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Study the Tumor Microenvironment With Flow Cytometry
Tumor microenvironments typically comprise multiple cell types including cancer, stromal, immune, endothelial, and non-pathogenic cells. Together, these cells create a complex and dynamic system through direct cell-to-cell contacts as well as by secreted proteins and low-molecular weight compounds. These cells and chemical factors are involved in cancer progression, metastasis, and evasion from host immunity. Understanding tumor microenvironments is therefore key to overcoming the resistance to therapeutic intervention that many tumors exhibit.
How it works: Immune surveillance in the tumor microenvironment
1. Mutations in tumor cells result in the formation of tumor-derived antigens (TDAs) that are released upon immunogenic cell death and are captured by antigen-presenting cells (APCs) such as dendritic cells
2. APCs prime and activate T cells by presenting the TDAs to naive T cells, initiating the immune response against cancer cells
3. Primed and activated effector T cells infiltrate the tumor tissue; T cells recognize, bind, and kill TDA-presenting malignant cells
4. Tumors can exhaust T cells with upregulation of immune checkpoint proteins or inactivate T cells with recruitment of regulatory T cells (Tregs)
Cell types typically found in the tumor microenvironment
Each immune cell has a specialized function in the recognition and elimination of tumor cells. Identifying immune cells in the lab often requires complex multicolor immunophenotyping experiments to distinguish subtle cell surface marker differences. Tissue preparation methods that involve mechanical isolation (and limit enzymatic dissociation as much as possible) typically result in better preservation of antigenic epitopes, but such preparations can cause clogging during flow cytometry runs. The Attune NxT Flow Cytometer is a non-pressure based instrument with the capability to analyze crudely prepared solid tumor samples with minimal clogging.
Typical solid tumor sample prep and analysis workflow
Isolate tumor sample
Mechanically dissociate tumor cells with a sterile razor blade in protein-containing medium such as FBS
Disperse cells in protein-containing medium such as FBS
Wash cells in PBS
Pass through fine-mesh sieve
Read on a flow cytometer with clog resistance capabilities such as the Attune NxT Flow Cytometer
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