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Cell Cycle Analysis and Relevance for Single‐Cell Gating in Mass Cytometry
Author(s) -
Rein Idun D.,
Notø Heidi Ø.,
Bostad Monica,
Huse Kanutte,
Stokke Trond
Publication year - 2020
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.23960
Subject(s) - cell cycle , flow cytometry , cytometry , single cell analysis , mass cytometry , cell , microbiology and biotechnology , interphase , mitosis , chemistry , biology , biochemistry , phenotype , gene
Cell cycle analysis by mass cytometry (MC) is hampered by the poor resolution of the Iridium‐labeled DNA intercalator compared to DNA‐specific fluorescent dyes. We report here a minimum cell cycle panel for MC consisting of Ir‐intercalator (DNA content), IdU (S phase), anti‐pS28HistoneH3 (mitosis), anti‐CDT1 (G 1 phase) and anti‐Geminin (non‐G 1 phases). Cell cycle distributions obtained by MC were not significantly different from fluorescence flow cytometry results ( r 2 = 0.98, P < 0.001). Further subdivision of the G 1 and G 2 phases could be done with anti‐pS780RB1 (late G 1 ) and anti‐PLK1 (late G 2 ), respectively. A disadvantage of MC is that aggregates of cells cannot easily be removed while retaining all single cells. We have developed an analysis pipeline including unsupervised clustering by FlowSOM and subsequent single‐cell gating. When performed on cells stained with the cell cycle panel, this analysis pipeline successfully identified debris, dead/apoptotic cells, nonsingle‐cell populations and the major cell cycle phases. The presented cell cycle panel and analysis pipeline thus achieves true single‐cell analysis at the same time as any additional channels in the panel are open for phenotyping and cell cycle‐resolved expression or modification analysis. © 2020 The Authors. Cytometry Part A published by Wiley Periodicals LLC. on behalf of International Society for Advancement of Cytometry.