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BD Rhapsody™ Single-Cell Analysis System Workflow: From Sample to Multimodal Single-Cell Sequencing Data

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Single Cell Transcriptomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2584))

Abstract

Advancements in single-cell sequencing have revolutionized our understanding of complex biological systems such as the immune system and allowed us to overcome limitations in various disciplines of life science research such as oncology, developmental biology, or neurobiology (Perkel, Nature 595. https://www.nature.com/articles/d41586-021-01994-w, 2021).

The BD Rhapsody™ Single-Cell Analysis System enables us to capture multimodal information from thousands of single cells in parallel (“Multiomics”) covering mRNA expression levels, protein expression levels, the immune repertoire for T-cell receptors (TCR) and B-cell receptors (BCR), and the identification of antigen-specific T cells and B cells using dCODE Dextramer® (RiO) from Immudex. The system utilizes microwell-based cartridges that allow to capture a broad range of single cells and an imaging device for sample quality control and workflow quality control (including viability and multiplets). The power of Multiomics relies on simultaneously measuring several aspects of single cells, including gene expression and protein abundance, using next generation sequencing (NGS) as a single readout.

Here we describe the complete BD Rhapsody™ Single-Cell Analysis System from the sample preparation including different options for the antibody and/or dCODE Dextramer® staining through to the data analysis.

For updated protocols, guides, and technical bulletins, please visit the BD Scomix page: https://scomix.bd.com/hc/en-us or the BDB webpage: https://www.bdbiosciences.com/en-eu.

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References

  1. Buus TB, Herrera A, Ivanova E, Mimitou E, Cheng A, Herati RS, Papagiannakopoulos T, Smibert P, Odum N, Koralov SB (2021) Improving oligo-conjugated antibody signal in multimodal single-cell analysis. eLife 10:e61973. https://doi.org/10.7554/eLife.61973

    Article  CAS  Google Scholar 

  2. Fan HC, Fu GK, Fodor SPA (2015) Combinatorial labeling of single cells for gene expression cytometry. Science 347:1258367. https://doi.org/10.1126/science.1258367

    Article  CAS  Google Scholar 

  3. Fu GK, Hu J, Wang PH, Fodor SP (2011) Counting individual DNA molecules by the stochastic attachment of diverse labels. Proc Natl Acad Sci U S A 108(22):9026–9031. https://doi.org/10.1073/pnas.1017621108

    Article  Google Scholar 

  4. Georg P, Astaburuaga-García R, Bonaguro L, Brumhard S, Michalick L, Lippert LJ, Kostevc T, Gäbel C, Schneider M, Streitz M, Demichev V, Gemünd I, Barone M, Tober-Lau P, Helbig ET, Hillus D, Petrov L, Stein J, Dey HP, Paclik D, et al; PA-COVID-19 Study Group (2022) Complement activation induces excessive T cell cytotoxicity in severe COVID-19. Cell 185(3):493–512.e25. https://doi.org/10.1016/j.cell.2021.12.040

    Article  CAS  Google Scholar 

  5. Haque A, Engel J, Teichmann SA, Lönnberg T (2017) A practical guide to single-cell RNA-sequencing for biomedical research and clinical applications. Genome Med 9(1):75. https://doi.org/10.1186/s13073-017-0467-4

    Article  CAS  Google Scholar 

  6. Huang J, Zhu Z, Ji D, Sun R, Yang Y, Liu L, Shao Y, Chen Y, Li L, Sun B (2022) Single-cell transcriptome profiling reveals neutrophil heterogeneity and functional multiplicity in the early stage of severe burn patients. Front Immunol 12:792122. https://doi.org/10.3389/fimmu.2021.792122

    Article  CAS  Google Scholar 

  7. Perkel JM (2021) Single-cell analysis enters the multiomics age. Nature 595. https://www.nature.com/articles/d41586-021-01994-w

  8. Junius S, Mavrogiannis AV, Lemaitre P, Gerbaux M, Staels F, Malviya V, Burton O, Gergelits V, Singh K, Tito Tadeo RY, Raes J, Humblet-Baron S, Liston A, Schlenner SM (2021) Unstable regulatory T cells, enriched for naïve and Nrp1neg cells, are purged after fate challenge. Sci Immunol 6(61). https://www.science.org/doi/10.1126/sciimmunol.abe4723?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub%20%200pubmed

  9. Ma KY, Schonnesen AA, He C, Xia AY, Sun E, Chen E, Sebastian KR, Guo YW, Balderas R, Kulkarni-Date M, Jiang N (2021) High-throughput and high-dimensional single-cell analysis of antigen-specific CD8+ T cells. Nat Immunol 22(12):1590–1598. https://doi.org/10.1038/s41590-021-01073-2

    Article  CAS  Google Scholar 

  10. Russo V, Brasu N, Pace L (2022) Combined measurement of RNA and protein expression on a single-cell level. Methods Mol Biol (Clifton, N.J.) 2386:263–288. https://doi.org/10.1007/978-1-0716-1771-7_16

