CellXVivo Human Treg Cell Differentiation Kit
CellXVivo Human Treg Cell Differentiation Kit Summary
For the differentiation of Human Regulatory T (Treg) Cells 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.
- Mouse Anti-Human CD3 Antibody
- Human Treg Reagent 1
- Human Treg Reagent 2
- Human Treg Reagent 3
- Human Treg Reagent 4
- Reconstitution Buffer 1
- Reconstitution Buffer 2
- Wash Buffer (20X)
Provides sufficient reagents for the differentiation of one 24-well plate.
Stability and Storage
Store the unopened kit at 2-8 °C. After opening the kit, the Mouse Anti-Human CD3 Antibody and Human Treg Reagents 1-4 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. Reconstitution Buffer 1, Reconstitution Buffer 2, and 20X Wash Buffer may be stored under sterile conditions for up to 3 months at 2-8 °C. Storage conditions are valid if used within the expiration date of the kit.
Specifications
Product Datasheets
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Scientific Data
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Differentiation of Human CD4+ T Cells into Treg Cells Confirmed by FoxP3 and CD25 Expression. Human peripheral blood naïve CD4+T cells were incubated with reagents included in the Human Treg Cell Differentiation Kit for 5 days. Cells were fixed, permeabilized, and stained using the FlowX™Human Regulatory T Cell Kit (Catalog # FMC021). Cells were imaged using flow cytometry. Quadrants were set based on isotope-stained samples.
Assay Procedure
Refer to the product datasheet for complete product details.
Briefly, mature human CD4+ T cells can be differentiated into Tregs using the following procedure:
- Coat a plate with Mouse Anti-Human CD3 Antibody
- Isolate naïve CD4+ T cells from Human PBMCs
- Culture naïve CD4+ T cells in Human Treg Differentiation Media for 5 days
- Verify differentiation into Treg cells by flow cytometry
View Graphic Procedure
Reagents Supplied in the CellXVivo™ Human T (Treg) Cell Differentiation Kit (Catalog # CDK006):
- Mouse Anti-Human CD3 Antibody
- Human Treg Reagent 1
- Human Treg Reagent 2
- Human Treg Reagent 3
- Human Treg Reagent 4
- Reconstitution Buffer 1
- Reconstitution Buffer 2
- Wash Buffer (20X)
Reagents
- Ficoll-Hypaque™
- MagCellect™ Human Naïve CD4+ T Cell Isolation Kit (R&D Systems, Catalog # MAGH115, or equivalent)
- X-VIVOTM15 Chemically Defined, Serum-free Hematopoietic Cell Medium (Lonza, or equivalent)
Equipment
- Tissue culture flasks and/or plates
- Sterile deionized water
- Microscope
- Hemocytometer
- 37 °C and 5% CO2 incubator
- Centrifuge
- Pipettes and pipette tips
- Coat wells of a 24-well plate with Mouse Anti-Human CD3 Antibody.
- Isolate PBMCs from human blood.
- Isolate human naïve CD4+ T cells from PBMCs (e.g., using magnetic cell selection).
- Perform a cell count.
- Suspend 1 - 2 x 105 naïve CD4+ T cells/mL in Human 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.
Citations for CellXVivo Human Treg Cell Differentiation Kit
R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.
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Citations: Showing 1 - 2
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Protein Kinase C Theta Modulates PCMT1 through hnRNPL to Regulate FOXP3 Stability in Regulatory T Cells
Authors: EI Ozay, S Shanthalin, JA Torres, BA Osborne, GN Tew, LM Minter
Mol. Ther., 2020-06-15;0(0):. 2020-06-15
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Cell-Penetrating Anti-Protein Kinase C Theta Antibodies Act Intracellularly to Generate Stable, Highly Suppressive Regulatory T Cells
Authors: EI Ozay, S Shanthalin, HL Sherman, JA Torres, BA Osborne, GN Tew, LM Minter
Mol. Ther., 2020-05-23;0(0):. 2020-05-23
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