scholarly journals DART‐ID Increases Single‐Cell Proteome Coverage

2019 ◽  
Vol 33 (S1) ◽  
Author(s):  
Albert Tian Chen ◽  
Alexander Franks ◽  
Nikolai Slavov
Keyword(s):  
Single Cell ◽  
Proteome Coverage ◽  
Cell Proteome

scholarly journals High-throughput and high-efficiency sample preparation for single-cell proteomics using a nested nanowell chip

2021 ◽  
Author(s):  
Jongmin Woo ◽  
Sarah M. Williams ◽  
Victor Aguilera-Vazquez ◽  
Ryan L. Sontag ◽  
Ronald J. Moore ◽  
...  
Keyword(s):  
Single Cell ◽  
High Efficiency ◽  
Single Cells ◽  
Protein Recovery ◽  
Processing Capacity ◽  
Protein Abundance ◽  
Proteome Coverage ◽  
Low Protein ◽  
Functional Differences ◽  
Cell Proteome

AbstractGlobal quantification of protein abundances in single cells would provide more direct information on cellular function phenotypes and complement transcriptomics measurements. However, single-cell proteomics (scProteomics) is still immature and confronts technical challenges, including limited proteome coverage, poor reproducibility, as well as low throughput. Here we describe a nested nanoPOTS (N2) chip to dramatically improve protein recovery, operation robustness, and processing throughput for isobaric-labeling-based scProteomics workflow. The N2 chip allows reducing cell digestion volume to <30 nL and increasing processing capacity to > 240 single cells in one microchip. In the analysis of ∼100 individual cells from three different cell lines, we demonstrate the N2 chip-based scProteomics platform can robustly quantify ∼1500 proteins and reveal functional differences. Our analysis also reveals low protein abundance variations (median CVs < 16.3%), highlighting the utility of such measurements, and also suggesting the single-cell proteome is highly stable for the cells cultured under identical conditions.

scholarly journals DART-ID increases single-cell proteome coverage

2019 ◽  
Vol 15 (7) ◽  
pp. e1007082 ◽  
Author(s):  
Albert Tian Chen ◽  
Alexander Franks ◽  
Nikolai Slavov
Keyword(s):  
Single Cell ◽  
Proteome Coverage ◽  
Cell Proteome

scholarly journals Improved Single-Cell Proteome Coverage Using Narrow-Bore Packed NanoLC Columns and Ultrasensitive Mass Spectrometry

2020 ◽  
Vol 92 (3) ◽  
pp. 2665-2671 ◽  
Author(s):  
Yongzheng Cong ◽  
Yiran Liang ◽  
Khatereh Motamedchaboki ◽  
Romain Huguet ◽  
Thy Truong ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Single Cell ◽  
Proteome Coverage ◽  
Cell Proteome

scholarly journals Improved proteome coverage by using high efficiency cysteinyl peptide enrichment: The human mammary epithelial cell proteome

PROTEOMICS ◽  
2005 ◽  
Vol 5 (5) ◽  
pp. 1263-1273 ◽  
Author(s):  
Tao Liu ◽  
Wei-Jun Qian ◽  
Wan-Nan U. Chen ◽  
Jon M. Jacobs ◽  
Ronald J. Moore ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Mammary Epithelial Cell ◽  
High Efficiency ◽  
Human Mammary Epithelial Cell ◽  
Mammary Epithelial ◽  
Proteome Coverage ◽  
Human Mammary Epithelial ◽  
Peptide Enrichment ◽  
Cell Proteome

scholarly journals Ultrasensitive single-cell proteomics workflow identifies >1000 protein groups per mammalian cell

2021 ◽  
Author(s):  
Yongzheng Cong ◽  
Khatereh Motamedchaboki ◽  
Santosh A. Misal ◽  
Yiran Liang ◽  
Amanda J. Guise ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Single Cell ◽  
Mammalian Cell ◽  
Ion Mobility Spectrometry ◽  
Ion Mobility ◽  
High Field ◽  
Low Flow ◽  
Proteome Profiling ◽  
Cell Proteome

The combination of nanodroplet sample preparation, ultra-low-flow nanoLC, high-field asymmetric ion mobility spectrometry (FAIMS) and latest-generation mass spectrometry instrumentation provides dramatically improved single-cell proteome profiling.

scholarly journals Global absolute quantification reveals tight regulation of protein expression in single Xenopus eggs

2014 ◽  
Vol 42 (15) ◽  
pp. 9880-9891 ◽  
Author(s):  
Arne H. Smits ◽  
Rik G.H. Lindeboom ◽  
Matteo Perino ◽  
Simon J. van Heeringen ◽  
Gert Jan C. Veenstra ◽  
...  
Keyword(s):  
Single Cell ◽  
Ribosomal Proteins ◽  
Single Cells ◽  
Transcript Level ◽  
Absolute Quantification ◽  
Protein Quantification ◽  
Protein Levels ◽  
Recent Developments ◽  
Basal Transcription Factors ◽  
Cell Proteome

