SpringerLink
Log in

Mass Cytometry for the Characterization of Individual Cell Types in Ovarian Solid Tumors

  • Protocol
  • First Online:
Ovarian Cancer

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2424))

Abstract

Mass cytometry aka Cytometry by Time-Of-Flight (CyTOF) is one of several recently developed multiparametric single-cell technologies designed to address cellular heterogeneity within healthy and diseased tissue. Mass cytometry is an adaptation of flow cytometry in which antibodies are labeled with stable heavy metal isotopes and the readout is by time-of-flight mass spectrometry. With minimal spillover between channels, mass cytometry enables readouts of up to 60 parameters per single cell. Critically, mass cytometry can identify minority cell populations that are lost in bulk tissue analysis. Mass cytometry has been used to great effect for the study of immune cells. We have extended its use to examine single cells within disaggregated solid tissues, specifically freshly resected tubo-ovarian high-grade serous tumors. Here we detail our protocols designed to ensure the production of high-quality single-cell datasets. The methodology can be modified to accommodate the study of other solid tissues.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free ship** worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free ship** worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

Change history

  • 16 December 2021

    The legends for figures 7 and 8 in chapter 4 were incorrect and have been updated now.

References

  1. Leelatian N, Doxie DB, Greenplate AR, Mobley BC, Lehman JM, Sinnaeve J, Kauffmann RM, Werkhaven JA, Mistry AM, Weaver KD, Thompson RC, Massion PP, Hooks MA, Kelley MC, Chambless LB, Ihrie RA, Irish JM (2017) Single cell analysis of human tissues and solid tumors with mass cytometry. Cytometry B Clin Cytom 92:68–78. https://doi.org/10.1002/cyto.b.21481

    Article  CAS  PubMed  Google Scholar 

  2. Gonzalez VD, Huang Y-W, Chen S-Y, Delgado-Gonzalez A, Donoso K, Gentles A, Sachs K, Porpiglia E, Fantl WJ (2021) High-grade serous ovarian tumor cells modulate NK cell function to create an immune-tolerant microenvironment. Cell Rep 36:109632

    Google Scholar 

  3. Gonzalez VD, Samusik N, Chen TJ, Savig ES, Aghaeepour N, Quigley DA, Huang YW, Giangarra V, Borowsky AD, Hubbard NE, Chen SY, Han G, Ashworth A, Kipps TJ, Berek JS, Nolan GP, Fantl WJ (2018) Commonly occurring cell subsets in high-grade serous ovarian tumors identified by single-cell mass cytometry. Cell Rep 22:1875–1888. https://doi.org/10.1016/j.celrep.2018.01.053

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Bendall SC, Nolan GP (2012) From single cells to deep phenotypes in cancer. Nat Biotechnol 30:639–647. https://doi.org/10.1038/nbt.2283

    Article  CAS  PubMed  Google Scholar 

  5. Bendall SC, Simonds EF, Qiu P, Amir el AD, Krutzik PO, Finck R, Bruggner RV, Melamed R, Trejo A, Ornatsky OI, Balderas RS, Plevritis SK, Sachs K, Pe’er D, Tanner SD, Nolan GP (2011) Single-cell mass cytometry of differential immune and drug responses across a human hematopoietic continuum. Science 332:687–696. https://doi.org/10.1126/science.1198704

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Bjornson ZB, Nolan GP, Fantl WJ (2013) Single-cell mass cytometry for analysis of immune system functional states. Curr Opin Immunol 25:484–494. https://doi.org/10.1016/j.coi.2013.07.004

    Article  CAS  PubMed  Google Scholar 

  7. Spitzer MH, Nolan GP (2016) Mass cytometry: single cells, many features. Cell 165:780–791. https://doi.org/10.1016/j.cell.2016.04.019

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Tanner SD, Baranov VI, Ornatsky OI, Bandura DR, George TC (2013) An introduction to mass cytometry: fundamentals and applications. Cancer Immunol Immunother 62:955–965. https://doi.org/10.1007/s00262-013-1416-8

    Article  PubMed  Google Scholar 

  9. Chang Q, Hedley D (2012) Emerging applications of flow cytometry in solid tumor biology. Methods 57:359–367. https://doi.org/10.1016/j.ymeth.2012.03.027

    Article  CAS  PubMed  Google Scholar 

  10. Mistry AM, Greenplate AR, Ihrie RA, Irish JM (2019) Beyond the message: advantages of snapshot proteomics with single-cell mass cytometry in solid tumors. FEBS J 286:1523–1539. https://doi.org/10.1111/febs.14730

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Anandan S, Thomsen LCV, Gullaksen S-E, Abdelaal T, Kleinmanns K, Skavland J, Bredholt G, Gjertsen BT, McCormack E, Bjørge L (2021) Phenotypic characterization by mass cytometry of the microenvironment in ovarian cancer and impact of tumor dissociation methods. Cancers 13:755