    Article  Google Scholar 

  11. Saigusa R, Ley K (2020) CITE-Seq hits vascular medicine. Clin Chem 66(6):751–753. https://doi.org/10.1093/clinchem/hvaa016

    Article  Google Scholar 

  12. Schulte-Schrepping J, Reusch N, Paclik D et al (2020) Severe COVID-19 is marked by a dysregulated myeloid cell compartment. Cell 182(6):1419–1440.e23. https://doi.org/10.1016/j.cell.2020.08.001

    Article  CAS  Google Scholar 

  13. Siwiak M, Zielenkiewicz P (2010) A comprehensive, quantitative, and genome-wide model of translation. PLoS Comput Biol 6(7):e1000865. https://doi.org/10.1371/journal.pcbi.1000865

    Article  CAS  Google Scholar 

  14. Triana S, Vonficht D, Jopp-Saile L, Raffel S, Lutz R, Leonce D, Antes M, Hernández-Malmierca P, Ordoñez-Rueda D, Ramasz B, Boch T, Jann JC, Nowak D, Hofmann WK, Müller-Tidow C, Hübschmann D, Alexandrov T, Benes V, Trumpp A, Paulsen M et al (2021) Single-cell proteo-genomic reference maps of the hematopoietic system enable the purification and massive profiling of precisely defined cell states. Nat Immunol 22(12):1577–1589. https://doi.org/10.1038/s41590-021-01059-0

    Article  CAS  Google Scholar 

  15. Trzupek D, Dunstan M, Cutler AJ, Lee M, Godfrey L, Jarvis L, Rainbow DB, Aschenbrenner D, Jones JL, Uhlig HH, Wicker LS, Todd JA, Ferreira RC (2020) Discovery of CD80 and CD86 as recent activation markers on regulatory T cells by protein-RNA single-cell analysis. Genome Med 12(1):55. https://doi.org/10.1186/s13073-020-00756-z

    Article  CAS  Google Scholar 

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Author information

Authors and Affiliations

  1. BD Biosciences GmbH, Heidelberg, Germany

    Jannes Ulbrich, Vadir Lopez-Salmeron & Ian Gerrard

Authors
  1. Jannes Ulbrich
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  2. Vadir Lopez-Salmeron
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  3. Ian Gerrard
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Corresponding author

Correspondence to Jannes Ulbrich .

Editor information

Editors and Affiliations

  1. Molecular Biotechnology Center, University of Torino, Torino, Italy

    Raffaele A. Calogero

  2. European Molecular Biology Laboratory, Heidelberg, Germany

    Vladimir Benes

Glossary

BAM

An alignment file in binary format. A binary SAM file.

Bioproduct

Identifiers for biologically derived products such as mRNA and protein. Examples of identifiers are gene name for mRNA or AbSeq identifier for Abseq.

Bioproduct Type

Type of bioproducts such as mRNA or Abseq.

CIGAR

Compact Idiosyncratic Gapped Alignment Report. A sequence of base lengths to indicate base alignments, insertions, and deletions with respect to the reference sequence.

CLS

Cell label sequence.

DBEC

Distribution-based error correction.

FASTA

Text-based format that contains one or more DNA or RNA sequences.

FASTQ

A file in standardized, text-based format that contains the output of read bases and per-base quality values from a sequencer.

L

Common sequence

Molecule

A unique combination of a cell label, UMI sequence, and a bioproduct. Without UMI adjustment methods, it is called raw molecule. With RSEC UMI adjustment, it is called RSEC-adjusted molecule. With additional DBEC UMI adjustment, it is called DBEC-adjusted molecule.

PhiX

Control library used for sequencing runs.

R1 reads

Contains information about the cell label and UMI.

R2 reads

Contains information about the bioproduct.

RSEC

Recursive substitution error correction.

SAM

Tab-delimited text file with sequence alignment data.

Singlet

A putative cell where more than 75% of sample tag reads are from a single tag.

Singleton

Clustering: Cell not assigned to any of the clusters. UMI correction/adjustment: Molecule that is represented by only one read.

UMI

Unique molecular identifier. A string of eight randomers immediately downstream of the cell label sequence (CLS) 3 of the R1 read that is used to uniquely label a molecule.

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Ulbrich, J., Lopez-Salmeron, V., Gerrard, I. (2023). BD Rhapsody™ Single-Cell Analysis System Workflow: From Sample to Multimodal Single-Cell Sequencing Data. In: Calogero, R.A., Benes, V. (eds) Single Cell Transcriptomics. Methods in Molecular Biology, vol 2584. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2756-3_2

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  • DOI: https://doi.org/10.1007/978-1-0716-2756-3_2

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2755-6

  • Online ISBN: 978-1-0716-2756-3

  • eBook Packages: Springer Protocols

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