A platinum‐based covalent viability reagent for single‐cell mass cytometry

In fluorescence‐based flow cytometry, cellular viability is determined with membrane‐impermeable fluorescent reagents that specifically enter and label plasma membrane‐compromised nonviable cells. A recent technological advance in flow cytometry uses antibodies conjugated to elemental metal isotopes...

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Published inCytometry. Part A Vol. 81A; no. 6; pp. 467 - 475
Main Authors Fienberg, Harris G., Simonds, Erin F., Fantl, Wendy J., Nolan, Garry P., Bodenmiller, Bernd
Format Journal Article
LanguageEnglish
Published Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.06.2012
Subjects
Antibodies
Cell Adhesion
Cell Membrane Permeability
Cell Survival
cisplatin
Cisplatin - chemistry
DNA - analysis
DNA - chemistry
Female
Flow Cytometry - methods
Fluorescent Dyes
Humans
mass cytometry
Mass Spectrometry - methods
Platinum - chemistry
Single-Cell Analysis
Staining and Labeling - methods
Tumor Cells, Cultured
viability reagent
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Summary:In fluorescence‐based flow cytometry, cellular viability is determined with membrane‐impermeable fluorescent reagents that specifically enter and label plasma membrane‐compromised nonviable cells. A recent technological advance in flow cytometry uses antibodies conjugated to elemental metal isotopes, rather than to fluorophores, to allow signal detection by atomic mass spectrometry. Unhampered by the limitations of overlapping emission fluorescence, mass cytometry increases the number of parameters that can be measured in single cells. However, mass cytometry is unable to take advantage of current fluorescent viability dyes. An alternative methodology was therefore developed here in which the platinum‐containing chemotherapy drug cisplatin was used to resolve live and dead cells by mass cytometry. In a 1‐min incubation step, cisplatin preferentially labeled nonviable cells from both adherent and suspension cultures, resulting in a platinum signal quantifiable by mass cytometry. This protocol was compatible with established sample processing steps for intracellular cytometry. Furthermore, the live/dead ratios were comparable between mass‐ and fluorescence‐based cytometry. Importantly, although cisplatin is a known DNA‐damaging agent, a 1‐min “pulse” of cisplatin did not induce observable DNA damage or apoptotic responses even within 6‐h post‐exposure. Cisplatin can therefore be used as a viability reagent for a wide range of mass cytometry protocols. © 2012 International Society for Advancement of Cytometry
Bibliography:Harris Fienberg and Erin F. Simonds contributed equally to this work.
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Current address: Institute of Molecular Life Sciences, University of Zurich, Switzerland
These authors contributed equally to this work.
ISSN:1552-4922
1552-4930
1552-4930
DOI:10.1002/cyto.a.22067