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Han G, Spitzer MH, Bendall SC, Fantl WJ, Nolan GP (2018) Metal-isotope-tagged monoclonal antibodies for high-dimensional mass cytometry. Nat Protoc 13:2121–2148. https://doi.org/10.1038/s41596-018-0016-7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Han G, Chen SY, Gonzalez VD, Zunder ER, Fantl WJ, Nolan GP (2017) Atomic mass tag of bismuth-209 for increasing the immunoassay multiplexing capacity of mass cytometry. Cytometry A 91:1150–1163. https://doi.org/10.1002/cyto.a.23283

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Behbehani GK, Thom C, Zunder ER, Finck R, Gaudilliere B, Fragiadakis GK, Fantl WJ, Nolan GP (2014) Transient partial permeabilization with saponin enables cellular barcoding prior to surface marker staining. Cytometry A 85:1011–1019. https://doi.org/10.1002/cyto.a.22573

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Zunder ER, Finck R, Behbehani GK, Amir el AD, Krishnaswamy S, Gonzalez VD, Lorang CG, Bjornson Z, Spitzer MH, Bodenmiller B, Fantl WJ, Pe’er D, Nolan GP (2015) Palladium-based mass tag cell barcoding with a doublet-filtering scheme and single-cell deconvolution algorithm. Nat Protoc 10:316–333. https://doi.org/10.1038/nprot.2015.020

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Fienberg HG, Simonds EF, Fantl WJ, Nolan GP, Bodenmiller B (2012) A platinum-based covalent viability reagent for single-cell mass cytometry. Cytometry A 81:467–475. https://doi.org/10.1002/cyto.a.22067

    Article  CAS  PubMed  Google Scholar 

  17. Ornatsky OI, Lou X, Nitz M, Schäfer S, Sheldrick WS, Baranov VI, Bandura DR, Tanner SD (2008) Study of cell antigens and intracellular DNA by identification of element-containing labels and metallointercalators using inductively coupled plasma mass spectrometry. Anal Chem 80:2539–2547. https://doi.org/10.1021/ac702128m

    Article  CAS  PubMed  Google Scholar 

  18. Behbehani GK, Bendall SC, Clutter MR, Fantl WJ, Nolan GP (2012) Single-cell mass cytometry adapted to measurements of the cell cycle. Cytometry A 81:552–566. https://doi.org/10.1002/cyto.a.22075

    Article  PubMed  PubMed Central  Google Scholar 

  19. Ascenzi P, Bettinelli M, Boffi A, Botta M, De Simone G, Luchinat C, Marengo E, Mei H, Aime S (2020) Rare earth elements (REE) in biology and medicine. Rendiconti Lincei Scienze Fisiche e Naturali 31:821–833. https://doi.org/10.1007/s12210-020-00930-w

    Article  Google Scholar 

  20. Stern AD, Rahman AH, Birtwistle MR (2017) Cell size assays for mass cytometry. Cytometry A 91:14–24. https://doi.org/10.1002/cyto.a.23000

    Article  CAS  PubMed  Google Scholar 

  21. Willis LM, Park H, Watson MWL, Majonis D, Watson JL, Nitz M (2018) Tellurium-based mass cytometry barcode for live and fixed cells. Cytometry A 93:685–694. https://doi.org/10.1002/cyto.a.23495

    Article  CAS  PubMed  Google Scholar 

  22. Mei HE, Leipold MD, Maecker HT (2016) Platinum-conjugated antibodies for application in mass cytometry. Cytometry A 89:292–300. https://doi.org/10.1002/cyto.a.22778

    Article  CAS  PubMed  Google Scholar 

  23. Lou X, Zhang G, Herrera I, Kinach R, Ornatsky O, Baranov V, Nitz M, Winnik MA (2007) Polymer-based elemental tags for sensitive bioassays. Angew Chem Int Ed Eng 46:6111–6114. https://doi.org/10.1002/anie.200700796

    Article  CAS  Google Scholar 

  24. Majonis D, Herrera I, Ornatsky O, Schulze M, Lou X, Soleimani M, Nitz M, Winnik MA (2010) Synthesis of a functional metal-chelating polymer and steps toward quantitative mass cytometry bioassays. Anal Chem 82:8961–8969. https://doi.org/10.1021/ac101901x

    Article  CAS  PubMed  Google Scholar 

  25. Bendall SC, Nolan GP, Roederer M, Chattopadhyay PK (2012) A deep profiler’s guide to cytometry. Trends Immunol 33:323–332. https://doi.org/10.1016/j.it.2012.02.010

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Bandura DR, Baranov VI, Ornatsky OI, Antonov A, Kinach R, Lou X, Pavlov S, Vorobiev S, Dick JE, Tanner SD (2009) Mass cytometry: technique for real time single cell multitarget immunoassay based on inductively coupled plasma time-of-flight mass spectrometry. Anal Chem 81:6813–6822. https://doi.org/10.1021/ac901049w

    Article  CAS  PubMed  Google Scholar 

  27. Keyes TJ, Domizi P, Lo YC, Nolan GP, Davis KL (2020) A cancer biologist’s primer on machine learning applications in high-dimensional cytometry. Cytometry A 97:782–799. https://doi.org/10.1002/cyto.a.24158