Abstract While recent developments in genomic sequencing technology have enabled comprehensive transcriptome analyses of single cells, single cell proteomics has thus far been restricted to targeted studies. Here, we perform global absolute protein quantification of fertilized Xenopus laevis eggs using mass spectrometry-based proteomics, quantifying over 5800 proteins in the largest single cell proteome characterized to date. Absolute protein amounts in single eggs are highly consistent, thus indicating a tight regulation of global protein abundance. Protein copy numbers in single eggs range from tens of thousands to ten trillion copies per cell. Comparison between the single-cell proteome and transcriptome reveal poor expression correlation. Finally, we identify 439 proteins that significantly change in abundance during early embryogenesis. Downregulated proteins include ribosomal proteins and upregulated proteins include basal transcription factors, among others. Many of these proteins do not show regulation at the transcript level. Altogether, our data reveal that the transcriptome is a poor indicator of the proteome and that protein levels are tightly controlled in X. laevis eggs.

scholarly journals Ultra-streamlined single cell proteomics by all-in-one chip and data-independent-acquisition mass spectrometry

2021 ◽  
Author(s):  
Hsiung-Lin Tu ◽  
Sofani Gebreyesus ◽  
Asad Ali Siyal ◽  
Reta Birhanu Kitata ◽  
Eric Sheng-Wen Chen ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Single Cell ◽  
Missing Values ◽  
Protein Quantification ◽  
Single Cell Level ◽  
Cell Level ◽  
Proteome Coverage ◽  
Data Independent Acquisition ◽  
General Utility ◽  
Ultrahigh Sensitivity

Abstract Single cell proteomics provides the ultimate resolution to reveal cellular phenotypic heterogeneity and functional network underlying biological processes. Here, we present an ultra-streamlined workflow combining an integrated proteomic chip (iProChip) and data-independent-acquisition (DIA) mass spectrometry for sensitive microproteomics analysis down to single cell level. The iProChip offers multiplexed and automated all-in-one station from cell isolation/counting/imaging to complete proteomic processing within a single device. By mapping to project-specific spectra libraries, the iProChip-DIA enables profiling of 1160 protein groups from triplicate analysis of a single mammalian cell. Furthermore, the applicability of iProChip-DIA was demonstrated using both adherent and non-adherent malignant cells, which reveals 5 orders of proteome coverage, highly consistent ~100-fold protein quantification (1-100 cells) and high reproducibility with low missing values (<16%). With the demonstrated all-in-one cell characterization, ultrahigh sensitivity, robustness, and versatility to add other functionalities, the iProChip-DIA is anticipated to offer general utility to realize advanced proteomics applications at single cell level.

scholarly journals High-throughput and high-efficiency sample preparation for single-cell proteomics using a nested nanowell chip

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jongmin Woo ◽  
Sarah M. Williams ◽  
Lye Meng Markillie ◽  
Song Feng ◽  
Chai-Feng Tsai ◽  
...  
Keyword(s):  
Single Cell ◽  
High Efficiency ◽  
Single Cells ◽  
Protein Recovery ◽  
Protein Abundance ◽  
Protein Markers ◽  
Low Protein ◽  
Tandem Mass Tag ◽  
Cellular Phenotypes ◽  
Cell Proteome

AbstractGlobal quantification of protein abundances in single cells could provide direct information on cellular phenotypes and complement transcriptomics measurements. However, single-cell proteomics is still immature and confronts many technical challenges. Herein we describe a nested nanoPOTS (N2) chip to improve protein recovery, operation robustness, and processing throughput for isobaric-labeling-based scProteomics workflow. The N2 chip reduces reaction volume to <30 nL and increases capacity to >240 single cells on a single microchip. The tandem mass tag (TMT) pooling step is simplified by adding a microliter droplet on the nested nanowells to combine labeled single-cell samples. In the analysis of ~100 individual cells from three different cell lines, we demonstrate that the N2 chip-based scProteomics platform can robustly quantify ~1500 proteins and reveal membrane protein markers. Our analyses also reveal low protein abundance variations, suggesting the single-cell proteome profiles are highly stable for the cells cultured under identical conditions.

scholarly journals Ultra-streamlined single cell proteomics by all-in-one chip and data-independent-acquisition mass spectrometry

2021 ◽  
Author(s):  
Hsiung-Lin Tu ◽  
Sofani Gebreyesus ◽  
Asad Ali Siyal ◽  
Reta Birhanu Kitata ◽  
Eric Sheng-Wen Chen ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Single Cell ◽  
Missing Values ◽  
Protein Quantification ◽  
Single Cell Level ◽  
Cell Level ◽  
Proteome Coverage ◽  
Data Independent Acquisition ◽  
General Utility ◽  
Ultrahigh Sensitivity

Abstract Single cell proteomics provides the ultimate resolution to reveal cellular phenotypic heterogeneity and functional network underlying biological processes. Here, we present an ultra-streamlined workflow combining an integrated proteomic chip (iProChip) and data-independent-acquisition (DIA) mass spectrometry for sensitive microproteomics analysis down to single cell level. The iProChip offers multiplexed and automated all-in-one station from cell isolation/counting/imaging to complete proteomic processing within a single device. By mapping to project-specific spectra libraries, the iProChip-DIA enables profiling of 1160 protein groups from triplicate analysis of a single mammalian cell. Furthermore, the applicability of iProChip-DIA was demonstrated using both adherent and non-adherent malignant cells, which reveals 5 orders of proteome coverage, highly consistent ~100-fold protein quantification (1-100 cells) and high reproducibility with low missing values (<16%). With the demonstrated all-in-one cell characterization, ultrahigh sensitivity, robustness, and versatility to add other functionalities, the iProChip-DIA is anticipated to offer general utility to realize advanced proteomics applications at single cell level.

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