MagCellect Mouse CD4+ CD25+ Regulatory T Cell Isolation Kit

Kit Summary

For the isolation of mouse regulatory T cells from a preparation of splenocytes.

Why Isolate Regulatory T cells?

Regulatory T cells (Treg), are a minor subset of CD4⁺ T cells that can suppress the development and progression of immune reactions. They also play an important role in the balance between tolerance and autoimmunity. Treg cells are characterized by the expression of CD4 and CD25 on the cell surface and the intracellular transcription factor FoxP3. The isolation of regulatory T cells facilitates research into their critical role in immune system regulation.

Subsets and Phenotypes

Treg cells generated in the thymus are known as natural Treg (nTreg) or thymic Treg (tTreg). Alternatively, Treg cells can differentiate in the periphery from naïve CD4⁺ T cells. These cells are known as induced Treg (iTreg) or peripherally-derived Treg (pTreg). pTreg cells can further differentiate into subsets with variable expression of CCR4, CCR5, CCR6, CCR10, and CXCR3. Differential chemokine receptor expression enables these cells to migrate to specific tissues and inflammatory sites. Some populations of Treg cells do not express FoxP3 but retain an immune suppressive function, while other populations express FoxP3 but do not exhibit suppressive activity. Additional Treg molecules that may aid in the discrimination between Treg cell populations include the cell surface proteins PD-1, Neuropilin, CD73, and LRRC32/GARP and the transcription factor Helios. There are significant differences in the generation and function of Treg cells in mice and in humans.

Induction and Mechanism of Action

The expression of FoxP3 is integral to the in vivo development of Treg cells. The presence of TGF-beta, IL-2, and IL-10 in tissue culture supports Treg cell differentiation and expansion in vitro. Dendritic cells (DC) support Treg cell development and expansion, and Treg cells in turn regulate the differentiation of DC precursors. Treg cells limit T effector cell differentiation, proliferation, and activation, as well as humoral B cell responses. Mechanistically, FoxP3 directly blocks transcription of the pro-inflammatory cytokines IL-17 and IFN-gamma. Cell surface CTLA-4 and LAG3 and secreted IL-10, IL-35, and TGF-beta are additionally implicated in Treg cell function.

Treg in Disease

Treg cell-mediated immune suppression is important for restraining immune reactions, but this activity can be damaging in the context of disease. For example, Treg cell generation in response to oral antigens or intestinal microbes promotes the development of immune tolerance. In contrast, gut Treg cells can contribute to chronic inflammatory states such as colitis. In cancer, elevated numbers of Treg cells are found within tumors. This imbalance is a result of the actions of local dendritic cells that recruit tTreg and also expand and induce pTreg. Intratumoral Treg cells then contribute to the suppression of host anti-tumor immunity. The immune dysregulatory syndrome IPEX results from mutations in FoxP3, and IPEX patients suffer from a variety of autoimmune disorders.

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Reagents Provided

The MagCellect Mouse CD4⁺CD25⁺ Regulatory T Cell Isolation Kit (Catalog # MAGM208) contains the following reagents that allow for isolation of highly pure mouse Treg cells.

• MagCellect Mouse CD4⁺ T Cell Biotinylated Antibody Cocktail; 1 mL in PBS containing BSA and preservative
• MagCellect Streptavidin Ferrofluid; 2 mL in a solution containing BSA and preservative
• Anti-Mouse CD25 Biotinylated Antibody; 0.5 mL in a solution containing BSA and preservative
• MagCellect Plus Buffer (10X); 25 mL of a 10X concentrated buffer

*This kit contains sufficient reagents to process 1 x 10⁹ total cells.

Stability and Storage

Store all reagents at 2 °C to 8 °C. DO NOT FREEZE.