CellXVivo Mouse Th2 Cell Differentiation Kit
CellXVivo Mouse Th2 Cell Differentiation Kit Summary
For the differentiation of Th2 cells from a preparation of CD4+ T cells isolated from mouse splenocytes.
Key Benefits
- Provides optimized reagents needed to induce Th2 differentiation
- Yields ~80 x 106 CD4+ T cells, of which > 40% are Th2 polarized
- Contains high quality bioactive proteins
- Includes straightforward procedures
- Does not require specialized instrumentation
Why Expand and Differentiate Th2 Cells Ex Vivo?
T helper type 2 (Th2) cells are a lineage of CD4+ effector T cells that provide host protection against intestinal parasites and extracellular bacteria. In addition, they provide support for B cell-dependent humoral responses. Pathological Th2 cell activity is a hallmark of allergic inflammation and asthma. Differentiation of CD4+ effector cells into the Th2 lineage is promoted by cytokines such as IL-4 in combination with either IL-2, IL-7, or TSLP. Th2 cells secrete IL-4, IL-5, IL-9, IL-13, and IL-17E/IL-25. In vitro differentiation of mouse Th2 cells from the larger naïve CD4+ T cell population provides increased numbers of Th2 cells to facilitate downstream research. The CellXVivo™ Mouse Th2 Cell Differentiation Kit contains all necessary components to differentiate mouse naïve CD4+ T cells into Th2 polarized cells.
This kit contains the following reagents for the ex vivo differentiation of mouse Th2 cells.
- Hamster Anti-Mouse CD3, Th2
- Rat Anti-Mouse CD28, Th2
- Mouse Th2 Reagent 1
- Mouse Th2 Reagent 2
- Mouse Th2 Reagent 3
- Reconstitution Buffer 1
- Reconstitution Buffer 2
- 20X Wash Buffer
The quantity of the components in the kit is sufficient to differentiate approximately 4 x 106 naïve CD4+ T cells, and generate 80 x 106 T cells, of which >40% are IL-4+ Th2 polarized cells.
Note: Results may vary due to strain, age, and/or the health of the mice used for isolation.
Stability and Storage
Store the unopened kit at ≤-20 °C. Do not use past the kit expiration date.
Limitations
- FOR LABORATORY RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.
- The safety and efficacy of this product in diagnostic or other clinical uses has not been established.
- This reagent should not be used beyond the expiration date indicated on the label.
- Results may vary due to variations among cells derived from different donors.
Specifications
Product Datasheets
Scientific Data
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Th2-differentiated Mouse CD4+ Cells Secrete High Levels of IL-4. Mouse naïve CD4+T cells were differentiated for 6 days using the reagents included in this kit. On day 6 of differentiation cells were harvested and stimulated with Anti-Mouse CD3 and Anti-Mouse CD28 overnight. Cell culture supernatant was collected and cytokine secretion determined using the Mouse IL-4 Quantikine®ELISA Kit (Catalog # M4000B), the Mouse IFN-gamma Quantikine®ELISA Kit (Catalog # MIF00), and the Mouse IL-17 Quantikine®ELISA Kit (Catalog # M1700).
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Intracellular Cytokine Staining of Differentiated Mouse Th2 Cells. Flow cytometry data of mouse naïve CD4+T cells (A, C) and differentiated Th2 cells (B, D) generated using reagents included in this kit. After 6 days of differentiation, naïve CD4+cells or differentiated Th2 cells were stimulated with Cell Activation Cocktail (Tocris®, Catalog # 5476) and stained with conjugated Anti-Mouse IL-4 (Clone 11B11), Anti-Mouse IFN-gamma (Catalog # IC485P), and Anti-Mouse IL-17A (Novus Biologicals®, Catalog # NBP1-72027) Monoclonal Antibodies. Quadrants were set based on isotype-stained controls.
Assay Procedure
Refer to the product datasheet for complete product details.
