Time-resolved assessment of single-cell protein secretion by sequencing

Wu, Tongjin; Womersley, Howard John; Wang, Jiehao Ray; Scolnick, Jonathan; Cheow, Lih Feng

doi:10.1038/s41592-023-01841-y

Cited by 13 publications

(2 citation statements)

References 72 publications

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“…28 Recently, droplet-based phenotyping sequencing technology was developed to capture secreted proteins on cell surfaces, which were then labeled with gene barcodes for extensive analysis of single-cell secretion over time. 29 However, massive single-cell sequencing and de-barcoding were time-consuming and costly for diagnosis. A summary of different main singlecell secretion assays is included for comparison (Supporting information 1).…”

Section: ■ Introductionmentioning

confidence: 99%

“…For example, immune probes with dual antibodies were anchored to cell-surface proteins, such as CD45, to capture secretions for high-throughput single-cell secretion screening through flow cytometry, − while dual antibodies proved difficult to extend to other secreted cells, such as plasma B cells, that do not highly express CD45 . Recently, droplet-based phenotyping sequencing technology was developed to capture secreted proteins on cell surfaces, which were then labeled with gene barcodes for extensive analysis of single-cell secretion over time . However, massive single-cell sequencing and de-barcoding were time-consuming and costly for diagnosis.…”

Section: Introductionmentioning

confidence: 99%

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Single-Cell Secretion Analysis via Microfluidic Cell Membrane Immunosorbent Assay for Immune Profiling

Xu,

Wu,

Jiang

et al. 2023

Anal. Chem.

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Single-cell multiplexed phenotypic analysis expands the biomarkers for diagnosis, heralding a new era of precision medicine. Cell secretions are the primary measures of immune function, but single-cell screening remains challenging. Here, a novel cell membrane-based assay was developed using cholesterollinked antibodies (CLAbs), integrating immunosorbent assays and droplet microfluidics to develop a flexible high-throughput singlecell secretion assay for multiplexed phenotyping. CLAb-grafted single cells were encapsulated in water-in-oil droplets to capture their own secretions. Subsequently, the cells were extracted from droplets for fluorescence labeling and screening. Multiple secretions and surface proteins were simultaneously measured from single cells by flow cytometry. To validate the approach, THP-1 cells, THP-1-derived M1 macrophages, and dendritic cells were assayed, indicating the differentiation efficiency of THP-1 cells under different chemical stimulations. Moreover, peripheral blood mononuclear cells from healthy donors under various stimuli showed varied active immune cell populations (6.62−47.14%). The peripheral blood mononuclear cells (PBMCs) of nasopharyngeal carcinoma patients were analyzed to identify a higher percentage of actively cytokine-secreted single cells in the basal state (2.82 ± 1.48%), compared with that in the health donors (0.70 ± 0.29%).

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Section: ■ Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Single-Cell Secretion Analysis via Microfluidic Cell Membrane Immunosorbent Assay for Immune Profiling

Xu,

Wu,

Jiang

et al. 2023

Anal. Chem.

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A Microfluidic 3D‐Tumor‐Spheroid Model for the Evaluation of Targeted Therapies from Angiogenesis‐Related Cytokines at the Single Spheroid Level

Liu,

Wang,

Wang

et al. 2024

Adv Healthcare Materials

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Angiogenesis is a key player in drug resistance to targeted therapies for breast cancer. The average expression of angiogenesis‐related cytokines is widely associated with the treatments of target therapies for a population of cells or spheroids, overlooking the distinct responses for individuals. In this work, a highly integrated microfluidic platform is developed for the generation of monodisperse multicellular tumor spheroids (MTSs), drug treatments, and the measurement of cytokines for individual MTSs in a single chip. The platform allows the correlation evaluation between cytokine secretion and drug treatment at the level of individual spheroids. For validation, quantities of six representative proangiogenic cytokines are tested against treatments with four model drugs at varying times and concentrations. By applying a linear regression model, significant correlations are established between cytokine secretion and the treated drug concentration for individual spheroids. The proposed platform provides a high‐throughput method for the investigation of the molecular mechanism of the cytokine response to targeted therapies and paves the way for future drug screening using predictive regression models at the single‐spheroid level.

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Integration of single‐cell transcriptome and proteome technologies: Toward spatial resolution levels

Wang

Dang

et al. 2023

VIEW

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Cells are basic building blocks of life with vast heterogeneity. Nowadays, the rapid development of single‐cell multiomics (scMulti‐Omics) has facilitated comprehensive understanding of gene regulatory networks, cellular characteristics, and temporal dynamics. However, simultaneous analysis of transcriptome and proteome at single‐cell level still faces huge challenges due to their differences in molecular modalities. Recent technological advances in single‐cell manipulations, barcoding, and ultrasensitive instrument recently offer unprecedented opportunities for the co‐profiling of genes and proteins. In this review, multiple types of single‐cell isolation, lysis, and molecular separation technologies are first introduced. Second, various approaches for co‐measurement of transcriptome and proteome in single‐cells are summarized, with their advantages, limitations, and capacity for targeted or unbiased deep analysis. Then we highlight the cutting‐edge spatial multiomics methodologies that operate at the single‐cell or subcellular resolution level, providing a comprehensive understanding of cell function and heterogeneity within the tissue spatial environment. The emerging biomedical applications of multiomics are also discussed. Finally, the challenges and prospects of this field are proposed.

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Time-resolved assessment of single-cell protein secretion by sequencing

Cited by 13 publications

References 72 publications

Single-Cell Secretion Analysis via Microfluidic Cell Membrane Immunosorbent Assay for Immune Profiling

Single-Cell Secretion Analysis via Microfluidic Cell Membrane Immunosorbent Assay for Immune Profiling

A Microfluidic 3D‐Tumor‐Spheroid Model for the Evaluation of Targeted Therapies from Angiogenesis‐Related Cytokines at the Single Spheroid Level

Integration of single‐cell transcriptome and proteome technologies: Toward spatial resolution levels

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