Multiplexed detection of proteins, transcriptomes, clonotypes and CRISPR perturbations in single cells

Mimitou, Eleni P.; Cheng, Anthony; Montalbano, Antonino; Hao, Stephanie; Stoeckius, Marlon; Legut, Mateusz; Roush, Timothy; Herrera, Alberto; Papalexi, Efthymia; Ouyang, Zhengqing; Satija, Rahul; Sanjana, Neville E.; Koralov, Sergei B.; Smibert, Peter

doi:10.1038/s41592-019-0392-0

Cited by 426 publications

(374 citation statements)

References 37 publications

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“…Recently, two similar approaches, CITE-seq (9) and REAP-seq (10), have been described to measure protein expression using oligo-conjugated antibodies in parallel with scRNA-seq data. Furthermore, other applications are currently being developed to integrate the growing portfolio of single-cell omics technologies (35,36). A fundamental difference with the approach described in this study is that these technologies all rely on whole-transcriptome data, providing a high-level cross-sectional representation of all polyA transcripts in the cell.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, other applications are currently being developed to integrate the growing portfolio of single-cell omics technologies [30,31]. A fundamental difference with the approach described in this study is that these technologies all rely on whole-transcriptome data, providing a high-level cross-sectional representation of all polyA mRNA transcripts in the cell.…”

Section: Discussionmentioning

confidence: 99%

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Discovery of CD80 and CD86 as recent activation markers on regulatory T cells by protein-RNA single-cell analysis

Trzupek

Dunstan

Cutler

et al. 2019

Preprint

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The transcriptomic and proteomic characterisation of CD4 + T cells at the single-cell level has been performed traditionally by two largely exclusive types of technologies: single cell RNAsequencing (scRNA-seq) technologies and antibody-based cytometry. Here we demonstrate that the simultaneous targeted quantification of mRNA and protein expression in single-cells provides a high-resolution map of human primary CD4 + T cells, and reveals precise trajectories of Th1, Th17 and regulatory T-cell differentiation in blood and tissue. We report modest correlation between mRNA and protein in primary CD4 + T cells at the single-cell level, highlighting the value of protein expression data to characterise cell effector function. This transcriptomic and proteomic hybrid technology provides a massively-parallel, cost-effective, solution to dissect the heterogeneity of immune cell populations and to immunophenotype rare and potentially pathogenic immune subsets, and could have important clinical applications to redefine differentiation and activation of circulating and tissue-resident human immune cells.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Discovery of CD80 and CD86 as recent activation markers on regulatory T cells by protein-RNA single-cell analysis

Trzupek

Dunstan

Cutler

et al. 2019

Preprint

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“…[6]) -are rapidly evolving, resulting in continuous churn in the field as new techniques and applications come on the scene (e.g. [7,8]). As the volume of data and diversity of applications continue to grow, so does the number of software libraries and tools for the analysis and visualization of these datasets.…”

Section: Introductionmentioning

confidence: 99%

NASQAR: A web-based platform for high-throughput sequencing data analysis and visualization

Yousif

Drou

Rowe

et al. 2019

Preprint

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Background: As high-throughput sequencing applications continue to evolve, the rapid growth in quantity and variety of sequence-based data calls for the development of new software libraries and tools for data analysis and visualization. Often, effective use of these tools requires computational skills beyond those of many researchers. To lower this computational barrier, we have created a dynamic web-based platform, NASQAR (Nucleic Acid SeQuence Analysis Resource). Results: NASQAR offers a collection of custom and publicly available open-source web applications that make extensive use of the Shiny R package to provide interactive data visualization. The platform is publicly accessible at http://nasqar.abudhabi.nyu.edu/. NASQAR is open-source and the code is available through GitHub at https://github.com/nasqar/NASQAR. NASQAR is also available as a Docker image at https://hub.docker.com/r/aymanm/nasqarall. NASQAR is a collaboration between the core bioinformatics teams of the NYU Abu Dhabi and NYU New York Centers for Genomics and Systems Biology.Conclusions: NASQAR provides an intuitive interface that allows users to easily and efficiently explore their data in an interactive way using popular tools for a variety of applications, including Transcriptome Data Preprocessing, RNAseq Analysis (including Single-cell RNAseq), Metagenomics, and Gene Enrichment.

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“…Focusing on a common human allele, HLA-A*02:01, we have extended the capability of our system by incorporating our multi-modal cellular indexing technology (ECCITE-seq) 18,19 .…”

mentioning

confidence: 99%

“…To link pMHC specificities with TCR V(D)J sequences present in polyclonal samples, we barcoded fluorophore-labelled pMHC tetramers, prepared using TAPBPR exchange, with biotinylated DNA oligonucleotides (oligos) 25 . We used an inhouse oligo design compatible with 10x Genomics gel bead oligos in the 5' V(D)J product ( Figure 1 and Supplementary Fig 6a) adding an additional modality of cellular information to our recently described ECCITE-seq method 18 . This method incorporates a cellular barcode into cDNA generated from both tetramer oligos and TCR mRNA, thus the pairing of cellular barcodes can connect TCR sequences and other mRNAs, with tetramer specificities.…”

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confidence: 99%

High Throughput pMHC-I Tetramer Library Production Using Chaperone Mediated Peptide Exchange

Overall

Toor

Hao

et al. 2019

Preprint

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ABSTRACTPeptide exchange technologies are essential for the generation of pMHC-multimer libraries, used to probe highly diverse, polyclonal TCR repertoires. Using the molecular chaperone TAPBPR, we present a robust method for the capture of stable, empty MHC-I molecules which can be readily tetramerized and loaded with peptides of choice in a high-throughput manner. Combined with tetramer barcoding using multi-modal cellular indexing technology (ECCITE-seq), our approach allows a combined analysis of TCR repertoires and other T-cell transcription profiles together with their cognate pMHC-I specificities in a single experiment.

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Multiplexed detection of proteins, transcriptomes, clonotypes and CRISPR perturbations in single cells

Cited by 426 publications

References 37 publications

Discovery of CD80 and CD86 as recent activation markers on regulatory T cells by protein-RNA single-cell analysis

Discovery of CD80 and CD86 as recent activation markers on regulatory T cells by protein-RNA single-cell analysis

NASQAR: A web-based platform for high-throughput sequencing data analysis and visualization

High Throughput pMHC-I Tetramer Library Production Using Chaperone Mediated Peptide Exchange

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