Briefly, CD4+ T cells isolated from mouse splenocytes are differentiated ex vivo into the Th2 phenotype with reagents provided in this kit and using the following procedure:
- Isolate CD4+ T cells from mouse splenocytes
- Culture CD4+ T cells in reagents provided in kit
- Verify Th2 cell expansion after 6 days in culture
Reagents Supplied in the CellXVivo™ Mouse Th2 Cell Differentiation Kit (Catalog # CDK019):
- Hamster Anti-Mouse CD3, Th2
- Rat Anti-Mouse CD28, Th2
- Mouse Th2 Reagent 1
- Mouse Th2 Reagent 2
- Mouse Th2 Reagent 3
- Reconstitution Buffer 1
- Reconstitution Buffer 2
- 20X Wash Buffer
Reagents
- Laboratory mice
- X-VIVO™ 15 Chemically Defined, Serum-free Hematopoietic Cell Medium (Lonza, or equivalent)
- MagCellect™ Mouse Naïve CD4+ T Cell Isolation Kit (R&D Systems, Catalog # MAGM205, or equivalent)
- Penicillin/Streptomycin (optional)
- Cell Activation Cocktail 500X (Tocris®, Catalog # 5476)
Equipment
- Tissue culture plates and/or flasks
- Sterile deionized water
- Pipettes and pipette tips
- Inverted microscope
- Hemocytometer
- 37 °C, 5% CO2 incubator
- Centrifuge
Protocol for mouse Th2 cell differentiation using the CellXVivo™ Mouse Th2 Cell Differentiation Kit.
Note: Results may vary due to strain, age, and/or the health of the mice used for isolation.
Coat the desired tissue culture plate with Hamster Anti-Mouse CD3 antibody.
Isolate mouse splenocytes.
Isolate mouse naïve CD4+ T cells from splenocytes (e.g., using magnetic cell selection).
Perform a cell count.
Suspend 1 x 106 naïve CD4+ T cells/mL in Mouse Th2 Differentiation Media. Culture the cells on plates pre-coated with Hamster Anti-Mouse CD3 antibody.
Harvest cells on day 3.
Dilute cells 1:20 with fresh Mouse Th2 Differentiation Media.
Culture cells in a new flask for an additional 3 days.
Verify Th2 cell differentiation by analyzing cytokine expression via flow cytometry or ELISA (optional).
Citations for CellXVivo Mouse Th2 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 - 5
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JAK3 inhibitor suppresses multipotent ILC2s and attenuates steroid-resistant asthma
Authors: Kim, J;Ham, J;Kang, HR;Bae, YS;Kim, T;Kim, HY;
Science advances 2023-12-22
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Inhibition of pyruvate dehydrogenase kinase 4 in CD4+ T cells ameliorates intestinal inflammation
Authors: H Lee, JH Jeon, YJ Lee, MJ Kim, WH Kwon, D Chanda, T Thoudam, HS Pagire, SH Pagire, JH Ahn, RA Harris, ES Kim, IK Lee
Cellular and Molecular Gastroenterology and Hepatology, 2022-10-10;0(0):. 2022-10-10
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Vaccination by Two DerG LEAPS Conjugates Incorporating Distinct Proteoglycan (PG, Aggrecan) Epitopes Provides Therapy by Different Immune Mechanisms in a Mouse Model of Rheumatoid Arthritis
Authors: DH Zimmerman, K Mikecz, A Markovics, RE Carambula, JC Ciemielews, DM Toth, TT Glant, KS Rosenthal
Vaccines, 2021-05-02;9(5):. 2021-05-02
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SOX-5 activates a novel ROR&gammat enhancer to facilitate experimental autoimmune encephalomyelitis by promoting Th17 cell differentiation
Authors: Y Tian, C Han, Z Wei, H Dong, X Shen, Y Cui, X Fu, Z Tian, S Wang, J Zhou, D Yang, Y Sun, J Yuan, B Ni, Y Wu
Nature Communications, 2021-01-20;12(1):481. 2021-01-20
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Ginsenoside Rh2 Ameliorates Atopic Dermatitis in NC/Nga Mice by Suppressing NF-kappaB-Mediated Thymic Stromal Lymphopoietin Expression and T Helper Type 2 Differentiation
Authors: E Ko, S Park, JH Lee, CH Cui, J Hou, MH Kim, SC Kim
Int J Mol Sci, 2019-12-04;20(24):. 2019-12-04
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