CD4+ T cells can differentiate into T helper (Th)1, Th2, Th17, and Regulatory T (Treg) cells by exposure to various cytokines and cellular interactions that induce expression of specific sets of transcription factors. Differentiation into Th1 cells is promoted through IL-12 and IFN-gamma. These cells are characterized by their secretion of IFN-gamma, IL-10, and TNF-alpha. Differentiation into Th2 cells is promoted by IL-4 in combination with either IL-2, IL-7, or TSLP. through IL-12 and IFN-gamma. These cells are characterized by their secretion of IFN-gamma, IL-10, and TNF-alpha. Th2 cells secrete IL-4, IL-5, IL-9, IL-13, and IL17E/IL-25. Differentiation into the Th17 lineage is promoted by cytokines such as TGF-beta and IL-6, while their survival and expansion are dependent on IL-21 and IL-23. Th17 cells secrete TNF-alpha, IL-6, IL-9, IL-17A, IL-17F, IL-21, IL-22, and (human) IL-26/AK155. The ability to differentiate CD4+ T cells ex vivo into Th1, Th2, Th17, or Treg cells is valuable to researchers who require specific T helper cell subsets for downstream applications, including studies for cell therapy and cancer immunotherapy.
CellXVivo Mouse Treg Cell Differentiation Kit
R&D Systems | Catalog # CDK007
Key Product Details
Species
Product Summary for CellXVivo Mouse Treg Cell Differentiation Kit
Kit Summary
For the differentiation of regulatory T Cells (Tregs) from a preparation of naïve CD4+ T cells.
Key Benefits
- Contains high quality bioactive proteins
- Yields a highly enriched population of FoxP3+CD25+ Treg cells
- Provides optimized reagents for inducing Tregs from CD4+ T cells
- Involves validated and straightforward procedures
- Does not require specialized instrumentation
Why Differentiate Treg Cells In Vitro?
Forkhead Box P3 (FoxP3)+ regulatory T (Treg) cells are a suppressive subset of CD4+ T cells that function to antagonize immune responses. Treg cells have the capacity to prevent potentially damaging autoimmune and protective immune responses, so the number of Treg cells is a crucial determinant of the regulatory burden on the immune system. In vitro differentiation of Treg cells from the larger naïve CD4+ T cell population provides increased numbers of Treg cells to facilitate downstream research.
This kit contains the following optimized proteins and reagents to drive efficient differentiation of naïve CD4+ T cells into FoxP3+CD25+ Treg cells.
- Hamster Anti-Mouse CD3 Antibody
- Mouse Treg Reagent 1
- Mouse Treg Reagent 2
- Mouse Treg Reagent 3
- Reconstitution Buffer 1
- Reconstitution Buffer 2
- Wash Buffer (20X)
Stability and Storage
Store the unopened kit at -20 °C. After opening the kit, the Human Anti-Mouse CD3 Antibody, Mouse Treg Reagent 1, and Mouse Treg Reagent 2 may be stored at 2-8 °C under sterile conditions for up to 30 days or at -20 °C to -70 °C in a manual defrost freezer for up to 3 months. Mouse Treg Reagent 3 may be stored at -20 °C to -70 °C in a manual defrost freezer for up to 3 months. Reconstitution Buffer 1, Reconstitution Buffer 2, and 20X Wash Buffer may be stored under sterile conditions for up to 3 months at 2 to 8 °C. Storage conditions are valid if used within the expiration date of the kit.
Scientific Data Images for CellXVivo Mouse Treg Cell Differentiation Kit
Differentiation of CD4+ T Cells into Treg Cells Confirmed by FoxP3 and CD25 Expression.
Naïve CD4+T cells were incubated without (A, B) or with (C, D) reagents included in the CellXVivo™ Mouse Treg Cell Differentiation Kit for 5 days. Cells were fixed, permeabilized, and stained using the FlowX Mouse Regulatory T Cell Kit (Catalog # FMC022). Cells were imaged using flow cytometry. Quadrants were set based on isotope-stained samples.
Formulation, Preparation, and Storage
Shipping
Storage
Background: T Cell Differentiation Kits
Additional T Cell Differentiation Kits Products
Product Documents for CellXVivo Mouse Treg Cell Differentiation Kit
Certificate of Analysis
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Note: Certificate of Analysis not available for kit components.
Product Specific Notices for CellXVivo Mouse Treg Cell Differentiation Kit
For research use only
Related Research Areas
Citations for CellXVivo Mouse Treg Cell Differentiation Kit
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Protocols
View specific protocols for CellXVivo Mouse Treg Cell Differentiation Kit (CDK007):
Refer to the product datasheet for complete product details.
Briefly, naïve mouse CD4+ T cells can be differentiated into Tregs using the following procedure:
- Coat a plate with Hamster Anti-Mouse CD3 Antibody
- Isolate naïve CD4+ T cells from mouse splenocytes
- Culture naïve CD4+ T cells in Mouse Treg Differentiation Media for 5 days
- Verify differentiation into Treg cells by flow cytometry
Reagents Supplied in the CellXVivo™ Mouse Treg Cell Differentiation Kit (Catalog # CDK007):
- Hamster Anti-Mouse CD3 Antibody
- Mouse Treg Reagent 1
- Mouse Treg Reagent 2
- Mouse Treg Reagent 3
- Reconstitution Buffer 1
- Reconstitution Buffer 2
- Wash Buffer (20X)
Reagents
- MagCellect™ Mouse Naïve CD4+ T Cell Isolation Kit (R&D Systems, Catalog # MAGH205, or equivalent).
- RPMI 1640
- Fetal Bovine Serum (FBS)
- β-Mercaptoethanol (2-ME)
- L-Glutamine–Penicillin–Streptomycin solution
Equipment
- Tissue culture flasks and/or plates
- Sterile deionized water
- Microscope
- Hemocytometer
- 37 °C and 5% CO2 incubator
- Centrifuge
Coat wells of a 24-well plate with Hamster Anti-Mouse CD3 Antibody.
Prepare a single cell suspension of mouse splenocytes.
Isolate human naïve CD4+ T cells from PBMCs (e.g., using magnetic cell selection).
Perform a cell count.
Suspend 0.5 - 1 x 106 naïve CD4+ T cells/mL in Mouse Treg Differentiation Media.
Culture the cells on plates pre-coated with CD3 antibody for 5 days.
Verify Treg cell differentiation by analyzing marker expression using flow cytometry.
Mature Treg cells are now ready for further downstream applications.
