CellXVivo Human Th1 Cell Differentiation Kit
R&D Systems, part of Bio-Techne | Catalog # CDK001
Key Product Details
Assay Procedure
Refer to the product datasheet for complete product details.
Briefly, human CD4+ T cells can be differentiated into Th1 cells using the following procedure:
- Coat a plate with Mouse Anti-Human CD3 Antibody
- Isolate human naïve CD4+ T cells from a PBMC preparation
- Culture naïve CD4+ T cells in Human Th1 Differentiation Media for 5 days
- Stimulate cells with monensin
- Analyze culture media and cells for a Th1-specific cytokine production profile
Reagents Supplied in the CellXVivo™ Human Th1 Cell Differentiation Kit (Catalog # CDK001):
- Mouse Anti-Human CD3 Antibody
- Human Th1 Reagents
- Reconstitution Buffers
- Wash Buffer (20X)
Reagents
- MagCellect™ Human Naïve CD4+ T Cell Isolation Kit (Catalog # MAGH115 or equivalent)
- Monensin
- PMA
- Ficoll-Hypaque™
- RPMI 1640
- L-Glutamine
- Ionomycin
- Penicillin
- Streptomycin
- Fetal Bovine Serum (FBS)
- beta-Mercaptoethanol
Equipment
- Tissue culture flasks and/or plates
- Hemocytometer
- 37 °C and 5% CO2 humidified incubator
- Centrifuge
- Pipettes and pipette tips
Coat wells of a 24-well plate with Mouse Anti-Human CD3 Antibody.
Isolate PBMCs from human blood (e.g., using Ficoll-Hypaque™ density centrifugation).
Isolate human naïve CD4+ T cells from PBMCs (e.g., using R&D Systems MagCellect™ Human Naïve CD4+ T Cell Isolation Kit).
Perform a cell count.
Suspend 1-2 x 105 naïve CD4+ T cells/mL in media supplemented with Human Th1 Reagent 1 and Human Th1 Reagent 2.
Culture the cells on plates pre-coated with Mouse Anti-Human CD3 Antibody for 5 days.
Refresh the Diferentiation Media every 3-4 days.
Stimulate the cells with mitogens.
Verify Th1 cell differentiaiton by analying cytokine expression via flow cytometry.
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