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Updated: Sep 6, 2025
Sample Preparation for Mass Cytometry Analysis
Published on: April 29, 2017
An End-to-End Workflow for Interrogating Tumor-Infiltrating Myeloid Cells Using Mass Cytometry
Maunish Barvalia1, Kenneth W Harder2
1Life Science Institute, Department of Microbiology and Immunology, University of British Columbia, Vancouver, BC, Canada.
Insights
This study details a mass cytometry workflow for deep phenotyping of tumor-infiltrating myeloid cells. It enables comprehensive analysis of myeloid cell heterogeneity and phosphoprotein interrogation in cancer research.
Area of Science:
- Immunology
- Cancer Biology
- Biotechnology
Background:
- Myeloid cells are key immune components with diverse roles in tumor progression.
- Specific myeloid subsets, like tumor-associated macrophages and myeloid-derived suppressive cells (MDSCs), promote tumor growth.
- Understanding myeloid cell heterogeneity is crucial for developing effective cancer immunotherapies.
Purpose of the Study:
- To provide a comprehensive end-to-end workflow for deep phenotyping of tumor-infiltrating myeloid cells.
- To enable detailed characterization of myeloid cell heterogeneity using mass cytometry.
- To facilitate the interrogation of phosphoproteins in myeloid cells within the tumor microenvironment.
Main Methods:
- Utilized mass cytometry for high-dimensional single-cell analysis of myeloid cells.
- Developed and validated an experimental protocol for sample preparation and staining.
- Implemented computational analysis including Phenograph clustering for myeloid cell subset identification.
Main Results:
- Established a robust workflow for deep phenotyping of tumor-infiltrating myeloid cells.
- Enabled comprehensive characterization of myeloid cell heterogeneity and functional states.
- Provided protocols for phosphoprotein analysis in circulating and tumor-infiltrating myeloid cells.
Conclusions:
- Mass cytometry offers an unprecedented opportunity to dissect myeloid cell heterogeneity in tumors.
- The provided workflow and computational scripts facilitate advanced myeloid cell profiling.
- This research supports the development of targeted cancer immunotherapies by elucidating myeloid cell functions.
Abstract:
Myeloid cells are a highly heterogeneous group of innate immune cells which include a diverse collection of cell types and cell states. Distinct subsets can impact tumor progression differently, with conventional type 1 DCs important in protective anti-tumor immune responses, while immunosuppressive tumor-associated macrophages and myeloid-derived suppressive cells (MDSCs) play tumor-promoting roles. Deep phenotyping of myeloid cells using single-cell technologies such as mass cytometry provides the unprecedented opportunity to comprehensively characterize the underlying heterogeneity of myeloid cells. Here we provide a detailed end-to-end workflow including both experimental and computational protocols enabling deep phenotyping of tumor-infiltrating myeloid cells using mass cytometry. A protocol that facilitates interrogation of phosphoproteins in circulating and tumor-infiltrating myeloid cells has been provided together with detailed scripts for Phenograph analysis of tumor-infiltrating myeloid cells.
