The Application of Flow Cytometry for Simultaneous and Multi-parametric Analysis of Heterogenous Cell Populations in Basic and Clinical Research

Schmit, Taylor; Klomp, Mitchell; Khan, M. Nadeem

doi:10.1007/978-1-0716-1001-5_14

Cited by 5 publications

(5 citation statements)

References 7 publications

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“…For such high dimensional data samples, extracting relevant biological information becomes challenging (Newell and Cheng, 2016). Moreover, in large clinical studies, possibly with some longitudinal and multicentric components, this difficult task is itself multiplied by the high number of samples (Hartmann et al, 2019, Schmit, Klomp, and Khan, 2021). In the last decade, this led to the development of (semi) automated data analysis pipelines (Saeys, Van Gassen, and Lambrecht, 2016), to complement the traditional manual gating approaches (Staats et al, 2019).…”

Section: Introductionmentioning

confidence: 99%

Visual quality control withCytoMDS, a Bioconductor package for low dimensional representation of cytometry sample distances

Hauchamps,

Delandre,

Temmerman

et al. 2024

Preprint

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Quality Control (QC) of samples is an essential preliminary step in cytometry data analysis. Notably, identification of potential batch effects and outlying samples is paramount, to avoid mistaking these effects for true biological signal in downstream analyses. However, this task can prove to be delicate and tedious, especially for datasets with many samples.Here, we presentCytoMDS, a Bioconductor package implementing a dedicated method for low dimensional representation of cytometry samples composed of marker expressions for up to millions of single cells. This method allows a global representation of all samples of a study, with one single point per sample, in such a way that projected distances can be visually interpreted. It usesEarth Mover’s Distancefor assessing dissimilarities between multi-dimensional distributions of marker expression, andMulti Dimensional Scalingfor low dimensional projection of distances. Some additional visualization tools, both for projection quality diagnosis and for user interpretation of the projection coordinates, are also provided in the package.We demonstrate the strengths and advantages ofCytoMDSfor QC of cytometry data on three real biological datasets, revealing the presence of low quality samples, batch effects and biological signal between sample groups.

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Section: Introductionmentioning

confidence: 99%

Visual quality control withCytoMDS, a Bioconductor package for low dimensional representation of cytometry sample distances

Hauchamps,

Delandre,

Temmerman

et al. 2024

Preprint

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“…Flow cytometry is a technology that provides rapid multi-parametric analysis of suspended single cells in solution . Over the last 30 years, flow cytometry has offered several unique advantages as it realized multi-parametric , quantitative analysis of cell population , at the single-cell level with high processing speed . As a result, flow cytometry has become a powerful and widely accepted measurement method in biology, life science, and, especially, clinical diagnostics .…”

Section: Introductionmentioning

confidence: 99%

“…As a result, flow cytometry has become a powerful and widely accepted measurement method in biology, life science, and, especially, clinical diagnostics . For instance, detailed characteristics of cell populations obtained by flow cytometry are crucially important for the diagnosis of blood diseases and infectious diseases. , Working on the principle of light scattering and fluorescence emission by the fluorescently labeled cells as they pass through a laser beam, the existing commercial flow cytometers (FCMs) often consist of sophisticated optical components and flow control devices, resulting in a relatively huge volume . Typical FCMs are at the meter scale, with volumes between 240 and 860 L .…”

Section: Introductionmentioning

confidence: 99%

“…8 For instance, detailed characteristics of cell populations obtained by flow cytometry are crucially important for the diagnosis of blood diseases 9 and infectious diseases. 3,10 Working on the principle of light scattering and fluorescence emission by the fluorescently labeled cells as they pass through a laser beam, 11 the existing commercial flow cytometers (FCMs) often consist of sophisticated optical components and flow control devices, resulting in a relatively huge volume. 8 Typical FCMs are at the meter scale, with volumes between 240 and 860 L. 12 Such a large dimension limits the application of commercial FCMs on point-of-care testing diagnostics, especially in resource-limited or requiring on-site analysis occasions.…”

Section: Introductionmentioning

confidence: 99%

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Ultraportable Flow Cytometer Based on an All-Glass Microfluidic Chip

Cui

Xie

et al. 2023

Anal. Chem.

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The flow cytometer has become a powerful and widely accepted measurement device in both biological studies and clinical diagnostics. The application of the flow cytometer in emerging point-of-care scenarios, such as instant detection in remote areas and emergency diagnosis, requires a significant reduction in physical dimension, cost, and power consumption. This requirement promotes studies to develop portable flow cytometers, mostly based on the utilization of polymer microfluidic chips. However, due to the relatively poor optical performance of polymer materials, existing microfluidic flow cytometers are incapable of accurate blood analysis, such as the four-part leukocyte differential count, which is necessary to monitor the immune system and to assess the risk of allergic inflammation or viral infection. To address this issue, an ultraportable flow cytometer based on an all-glass microfluidic chip (AG-UFCM) has been developed in this study. Compared with that of a typical commercial flow cytometer (BD FACSAria III), the volume of the AG-UFCM was reduced by 90 times (from 720 to 8 L). A two-step laser processing was employed to fabricate an all-glass microfluidic chip with a surface roughness of less than 1 nm, significantly improving the optical performance of on-chip micro-lens. The signal-tonoise ratio was enhanced by 3 dB, compared with that of polymer materials. For the first time, a four-part leukocyte differential count based on single fluorescence staining was realized using a miniaturized flow cytometer, laying a foundation for the point-of-care testing of miniaturized flow cytometers.

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“…As the golden approach of quantitative single-cell protein analysis, fluorescence flow cytometry measures fluorescence intensities of single cells stained with antibodies tagged with fluorescence. , By combining with other technologies such as spectrometers , or cameras, , state-of-the-art fluorescence flow cytometry can provide quantitative insights in complex single-cell protein analysis associated with varied types of signals with diverse localizations. However, without a standardized calibration of fluorescence intensities, results of fluorescence flow cytometry can only be described in relative terms (e.g., negative/positive or dim/intermediate/bright) rather than protein numbers, severely compromising the developments of quantitative analysis of single-cell proteins. , …”

Section: Introductionmentioning

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.

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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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The Application of Flow Cytometry for Simultaneous and Multi-parametric Analysis of Heterogenous Cell Populations in Basic and Clinical Research

Cited by 5 publications

References 7 publications

Visual quality control withCytoMDS, a Bioconductor package for low dimensional representation of cytometry sample distances

Visual quality control withCytoMDS, a Bioconductor package for low dimensional representation of cytometry sample distances

Ultraportable Flow Cytometer Based on an All-Glass Microfluidic Chip

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

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