High-Dimensional Immunophenotyping with Fluorescence-Based Cytometry: A Practical Guidebook

Area of Science:

• Immunology
• Biotechnology
• Cell Biology

Background:

• High-dimensional single-cell immunophenotyping utilizes advanced technologies like mass cytometry, single-cell RNA sequencing, and flow cytometry.
• Fluorescence-based flow cytometry is the most prevalent method, with modern instruments measuring up to 30 parameters.
• These techniques provide profound insights into the intricate nature of the immune system.

Purpose of the Study:

• To provide a practical guidebook for designing and executing complex fluorescence-based immunophenotyping panels.
• To address common misconceptions and potential pitfalls in panel design and execution.
• To discuss challenges related to quality control and data analysis in high-dimensional flow cytometry.

Main Methods:

• The study focuses on fluorescence-based immunophenotyping panels.
• It details practical aspects of panel design and execution.
• It covers quality control and data analysis strategies for complex datasets.

Main Results:

• The guidebook addresses common misconceptions and caveats in fluorescence-based immunophenotyping.
• It highlights challenges in quality control and data analysis for high-dimensional datasets.
• Practical strategies for successful panel design and execution are provided.

Conclusions:

• Effective design and execution of complex immunophenotyping panels are crucial for accurate immune system analysis.
• Addressing common challenges in quality control and data analysis enhances the reliability of flow cytometry data.
• This guidebook serves as a valuable resource for researchers utilizing high-dimensional flow cytometry.