FACS to Identify Immune Subsets in Mouse Brain and Spleen
Mary K Malone1, Thomas A Ujas1, Katherine M Cotter1
1Department of Neurology, Department of Neuroscience, The University of Kentucky, Lexington, KY, USA.
Insights
This study details protocols for preparing single-cell suspensions from mouse spleen and brain tissues. These methods facilitate immune cell analysis using fluorescently activated cell sorting (FACS) for disease research.
Area of Science:
- Immunology
- Neuroscience
- Cell Biology
Background:
- Flow cytometry is crucial for multi-parametric cell quantification, particularly in immunophenotyping immune cell subsets.
- Accurate analysis requires high-quality single-cell suspensions from various tissues.
- Understanding immune responses in diseases like neuroinflammation is vital.
Purpose of the Study:
- To describe standardized protocols for preparing single-cell suspensions from mouse spleen and brain.
- To outline the steps for fluorescently activated cell sorting (FACS) analysis.
- To enable the isolation of lymphocytes for studying immune responses in disease models.
Main Methods:
- Development of protocols for dissociating mouse spleen and brain tissues into single-cell suspensions.
- Optimization of sample preparation for fluorescently activated cell sorting (FACS).
- Application of the protocol for isolating lymphocytes.
Main Results:
- Successful generation of single-cell suspensions from mouse spleen and brain tissues.
- Demonstrated utility of the protocol for lymphocyte isolation.
- Facilitation of immune cell subset analysis for disease research.
Conclusions:
- The described protocols provide a reliable method for preparing single-cell suspensions for flow cytometry and FACS.
- These methods are applicable to studying immune responses in conditions such as neuroinflammation post-ischemic stroke.
- This work supports research into immune cell roles in various pathologies.
Abstract:
Flow cytometry enables the multi-parametric quantification of cell types, especially in immunophenotyping of unique immune cell subsets that can either contribute to or ameliorate pathology. For tissues to be used in such analyses, single-cell suspensions must be created. Here we describe protocols for preparing single-cell suspensions of mouse spleen and brain tissue, as well as the steps for fluorescently activated cell staining/sorting (FACS). Specifically, this protocol enables the isolation of lymphocytes for the study of immune responses during various diseases, such as long-term neuroinflammation following ischemic stroke.
