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High-throughput mass cytometry staining for deep phenotyping of human natural killer cells
Amira Ben Amara1, Marie-Sarah Rouviere1, Stéphane Fattori1
1Immunomonitoring Department, Institut Paoli-Calmettes, 13009 Marseille, France; Team Immunity and Cancer, Centre de Recherche en Cancérologie de Marseille (CRCM), Inserm, U1068, CNRS, UMR7258, Institut Paoli-Calmettes, Aix-Marseille University, UM105, 13009 Marseille, France.
Insights
This protocol outlines immune phenotyping of natural killer (NK) cells in blood samples using mass cytometry. It details extracellular and intracellular staining for detailed NK cell subset analysis.
Area of Science:
- Immunology
- Cell Biology
- Biotechnology
Background:
- Natural killer (NK) cells are crucial immune cells involved in innate immunity.
- Detailed phenotyping of NK cells is essential for understanding immune responses and diseases.
- Current methods may require optimization for comprehensive analysis of clinical samples.
Purpose of the Study:
- To provide a detailed protocol for immune phenotyping of peripheral blood NK cells.
- To enable high-dimensional analysis of NK cell subsets using mass cytometry.
- To facilitate the study of NK cell characteristics in clinical settings.
Main Methods:
- The protocol involves peripheral blood mononuclear cell (PBMC) isolation.
- Extracellular and intracellular staining using metal-conjugated antibodies.
- Sample acquisition and analysis on a mass cytometer.
Main Results:
- The protocol allows for in-depth immune phenotyping of NK cells.
- High-dimensional analysis visualizes distinct NK cell subsets and their phenotypes.
- This method is applicable to clinical samples.
Conclusions:
- This mass cytometry protocol provides a robust method for NK cell immune phenotyping.
- It enables detailed characterization of NK cell populations.
- The protocol supports research in immunology and clinical diagnostics.
Abstract:
This protocol details the step-by-step procedure for in-depth immune phenotyping of peripheral blood natural killer (NK) cells from clinical samples by mass cytometry. The protocol consists of three main steps: PBMC incubation with a mix of metal-conjugated antibodies for extracellular phenotyping followed by fixation, permeabilization and incubation with a mix of metal-conjugated antibodies for staining of intracellular proteins, and sample acquisition on a mass cytometer. High-dimensional analysis enables the visualization of NK cell subsets and their phenotypical characteristics. For complete details on the use and execution of this protocol, please refer to Chretien et al. (2021).
