A 38-colour high dimensional immunophenotyping panel for human peripheral blood mononuclear cells

Area of Science:

• Immunology
• Cell Biology
• Biotechnology

Background:

• High-dimensional immunophenotyping is crucial for comprehensive immune cell analysis.
• Peripheral blood mononuclear cells (PBBMCS) are key targets for immune profiling.
• Existing methods may have limitations in analyzing rare immune cell subsets.

Purpose of the Study:

• To design and optimize a 38-colour high-dimensional phenotyping panel for Cytek Aurora.
• To enable detailed characterization of innate, T, and B cell subsets and their activation status.
• To provide a valuable tool for ex vivo studies, particularly with limited sample volumes.

Main Methods:

• Development of a 38-colour flow cytometry panel.
• Optimization of fluorochrome combinations and titres to minimize spectral spread and autofluorescence.
• Testing the panel on peripheral blood mononuclear cells (PBMCs) from 15 healthy adults.
• Utilizing a 5-laser Cytek Aurora cytometer for high-resolution analysis.

Main Results:

• Successful design and validation of a comprehensive 38-colour immunophenotyping panel.
• Demonstrated ability to measure diverse immune cell populations including monocytes, dendritic cells, NK cells, T cell subsets (e.g., MAIT, Treg), and B cell subsets (e.g., plasma cells, transitional B cells).
• Optimized panel effectively minimizes spread and autofluorescence, crucial for rare cell population detection.

Conclusions:

• The developed high-dimensional panel is a powerful resource for extensive immune cell phenotyping.
• This tool facilitates direct ex vivo studies for evaluating immune status in human health and disease.
• The panel is particularly advantageous when working with limited sample quantities, enhancing research efficiency.