Single‐Calibration Cell Size Measurement With Flow Cytometry
ABSTRACT Measuring the size of individual cells in high‐throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools for high‐throughput single‐cell biology, such as flow cytometers, only offer proxies of a cell's size, typically reported in arb...
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| Published in: | Cytometry. Part A Vol. 107; no. 4; pp. 263 - 270 |
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| Language: | English |
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Hoboken, USA
John Wiley & Sons, Inc
01.04.2025
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| Abstract | ABSTRACT
Measuring the size of individual cells in high‐throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools for high‐throughput single‐cell biology, such as flow cytometers, only offer proxies of a cell's size, typically reported in arbitrary scales and often subject to changes in the instrument's settings as selected by multiple users. In this paper, we demonstrate that it is possible to calibrate flowcytometry laser scatter signals with accurate measures of cell diameter from separate devices and that the calibration can be conserved upon changes in the laser settings. We demonstrate our approach based on flow cytometric sorting of cells of a mammalian cell line according to a selection of scatter parameters, followed by cell size determination with a Coulter counter. A straightforward procedure is presented that relates the flow cytometric scatter parameters to the absolute size measurements using linear models, along with a linear transformation that converts between different instrument settings on the flow cytometer. Our method makes it possible to record on a flow cytometer a cell's size in absolute units and correlate it with other features that are recorded in parallel in the fluorescence detection channels. |
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| AbstractList | Measuring the size of individual cells in high-throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools for high-throughput single-cell biology, such as flow cytometers, only offer proxies of a cell's size, typically reported in arbitrary scales and often subject to changes in the instrument's settings as selected by multiple users. In this paper, we demonstrate that it is possible to calibrate flowcytometry laser scatter signals with accurate measures of cell diameter from separate devices and that the calibration can be conserved upon changes in the laser settings. We demonstrate our approach based on flow cytometric sorting of cells of a mammalian cell line according to a selection of scatter parameters, followed by cell size determination with a Coulter counter. A straightforward procedure is presented that relates the flow cytometric scatter parameters to the absolute size measurements using linear models, along with a linear transformation that converts between different instrument settings on the flow cytometer. Our method makes it possible to record on a flow cytometer a cell's size in absolute units and correlate it with other features that are recorded in parallel in the fluorescence detection channels.Measuring the size of individual cells in high-throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools for high-throughput single-cell biology, such as flow cytometers, only offer proxies of a cell's size, typically reported in arbitrary scales and often subject to changes in the instrument's settings as selected by multiple users. In this paper, we demonstrate that it is possible to calibrate flowcytometry laser scatter signals with accurate measures of cell diameter from separate devices and that the calibration can be conserved upon changes in the laser settings. We demonstrate our approach based on flow cytometric sorting of cells of a mammalian cell line according to a selection of scatter parameters, followed by cell size determination with a Coulter counter. A straightforward procedure is presented that relates the flow cytometric scatter parameters to the absolute size measurements using linear models, along with a linear transformation that converts between different instrument settings on the flow cytometer. Our method makes it possible to record on a flow cytometer a cell's size in absolute units and correlate it with other features that are recorded in parallel in the fluorescence detection channels. Measuring the size of individual cells in high‐throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools for high‐throughput single‐cell biology, such as flow cytometers, only offer proxies of a cell's size, typically reported in arbitrary scales and often subject to changes in the instrument's settings as selected by multiple users. In this paper, we demonstrate that it is possible to calibrate flowcytometry laser scatter signals with accurate measures of cell diameter from separate devices and that the calibration can be conserved upon changes in the laser settings. We demonstrate our approach based on flow cytometric sorting of cells of a mammalian cell line according to a selection of scatter parameters, followed by cell size determination with a Coulter counter. A straightforward procedure is presented that relates the flow cytometric scatter parameters to the absolute size measurements using linear models, along with a linear transformation that converts between different instrument settings on the flow cytometer. Our method makes it possible to record on a flow cytometer a cell's size in absolute units and correlate it with other features that are recorded in parallel in the fluorescence detection channels. ABSTRACT Measuring the size of individual cells in high‐throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools for high‐throughput single‐cell biology, such as flow cytometers, only offer proxies of a cell's size, typically reported in arbitrary scales and often subject to changes in the instrument's settings as selected by multiple users. In this paper, we demonstrate that it is possible to calibrate flowcytometry laser scatter signals with accurate measures of cell diameter from separate devices and that the calibration can be conserved upon changes in the laser settings. We demonstrate our approach based on flow cytometric sorting of cells of a mammalian cell line according to a selection of scatter parameters, followed by cell size determination with a Coulter counter. A straightforward procedure is presented that relates the flow cytometric scatter parameters to the absolute size measurements using linear models, along with a linear transformation that converts between different instrument settings on the flow cytometer. Our method makes it possible to record on a flow cytometer a cell's size in absolute units and correlate it with other features that are recorded in parallel in the fluorescence detection channels. |
| Author | Cavallaro, Massimo Hebenstreit, Daniel Davies, Philip |
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| Cites_doi | 10.1371/journal.pbio.3002453 10.1038/msb.2011.64 10.1002/0471142956.cy0123s50 10.1007/978-3-031-10836-5 10.18637/jss.v033.i01 10.1534/genetics.119.301012 10.1128/JB.00469-20 10.1126/sciadv.abk0271 10.1039/b404251b 10.1016/j.molcel.2015.03.005 10.1371/journal.pone.0016053 10.1186/s13059-020-02227-5 10.1038/s41467-018-06912-9 10.1016/j.molcel.2022.07.017 10.1146/annurev-cellbio-120219-040142 10.52601/bpr.2022.210036 |
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| Copyright | 2025 The Author(s). published by Wiley Periodicals LLC on behalf of International Society for Advancement of Cytometry. 2025 The Author(s). Cytometry Part A published by Wiley Periodicals LLC on behalf of International Society for Advancement of Cytometry. 2025. This article is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License. |
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| Notes | Philip Davies and Massimo Cavallaro contributed equally to this work. This work was supported by EPSRC grant EP/T002794/1 and BBSRC grant BB/M017982/1. Funding ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 14 content type line 23 |
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Measuring the size of individual cells in high‐throughput experiments is often important in biomedical research and applications. Nevertheless,... Measuring the size of individual cells in high‐throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools... Measuring the size of individual cells in high-throughput experiments is often important in biomedical research and applications. Nevertheless, popular tools... |
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| SubjectTerms | Animals Calibration Cell Size computational biology Flow Flow cytometry Flow Cytometry - instrumentation Flow Cytometry - methods Flow Cytometry - standards Humans Lasers Linear Models Linear transformations Medical research Parameters Scattering Single-Cell Analysis - methods Size determination |
| Title | Single‐Calibration Cell Size Measurement With Flow Cytometry |
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