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Immunometabolism pp 31–45Cite as

The Conjugation of Antibodies for the Simultaneous Detection of Surface Proteins and Transcriptome Analysis at a Single-Cell Level

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 2184))

Abstract

Transcriptome analysis at a single-cell level with single-cell RNA sequencing (scRNA-seq) is a powerful method for detailed characterization of heterogeneous cell populations. Recent developments have enabled parallel analysis of both transcript and protein levels by using antibodies conjugated to barcoded oligonucleotides. These antibodies enable protein levels to be converted into nucleotide format, allowing the sequencing-based detection of both modalities at single-cell level. Here we present a simple and reliable method for conjugation of oligonucleotides with antibodies and a protocol for their use in single-cell transcriptome sequencing.

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References

  1. Chen H, Ye F, Guo G (2019) Revolutionizing immunology with single-cell RNA sequencing. Cell Mol Immunol 16:242–249

    Article  CAS  Google Scholar 

  2. Papalexi E, Satija R (2018) Single-cell RNA sequencing to explore immune cell heterogeneity. Nat Rev Immunol 18:35–45

    Article  CAS  Google Scholar 

  3. Vegh P, Haniffa M (2018) The impact of single-cell RNA sequencing on understanding the functional organization of the immune system. Brief Funct Genomics 17:265–272

    Article  CAS  Google Scholar 

  4. Azizi E, Carr AJ, Plitas G, Cornish AE, Konopacki C, Prabhakaran S et al (2018) Single-cell map of diverse immune phenotypes in the breast tumor microenvironment. Cell 174:1293–1308.e36

    Article  CAS  Google Scholar 

  5. Savas P, Virassamy B, Ye C, Salim A, Mintoff CP, Caramia F, Salgado R, Byrne DJ, Teo ZL, Dushyanthen S, Byrne A, Wein L, Luen SJ, Poliness C, Nightingale SS, Skandarajah AS, Gyorki DE, Thornton CM, Beavis PA, Fox SB, Kathleen Cuningham Foundation Consortium for Research into Familial Breast Cancer (kConFab), Darcy PK, Speed TP, Mackay LK, Neeson PJ, Loi S (2018) Single-cell profiling of breast cancer T cells reveals a tissue-resident memory subset associated with improved prognosis. Nat Med 24:986–993

    Article  CAS  Google Scholar 

  6. Villani A-C, Satija R, Reynolds G, Sarkizova S, Shekhar K, Fletcher J et al (2017) Single-cell RNA-seq reveals new types of human blood dendritic cells, monocytes, and progenitors. Science 356(6335):eaah4573

    Article  Google Scholar 

  7. Song Q, Hawkins GA, Wudel L, Chou P-C, Forbes E, Pullikuth AK et al (2019) Dissecting intratumoral myeloid cell plasticity by single cell RNA-seq. Cancer Med 8:3072–3085

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Peterson VM, Zhang KX, Kumar N, Wong J, Li L, Wilson DC, Moore R et al (2017) Multiplexed quantification of proteins and transcripts in single cells. Nat Biotechnol 35:936–939

    Article  CAS  Google Scholar 

  9. Vogel C, Marcotte EM (2012) Insights into the regulation of protein abundance from proteomic and transcriptomic analyses. Nat Rev Genet 13:227–232

    Article  CAS  Google Scholar 

  10. Liu Y, Beyer A, Aebersold R (2016) On the dependency of cellular protein levels on mRNA abundance. Cell 165:535–550

    Article  CAS  Google Scholar 

  11. Stoeckius M, Hafemeister C, Stephenson W, Houck-Loomis B, Chattopadhyay PK, Swerdlow H et al (2017) Simultaneous epitope and transcriptome measurement in single cells. Nat Methods 14:865–868

    Article  CAS  Google Scholar 

  12. Cossarizza A, Chang HD, Radbruch A et al (2017) Guidelines for the use of flow cytometry and cell sorting in immunological studies. Eur J Immunol 47:1584–1797

    Article  CAS  Google Scholar 

  13. De Simone M, Rossetti G, Pagani M (2019) Chromium 10× single-cell 3′ mRNA sequencing of tumor-infiltrating lymphocytes. Methods Mol Biol 1979:87–110

    Article  Google Scholar 

  14. Kotagiri N, Li Z, Xu X, Mondal S, Nehorai A, Achilefu S (2014) Antibody quantum dot conjugates developed via copper-free click chemistry for rapid analysis of biological samples using a microfluidic microsphere array system. Bioconjug Chem 25:1272–1281

    Article  CAS  Google Scholar 

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Acknowledgments

The Academy of Finland has supported EK (decision 308913) and TL (decision 311081). This work was supported by the Foundation for Pediatric Research in Finland and the Väre Foundation. Eliisa Kekäläinen, and Tapio Lönnberg contributed equally to this work.

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Author notes

  1. Eliisa Kekäläinen and Tapio Lönnberg contributed equally to this work.

Authors and Affiliations

  1. Translational Immunology Research Program, University of Helsinki, Helsinki, Finland

    Iivari Kleino & Eliisa Kekäläinen

  2. Department of Bacteriology and Immunology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland

    Iivari Kleino & Eliisa Kekäläinen

  3. Turku Bioscience Centre, University of Turku and Åbo Akademi University, Turku, Finland

    Tapio Lönnberg

Authors
  1. Iivari Kleino
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  2. Eliisa Kekäläinen
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  3. Tapio Lönnberg
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Corresponding author

Correspondence to Iivari Kleino .

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Editors and Affiliations

  1. Faculty of Health Sciences, Department of Internal Medicine, University of Manitoba, Winnipeg, MB, Canada

    Suresh Mishra

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Kleino, I., Kekäläinen, E., Lönnberg, T. (2020). The Conjugation of Antibodies for the Simultaneous Detection of Surface Proteins and Transcriptome Analysis at a Single-Cell Level. In: Mishra, S. (eds) Immunometabolism. Methods in Molecular Biology, vol 2184. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0802-9_3

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  • DOI: https://doi.org/10.1007/978-1-0716-0802-9_3

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0801-2

  • Online ISBN: 978-1-0716-0802-9

  • eBook Packages: Springer Protocols

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