Cell size assays for mass cytometry

Stern, Ari; Rahman, Adeeb; Birtwistle, Marc R.

doi:10.1002/cyto.a.23000

Cited by 55 publications

(47 citation statements)

References 36 publications

(45 reference statements)

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“…See protocol [46] for conjugating antibodies with these heavy-metals. Beyond this, users can employ quantum dot labeled antibodies to gain additional channels [47], iodine for cell cycle tracking [124], RNA transcripts [131][132][133], platinum drug uptake [128,129], cell cycle status and DNA synthesis [130], cell size [125], apoptosis [48], and viability [27] Proteins, phospho-proteins, chromatin modifications [145], RNA transcripts [146], fluorescent drug uptake [147], metabolism and redox state [148,149], cell cycle status and DNA synthesis [150], cell size and granularity, apoptosis [151] and viability [152] Poly-adenylated RNA transcripts, CITE-seq probes [144] Minimum cells per sample needed at start of protocol Max is an example maximum range of the instrument scale; error is the typical error for that scale. c Includes capabilities of novel instruments such as Cytek TM Aurora (Cytek Biosciences), ZE5 TM (Bio-Rad), and FACSymphony TM (BD).…”

Section: Multiplexing Capabilitymentioning

confidence: 99%

Beyond the message: advantages of snapshot proteomics with single‐cell mass cytometry in solid tumors

et al. 2019

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Single‐cell technologies that can quantify features of individual cells within a tumor are critical for treatment strategies aiming to target cancer cells while sparing or activating beneficial cells. Given that key players in protein networks are often the primary targets of precision oncology strategies, it is imperative to transcend the nucleic acid message and read cellular actions in human solid tumors. Here, we review the advantages of multiplex, single‐cell mass cytometry in tissue and solid tumor investigations. Mass cytometry can quantitatively probe nearly any cellular feature or target. In discussing the ability of mass cytometry to reveal and characterize a broad spectrum of cell types, identify rare cells, and study functional behavior through protein signaling networks in millions of individual cells from a tumor, this review surveys publications of scientific advances in solid tumor biology made with the aid of mass cytometry. Advances discussed include functional identification of rare tumor and tumor‐infiltrating immune cells and dissection of cellular mechanisms of immunotherapy in solid tumors and the periphery. The review concludes by highlighting ways to incorporate single‐cell mass cytometry in solid tumor precision oncology efforts and rapidly developing cytometry techniques for quantifying cell location and sequenced nucleic acids.

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Section: Multiplexing Capabilitymentioning

confidence: 99%

Beyond the message: advantages of snapshot proteomics with single‐cell mass cytometry in solid tumors

et al. 2019

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“…In a recent publication, Stern et al showed that the lipid stain osmium tetroxide (which is detectable on a mass cytometer) correlates with forward light scatter and can be used as a proxy for cell size. There is of course no guarantee that the amount of lipids would always be proportionate to cell volume and, indeed, eosinophils exhibited disproportionately strong staining compared to other blood cell types.…”

Section: Indirect Methods: Light Scattering and Cell Stainingmentioning

confidence: 99%

Methods for cell volume measurement

Model

2017

Cytometry Pt A

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Volume is an essential characteristic of a cell, and this review describes the main methods of its measurement that have been used in the past several decades. The discussed methods include various implementations of light scattering, estimates based on one or two cell dimensions, surface scanning, fluorescence confocal and transmission slice-by-slice imaging, intracellular volume markers, displacement of extracellular solution, quantitative phase imaging, radioactive methods, and some others. Suitability of these methods to some typical samples and applications is discussed. © 2017 International Society for Advancement of Cytometry.

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“…Accurately measuring single-cell size remains a challenge for mammalian cells due to their irregular shape. Existing techniques require specialized hardware, fluorescent labeling 10,11 and/or cell suspension [12][13][14][15][16] . Fluorescent labeling or over-expression of a target marker can alter cell function.…”

mentioning

confidence: 99%

CTRL: a label-free method for dynamic measurement of single-cell volume

Yao

Rochman

Sun

2019

Preprint

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Measuring the physical size of the cell is valuable in understanding cell growth control. Current singlecell volume measurement methods for mammalian cells are labor-intensive, inflexible, and can cause cell damage. We introduce CTRL: Cell Topography Reconstruction Learner, a label-free technique incorporating Deep Learning and Fluorescence Exclusion for reconstructing cell topography and estimating mammalian cell volume from DIC microscopy images alone. The method achieves quantitative accuracy, requires minimal sample preparation, and applies to extensive biological and experimental conditions. Using this method, we observe a noticeable reduction in cell size fluctuations during cell cycle, which is consistent with the presence of a cell size checkpoint.(https://GitHub.com/sxslabjhu/CTRL) Main textCell size plays a critical role during cell growth, division, and proliferation 1-5 . Abnormalities in cell size regulation and growth control are thought to promote disease development 2,5-9 . Accurately measuring single-cell size remains a challenge for mammalian cells due to their irregular shape. Existing techniques require specialized hardware, fluorescent labeling 10,11 and/or cell suspension [12][13][14][15][16] . Fluorescent labeling or over-expression of a target marker can alter cell function. Cell suspension alters the cell shape and biochemical signaling from the extracellular matrix, and also potentially affecting cell size. None of these methods has been successfully applied to measure mammalian cell growth at the single-cell level.While sensitive and accurate methods have been developed to measure single-cell mass over time 17 , the relationship between cell size and mass is not always clear.An accurate and high throughput method of cell volume quantification is the Fluorescence Exclusion method (FXm), first proposed in 1983 14 and subsequently developed and refined by several groups [18][19][20] . Cells are seeded in a micro-fabricated chamber and a membrane-impermeable high molecular weight fluorescent dye (e.g. FITC-dextran) is injected into the microchamber (Fig. 1a). The cell excludes its volume in the microchamber, therefore the total fluorescence loss is proportional to the cell volume.The FXm method obtains the cell volume from a single epi-fluorescence image, and therefore is high throughput 3,18-20 . However, due to endocytosis 3,20 that is common in many cell types, the dye eventually enters the cytoplasm, and therefore FXm generally cannot accurately report cell volume in time-lapse without careful controls. Fluorescent imaging also introduces photobleaching, which alters the signal during time-lapse measurements. Moreover, microfluidic fabrication is needed to perform the experiment and the confinement of the microchamber may alter cell physiological processes over long periods. These drawbacks limit the use and applicability of FXm for studying cell growth.Convolutional Neural Networks (CNN) have been applied to microscopy images for both phenotype classification 21,22 and image segmentati...

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Cell size assays for mass cytometry

Cited by 55 publications

References 36 publications

Beyond the message: advantages of snapshot proteomics with single‐cell mass cytometry in solid tumors

Beyond the message: advantages of snapshot proteomics with single‐cell mass cytometry in solid tumors

Methods for cell volume measurement

CTRL: a label-free method for dynamic measurement of single-cell volume

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