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Updated: Mar 27, 2026
Isolation of Uterine Innate Lymphoid Cells for Analysis by Flow Cytometry
Published on: October 14, 2021
Multi-parameter flow cytometric analysis of uterine immune cell fluctuations over the murine estrous cycle
Kerrilyn R Diener1, Sarah A Robertson2, John D Hayball1
1Experimental Therapeutics Laboratory, Hanson Institute, Royal Adelaide Hospital, and Sansom Institute, School of Pharmacy and Medical Science, University of South Australia, Adelaide, S.A., 5000, Australia; Robinson Research Institute, Discipline of Obstetrics and Gynaecology, School of Medicine, University of Adelaide, Adelaide, S.A., 5005, Australia.
Insights
Immune cell populations in mouse uterus change dynamically throughout the estrous cycle. Specific markers like anti-SiglecF help differentiate immune cells, revealing key changes during estrus and metestrus.
Area of Science:
- Reproductive immunology
- Flow cytometry
- Cellular immunology
Background:
- Flow cytometry is crucial for analyzing immune cells in reproductive tissues.
- Advances in technology allow for multi-parameter analysis of immune cell populations.
- Understanding uterine immune cell dynamics is vital for reproductive health.
Purpose of the Study:
- To characterize proportional changes in immune cell populations within mouse uterine tissue across the four stages of the estrous cycle.
- To identify reliable methods for distinguishing specific immune cell subsets, particularly monocyte/macrophages and eosinophils.
Main Methods:
- A flow cytometry protocol optimized for mouse uterine tissue was employed.
- Analysis included dendritic cells, monocyte/macrophages, T cells, B cells, NK cells, NK T cells, neutrophils, and eosinophils.
- The utility of anti-SiglecF and FSC/SSC plots for cell differentiation was assessed.
Main Results:
- Significant proportional changes in uterine immune cells were observed across the estrous cycle.
- Peak abundance for many immune cell types occurred during the estrus and metestrus phases.
- Anti-SiglecF and FSC/SSC plots proved effective in distinguishing monocyte/macrophage and eosinophil populations from F4/80+CD11b+ cells.
Conclusions:
- The mouse uterus harbors a dynamic immune cell population that fluctuates with the estrous cycle.
- Immune cell dynamics are likely important for responding to insemination-related antigens and pathogens.
- Novel differentiation strategies enhance the accuracy of uterine immune cell profiling.
Abstract:
Investigating immune cell populations within various reproductive tissues commonly utilises flow cytometric methods. With advances in fluorophore technology and equipment capabilities, multiple cell types from a single tissue sample can be identified by using different combinations of cell surface markers to distinguish specific cell populations. Here a protocol optimized for mouse uterine tissue was used to show the proportional changes in dendritic cells, monocyte/macrophages, T and B cells, NK and NK T cells, and the granulocytes, neutrophils and eosinophils at each of the four stages of the estrous cycle. Importantly, we demonstrate that use of anti-SiglecF or assessment of FSC/SSC plots could be used to differentiate monocyte/macrophage and eosinophil populations that otherwise cannot be distinguished by use of the common combination of antibodies against F4/80 and CD11b. Our results clearly indicate that within the uterus a dynamic population of immune cells resides, with many cell types reaching peak abundance at estrus and metestrus phases of the cycle, consistent with their importance in the response to paternal antigens and/or pathogens encountered after insemination.
