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Whole-mount immunofluorescence imaging and isolation of mesothelium-bound immune cell aggregates during mouse
Margarita Ferriz1, Adrián Vega-Pérez1, Alejandra Gutiérrez-González1
1Departamento de Inmunología y Oncología, Centro Nacional de Biotecnología/CSIC, Darwin 3, 28049 Madrid, Spain.
Abstract:
Resident peritoneal macrophages (resMØs) are crucial for repairing peritoneal injuries and controlling infections by forming mesothelium-bound resMØ-aggregates in the peritoneal wall and omentum. Here we present a protocol to analyze these structures in mouse models of peritoneal inflammation. We describe the dissection, fixation, immunofluorescent staining, and mounting of whole peritoneal wall and omentum samples and subsequent confocal microscopy imaging of resMØ-aggregates. We also detail the steps to isolate resMØ-aggregates for additional studies, including flow cytometry and electron-microscopy-based analysis. For complete details on the use and execution of this protocol, please refer to Vega-Pérez et al. (2021).1.
Insights
This study details a protocol for analyzing resident peritoneal macrophages (resMØs) and their aggregates in mouse models. The method aids in understanding peritoneal injury repair and infection control.
Area of Science:
- Immunology
- Cell Biology
- Microscopy
Background:
- Resident peritoneal macrophages (resMØs) are vital for peritoneal tissue repair and infection defense.
- These macrophages form aggregates crucial for their function in the peritoneum and omentum.
Purpose of the Study:
- To present a detailed protocol for analyzing mesothelium-bound resMØ-aggregates in mouse models.
- To facilitate the study of peritoneal inflammation and macrophage roles.
Main Methods:
- Dissection, fixation, and immunofluorescent staining of whole peritoneal wall and omentum samples.
- Confocal microscopy for imaging resMØ-aggregates.
- Isolation of resMØ-aggregates for flow cytometry and electron microscopy.
Main Results:
- The protocol enables comprehensive analysis of resMØ-aggregate structures.
- It allows for detailed examination of macrophage behavior in peritoneal inflammation models.
Conclusions:
- This protocol provides a robust framework for investigating resident peritoneal macrophages and their aggregates.
- It supports further research into peritoneal immunity and repair mechanisms.
