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StemXVivo Serum-Free Dendritic Cell Base Media
Catalog # CCM003 | R&D Systems, Inc. a Bio-Techne Brand
Key Product Details
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Differentiation of CD14+ Monocytes into Dendritic Cells. LPS-matured monocyte-derived dendritic cells were obtained after CD14+-enriched monocytes were cultured for nine days in StemXVivo Serum-Free Dendritic Cell Base Media (Catalog # CCM003) supplemented with GM-CSF (Catalog # 215-GM) and IL-4 (Catalog # 204-IL). |
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Phenotypic Analysis of Cultured Monocyte-derived Dendritic Cells Before and After LPS-induced Maturation. Immature monocyte-derived dendritic cells (MoDCs) (open histograms; green line) were obtained after CD14+-enriched monocytes were cultured for seven days in StemXVivo Serum-Free Dendritic Cell Base Media (Catalog # CCM003) supplemented with 50 ng/mL Recombinant Human GM-CSF (Catalog # 215-GM) and 35 ng/mL Recombinant Human IL-4 (Catalog # 204-IL). Mature monocyte-derived dendritic cells (filled histograms) were cultured under the same conditions for seven days and then induced with LPS for an additional 48 hours. Day seven immature MoDCs and day nine LPS-treated MoDCs were stained with a FITC-conjugated Mouse Anti-Human B7-1/CD80 Monoclonal Antibody (Catalog # FAB140F), a FITC-conjugated Mouse Anti-Human CD83 Monoclonal Antibody (Catalog # FAB1774F), a FITC-conjugated Mouse Anti-Human B7-2/CD86 Monoclonal Antibody (Catalog # FAB141F), an Anti-MHC Class II Antibody, or an appropriate isotype control antibody (empty histogram with solid line). |
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Mature Monocyte-derived Dendritic Cells Induce Proliferation of Allogenic T Cells. CD14+monocytes were cultured for seven days in Human StemXVivo Serum-free Dendritic Cell Base Media (Catalog # CCM003) supplemented with Recombinant Human GM-CSF (Catalog # 215-GM), Recombinant Human IL-4 (Catalog # 204-IL) and Gentamycin. The cells were subsequently treated with LPS for an additional 48 hours to induce dendritic Cell Differentiation/Maturation. Graded doses of mature monocyte-derived dendritic cells were incubated with 1 x 105autologous or allogenic CD3+T cells for five days.3H-thymidine (3H-TdR) was added to the culture for the final 18 hours and T cell proliferation was measured using a scintillation counter. Results are presented as the mean cpm obtained from three experiments. |
Assay Procedure
Refer to the product datasheet for complete product details.
Briefly, CD14+ monocytes are differentiated into dendritic cells using the following procedure:
- Culture CD14+ cells in StemXVivo® Serum-Free Dendritic Cell Base Media supplemented with GM-CSF, IL-4, and Gentamicin.
- Replace the media every three days.
- Harvest immature MoDCs or use LPS to induce maturation
Reagents Provided
Reagents supplied in the StemXVivo® Serum-Free Dendritic Cell Base Media (Catalog # CCM003):
- 250 mL StemXVivo® Serum-Free Dendritic Cell Base Media
Other Supplies Required
Reagents
- Human CD14+ monocytes isolated using the MagCellect Human CD14+ Cell Isolation Kit (Catalog # MAGH105) or equivalent
- Recombinant Human GM-CSF (Catalog # 215-GM)
- Recombinant Human IL-4 (Catalog # 204-IL)
- LPS
- Gentamicin (Catalog # B20192)
Materials
- Tissue culture flasks (25 cm2 or 75 cm2) or plates (6-well or 24-well)
- 15 mL centrifuge tubes
- Serological pipette
- Pipettes and pipette tips
- Hemocytometer
Equipment
- 37 °C and 5% CO2 incubator
- Centrifuge
- Vortex mixer
- Inverted Microscope
Procedure Overview
Differentiation of CD14+ Monocytes into Monocyte-derived Dendritic Cells Using StemXVivo® Serum-Free Dendritic Cell Base Media (Catalog # CCM003)
Thaw Human StemXVivo® Serum-Free Dendritic Cell Base Media.
Mix the thawed media by vortexing and warm the media in a 37 °C incubator.
Prepare the CD14+ cells from peripheral blood mononuclear cells.
Resuspend the purified CD14+ cells in Human StemXVivo® Serum-Free Dendritic Cell Base Media.
Perform a cell count and adjust the cell density to 1 x 106 cells/mL.
Supplement the media with Gentamicin, GM-CSF, and IL-4.
Add the cell suspension to the tissue culture flask or tissue culture plate.
Incubate the cells in a 37 °C and 5% CO2 humidified incubator for 7 days.
Change the media every three days.
Observe immature monocyte-derived dendritic cells (MoDCs) from day 5 to day 7.
Induce maturation of MoDCs by adding LPS to the cell suspension on day 7 and incubating for 48 hours.
Citations for StemXVivo Serum-Free Dendritic Cell Base Media (3)
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Citations are publications that use Bio-Techne products. Selected citations for StemXVivo Serum-Free Dendritic Cell Base Media include:
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Species: Human
Sample Types: Whole Cells
Applications: BioassayJS Moon et al. (2023-01-19), Cytotoxic CD8+ T cells target citrullinated antigens in rheumatoid arthritis Nature Communications, 2023-014(1):319.
PMID: 36658110 -
Species: Human
Sample Types: Whole Cells
Legat A et al. (2010-03-01), CD14-independent responses induced by a synthetic lipid A mimetic. Eur J Immunol, 2010-040(3):797-802.
PMID: 20039305 -
Species: Human
Sample Types: Whole Cells
Duraisingham SS et al. (2009-07-22), TLR-stimulated CD34 stem cell-derived human skin-like and monocyte-derived dendritic cells fail to induce Th17 polarization of naive T cells but do stimulate Th1 and Th17 memory responses. J. Immunol., 2009-0183(4):2242-51.
PMID: 19625644
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