    Article  PubMed  PubMed Central  Google Scholar 

  28. Olsen LR, Leipold MD, Pedersen CB, Maecker HT (2019) The anatomy of single cell mass cytometry data. Cytometry A 95:156–172. https://doi.org/10.1002/cyto.a.23621

    Article  PubMed  Google Scholar 

  29. Bodenmiller B, Zunder ER, Finck R, Chen TJ, Savig ES, Bruggner RV, Simonds EF, Bendall SC, Sachs K, Krutzik PO, Nolan GP (2012) Multiplexed mass cytometry profiling of cellular states perturbed by small-molecule regulators. Nat Biotechnol 30:858–867. https://doi.org/10.1038/nbt.2317

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Mei HE, Leipold MD, Schulz AR, Chester C, Maecker HT (2015) Barcoding of live human peripheral blood mononuclear cells for multiplexed mass cytometry. J Immunol 194:2022–2031. https://doi.org/10.4049/jimmunol.1402661

    Article  CAS  PubMed  Google Scholar 

  31. Van Gassen S, Gaudilliere B, Angst MS, Saeys Y, Aghaeepour N (2020) CytoNorm: a normalization algorithm for cytometry data. Cytometry A 97:268–278. https://doi.org/10.1002/cyto.a.23904

    Article  PubMed  Google Scholar 

  32. Domcke S, Sinha R, Levine DA, Sander C, Schultz N (2013) Evaluating cell lines as tumour models by comparison of genomic profiles. Nat Commun 4:2126. https://doi.org/10.1038/ncomms3126

    Article  CAS  PubMed  Google Scholar 

  33. Finck R, Simonds EF, Jager A, Krishnaswamy S, Sachs K, Fantl W, Pe’er D, Nolan GP, Bendall SC (2013) Normalization of mass cytometry data with bead standards. Cytometry A 83:483–494. https://doi.org/10.1002/cyto.a.22271

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Dasari S, Tchounwou PB (2014) Cisplatin in cancer therapy: molecular mechanisms of action. Eur J Pharmacol 740:364–378. https://doi.org/10.1016/j.ejphar.2014.07.025

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Hartmann FJ, Simonds EF, Vivanco N, Bruce T, Borges L, Nolan GP, Spitzer MH, Bendall SC (2019) Scalable conjugation and characterization of immunoglobulins with stable mass isotope reporters for single-cell mass cytometry analysis. Methods Mol Biol 1989:55–81. https://doi.org/10.1007/978-1-4939-9454-0_5

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Pichaandi J, Zhao G, Bouzekri A, Lu E, Ornatsky O, Baranov V, Nitz M, Winnik MA (2019) Lanthanide nanoparticles for high sensitivity multiparameter single cell analysis. Chem Sci 10:2965–2974. https://doi.org/10.1039/c8sc04407d

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Illy N, Majonis D, Herrera I, Ornatsky O, Winnik MA (2012) Metal-chelating polymers by anionic ring-opening polymerization and their use in quantitative mass cytometry. Biomacromolecules 13:2359–2369. https://doi.org/10.1021/bm300613x

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors thank Professor Scott Tanner, the inventor of mass cytometry, for his critical reading of the manuscript. Furthermore, we recognize the outstanding contribution Professor Tanner and his team have made to the advancement of biomedical sciences. We thank Shawn Hollahan at paradigm_shiftin@mac.com for artwork, Dr. Zach Bjornson, and Astraea Jaeger for critical reading of the protocols.

This work was supported by funding from Department of Defense W81XWH-14-1-0180, NCI R21CA231280, NCI 1R01CA234553-01A1, NHLBI P01HL10879709, NIAID U1911057229, The 2019 Cancer Innovation Award, supported by the Stanford Cancer Institute, an NCI-designated Comprehensive Cancer Center, The Department of Urology at Stanford University and The BRCA Foundation and the V Foundation for Cancer Research and a gift from the Gray Foundation, and PICI Bedside to Bench grant.

Author information

Author notes

  1. Veronica D. Gonzalez

    Present address: 10X Genomics, Pleasanton, CA, USA

Authors and Affiliations

  1. Baxter Laboratory for Stem Cell Biology, Department of Microbiology & Immunology, Stanford University School of Medicine, Stanford, CA, USA

    Veronica D. Gonzalez

  2. Department of Urology, Stanford University School of Medicine, Stanford, CA, USA

    Ying-Wen Huang

  3. Department of Urology, Department of Obstetrics and Gynecology, Stanford Comprehensive Cancer Institute, Stanford, CA, USA

    Wendy J. Fantl

Authors
  1. Veronica D. Gonzalez
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ying-Wen Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Wendy J. Fantl
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Wendy J. Fantl .

Editor information

Editors and Affiliations

  1. Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI, USA

    Pamela K. Kreeger

Rights and permissions

Reprints and permissions

Copyright information

© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Gonzalez, V.D., Huang, YW., Fantl, W.J. (2022). Mass Cytometry for the Characterization of Individual Cell Types in Ovarian Solid Tumors. In: Kreeger, P.K. (eds) Ovarian Cancer. Methods in Molecular Biology, vol 2424. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1956-8_4

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-1956-8_4

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1955-1

  • Online ISBN: 978-1-0716-1956-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation