Progress Towards Global Standardization for Quantitative Flow Cytometry

Gonneau, Christèle; Wang, Lili; Mitra-Kaushik, Shibani; Trampont, Paul C.; Litwin, Virginia

doi:10.4155/bio-2021-0148

Cited by 6 publications

(3 citation statements)

References 11 publications

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“…From one perspective, in quantitative flow cytometry, calibration microbeads with well-defined fluorescent probes were synthesized to form calibration curves of translating preliminary fluorescence intensities into protein expressions. However, up to now, since only microbeads with surface binding sites can be accurately defined, this approach can only be used to quantify membrane proteins while it was unable to accurately estimate targeted proteins with arbitrary distribution patterns within biological cells (e.g., a specific type of protein with arbitrary expressions in a variety of cell organelles such as the membrane, cytoplasm, and nucleus). , …”

Section: Resultsmentioning

confidence: 99%

“…However, up to now, since only microbeads with surface binding sites can be accurately defined, this approach can only be used to quantify membrane proteins while it was unable to accurately estimate targeted proteins with arbitrary distribution patterns within biological cells (e.g., a specific type of protein with arbitrary expressions in a variety of cell organelles such as the membrane, cytoplasm, and nucleus). 22 , 23 …”

Section: Resultsmentioning

confidence: 99%

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Development of a Microfluidic Flow Cytometer with a Uniform Optical Field (Uni-μFCM) Enabling Quantitative Analysis of Single-Cell Proteins and Its Applications in Leukemia Gating, Tumor Classification, and Hierarchy of Cancer Stem Cells

Gao,

Zhang,

Wei

et al. 2023

ACS Sens.

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Fast and quantitative estimation of single-cell proteins with various distribution patterns remains a technical challenge. Here, a microfluidic flow cytometer with a uniform optical field (Uni-μFCM) was developed, which enabled the translation of multicolor fluorescence signals of bound antibodies into targeted protein numbers with arbitrary distributions of biological cells. As the core of Uni-μFCM, a uniform optical field for optical excitation and fluorescence detection was realized by adopting a microfabricated metal window to shape the optical beam for excitation, which was modeled and validated by both numerical simulation and experimental characterization. After the validation of Uni-μFCM in single-cell protein quantification by measuring single-cell expressions of three transcriptional factors from four cell lines of variable sizes and origins, Uni-μFCM was applied to (1) quantify membrane and cytoplasmic markers of myeloid and lymphocytic leukocytes to classify cell lines and normal and patient blood samples; (2) measure single-cell expressions of key cytokines affiliated with gene stabilities, differentiating paired oral and colon tumor cell lines with varied malignancies, and (3) quantify single-cell stemming markers of liver tumor cell lines, cell subtypes, and liver patient samples to determine a variety of lineage hierarchy. By quantitatively assessing complex cellular phenotypes, Uni-μFCM substantially expanded the phenotypic space accessible to single-cell applications in leukemia gating, tumor classification, and hierarchy determination of cancer stem cells.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Development of a Microfluidic Flow Cytometer with a Uniform Optical Field (Uni-μFCM) Enabling Quantitative Analysis of Single-Cell Proteins and Its Applications in Leukemia Gating, Tumor Classification, and Hierarchy of Cancer Stem Cells

Gao,

Zhang,

Wei

et al. 2023

ACS Sens.

View full text Add to dashboard Cite

show abstract

“…In conventional flow cytometry, results are usually described in relative terms (e.g., negative/positive, dim/intermediate/bright), or in arbitrary units of fluorescent intensities. In order to address this issue, quantitative flow cytometry has been developed where fluorescent intensities of labeled cells are translated into numbers of fluorescent probes leveraging calibration curves enabled by “calibration materials.” Since the optical intensities in the detection region of flow cytometry are not evenly distributed, the fluorescent distributions of calibration materials should be comparable with labeled biological cells [6–8].…”

Section: Introductionmentioning

confidence: 99%

Quantitative flow cytometry leveraging droplet‐based constriction microchannels with high reliability and high sensitivity

Yang

Zhang

et al. 2022

Cytometry Pt A

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This study presented a quantitative flow cytometry leveraging droplet-based constriction microchannels with high reliability and high sensitivity. Droplets encapsulating single cells and even distribution of fluorescein labeled antibodies removed from targeted cells deformed through the constriction microchannel where the excited fluorescent signals were sampled and interpreted into numbers of proteins based on volume equivalence in measurement of droplets and calibration of fluorescence. To

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Demands and technical developments of clinical flow cytometry with emphasis in quantitative, spectral, and imaging capabilities

Zhang

Gao

Chen

et al. 2022

Nanotechnology and Precision Engineering

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As the gold-standard method for single-cell analysis, flow cytometry enables high-throughput and multiple-parameter characterization of individual biological cells. This review highlights the demands for clinical flow cytometry in laboratory hematology (e.g., diagnoses of minimal residual disease and various types of leukemia), summarizes state-of-the-art clinical flow cytometers (e.g., FACSLyricTM by Becton Dickinson, DxFLEX by Beckman Coulter), then considers innovative technical improvements in flow cytometry (including quantitative, spectral, and imaging approaches) to address the limitations of clinical flow cytometry in hematology diagnosis. Finally, driven by these clinical demands, future developments in clinical flow cytometry are suggested.

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Progress Towards Global Standardization for Quantitative Flow Cytometry

Cited by 6 publications

References 11 publications

Development of a Microfluidic Flow Cytometer with a Uniform Optical Field (Uni-μFCM) Enabling Quantitative Analysis of Single-Cell Proteins and Its Applications in Leukemia Gating, Tumor Classification, and Hierarchy of Cancer Stem Cells

Development of a Microfluidic Flow Cytometer with a Uniform Optical Field (Uni-μFCM) Enabling Quantitative Analysis of Single-Cell Proteins and Its Applications in Leukemia Gating, Tumor Classification, and Hierarchy of Cancer Stem Cells

Quantitative flow cytometry leveraging droplet‐based constriction microchannels with high reliability and high sensitivity

Demands and technical developments of clinical flow cytometry with emphasis in quantitative, spectral, and imaging capabilities

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