Cell surface profiling using high-throughput flow cytometry: a platform for biomarker discovery and analysis of cellular heterogeneity

Craig A Gedye, Ali Hussain, Joshua Paterson, Alannah Smrke, Harleen Saini, Danylo Sirskyj, Keira Pereira, Nazleen Lobo, Jocelyn Stewart, Christopher Go, Jenny Ho, Mauricio Medrano, Elzbieta Hyatt, Julie Yuan, Stevan Lauriault, Mona Meyer, Maria Kondratyev, Twan van den Beucken, Michael Jewett, Peter DirksCynthia J Guidos, Jayne Danska, Jean Wang, Bradly Wouters, Benjamin Neel, Robert Rottapel, Laurie E Ailles

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Cell surface proteins have a wide range of biological functions, and are often used as lineage-specific markers. Antibodies that recognize cell surface antigens are widely used as research tools, diagnostic markers, and even therapeutic agents. The ability to obtain broad cell surface protein profiles would thus be of great value in a wide range of fields. There are however currently few available methods for high-throughput analysis of large numbers of cell surface proteins. We describe here a high-throughput flow cytometry (HT-FC) platform for rapid analysis of 363 cell surface antigens. Here we demonstrate that HT-FC provides reproducible results, and use the platform to identify cell surface antigens that are influenced by common cell preparation methods. We show that multiple populations within complex samples such as primary tumors can be simultaneously analyzed by co-staining of cells with lineage-specific antibodies, allowing unprecedented depth of analysis of heterogeneous cell populations. Furthermore, standard informatics methods can be used to visualize, cluster and downsample HT-FC data to reveal novel signatures and biomarkers. We show that the cell surface profile provides sufficient molecular information to classify samples from different cancers and tissue types into biologically relevant clusters using unsupervised hierarchical clustering. Finally, we describe the identification of a candidate lineage marker and its subsequent validation. In summary, HT-FC combines the advantages of a high-throughput screen with a detection method that is sensitive, quantitative, highly reproducible, and allows in-depth analysis of heterogeneous samples. The use of commercially available antibodies means that high quality reagents are immediately available for follow-up studies. HT-FC has a wide range of applications, including biomarker discovery, molecular classification of cancers, or identification of novel lineage specific or stem cell markers.

Original languageEnglish
Article numbere105602
JournalPLOS ONE
Volume9
Issue number8
DOIs
Publication statusPublished - 2014

Keywords

  • Antigens, Surface
  • Biomarkers
  • Cell Line, Tumor
  • Cells, Cultured
  • Cluster Analysis
  • Flow Cytometry
  • Humans
  • Jurkat Cells
  • MCF-7 Cells
  • Microscopy, Fluorescence
  • Proteome
  • Proteomics
  • Reproducibility of Results

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