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Updated: Jul 12, 2026
Assessing Retinal Microglial Phagocytic Function In Vivo Using a Flow Cytometry-based Assay
Published on: October 18, 2016
Protocol for evaluating microglial phagocytosis in situ in mouse hippocampal slices
Summer G Paulson1, Fritz W Lischka2, Jeremy D Rotty3
1Uniformed Services University of the Health Sciences, Department of Biochemistry, Bethesda, MD 20814, USA; The Henry M. Jackson Foundation for the Advancement of Military Medicine, Bethesda, MD 20817, USA.
Abstract:
Microglia are the primary phagocytes of the central nervous system (CNS). Here, we present a protocol for microinjecting opsonized particles into mouse hippocampal slices followed by two-photon time-lapse microscopy. We describe steps for preparing components, removing the brain, creating coronal slices, and incubating samples. We then detail procedures for analyzing microglial morphology and cell process dynamics with microinjected particles. We also provide several common potential problems encountered during the protocol alongside troubleshooting strategies to counter them. For complete details on the use and execution of this protocol, please refer to Paulson et al.1.
Insights
This study details a protocol for studying microglia, the brain's immune cells, using microinjected particles in mouse brain slices. It provides methods for analyzing microglial phagocytosis and cell dynamics.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglia are the primary immune cells and phagocytes within the central nervous system (CNS).
- Understanding microglial function is crucial for CNS health and disease research.
- Existing methods may not fully capture dynamic microglial interactions in situ.
Purpose of the Study:
- To present a detailed protocol for investigating microglial phagocytosis and cell dynamics.
- To enable the study of microglial responses to opsonized particles in a controlled ex vivo environment.
- To provide a reproducible method for analyzing microglial morphology and process behavior.
Main Methods:
- Microinjection of opsonized particles into ex vivo mouse hippocampal slices.
- Two-photon time-lapse microscopy for live imaging of microglial activity.
- Detailed procedures for sample preparation, brain slicing, and incubation.
- Quantitative analysis of microglial morphology and cell process dynamics.
Main Results:
- The protocol allows for visualization and analysis of microglial phagocytic activity.
- It enables the assessment of dynamic changes in microglial cell processes.
- Troubleshooting strategies are provided for common experimental challenges.
Conclusions:
- This protocol offers a robust method for studying microglial function in the CNS.
- It facilitates the investigation of microglial phagocytosis and cellular dynamics in a relevant ex vivo model.
- The described techniques can advance research into neuroinflammation and neurodegenerative diseases.
