribution of selected HTC and MSC markers for cell lines in e. Horizontal lines (grey) represent overall median values across the samples. g Heatmap showing Earth Mover’s Distance (EMD) of OSA and CCA markers in MCF7 cells treated for 1 h with either vistusertib (1μM) or palbociclib (1μM) compared to the same cells in DMSO as a control (N_cells = 36,582). Expected changes in a subset of markers are indicated for each compound.