Primary normal human pre-adipocytes (ATCC PCS-210–010, American Type Culture Collection, Manassas, VA, USA) were differentiated into mature adipocytes in wells of 24-well-plates containing adipocyte differentiation medium (Cell Applications, Inc. San Diego, CA) in a 5% CO2 atmosphere at 370 C29 (link). These cells can be expanded in an undifferentiated state for future differentiation to mature adipocytes and show higher efficiency of adipogenesis compared to mesenchymal stem cells. In this study, the cells were cultured in adipocyte differentiation medium with increasing doses of glucose (8.4mM to 19.3mM, Sigma-Aldrich, St. Louis, MO), or ADM (1×10− 10M to 1×10− 8M, Sigma-Aldrich) for 24 hours. Total RNA was isolated from the cells using TRIzol (Life Technologies, Grand Island, NY) and RT was performed for further Quantitative Real-time-PCR analysis.
Adipocyte differentiation medium
Manufactured by Merck Group
Sourced in United States
Adipocyte differentiation medium is a specialized cell culture medium designed to support the differentiation of preadipocytes into mature adipocytes (fat cells) in vitro. The medium provides the necessary nutrients, growth factors, and hormones to facilitate the adipogenic differentiation process.
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Lab products found in correlation
Osteogenic differentiation medium, by Merck Group (2 mentions)
Alizarin red, by Merck Group (2 mentions)
Adipocyte differentiation medium, by Cell Applications (1 mentions)
Trizol, by Thermo Fisher Scientific (1 mentions)
Atcc pcs210010, by American Type Culture Collection (1 mentions)
Chondrogenic differentiation medium, by Merck Group (1 mentions)
Alcian blue, by Merck Group (1 mentions)
Oil red o, by Merck Group (1 mentions)
Oil red, by Merck Group (1 mentions)
3 protocols using adipocyte differentiation medium
1
Adipocyte Differentiation and Glucose/ADM Response
2
Evaluating EMSC Differentiation Potential
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The capacity of EMSCs to differentiate in vitro was evaluated. EMSCs were seeded at 1 × 105/cm2 and cultured for 3 weeks in osteogenic differentiation medium (0.01 μM 1α-25-dihydroxyvitamin-D3, 10 mM β-glycerophosphate, and 50 μM ascorbate-2) (Sigma-Aldrich, St. Louis, MO, USA), adipocyte differentiation medium (500 μM isobutyl-methylxanthine, 1 μM dexamethasone, 10 μM insulin, and 200 μM indomethacin) (Sigma-Aldrich, St. Louis, MO, USA), or chondrogenic differentiation medium (6.25 g/ml insulin, 50 μM ascorbate-2) (Sigma-Aldrich, St. Louis, MO, USA). Cells cultured in normal medium were used as the control. The medium was changed every 3 days. After 3 weeks, the cells in each group were stained with alizarin red, oil red O, and alcian blue (Sigma-Aldrich, St. Louis, MO, USA) to assess osteogenesis, adipogenesis, and chondrogenesis, respectively [25 (link)].
3
Differentiation of hWJ-MSCs into Osteocytes and Adipocytes
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The differentiation potential of cells was examined on third passage of the hWJ-MSCs. For induction of osteogenic differentiation, the hWJ-MSCs were plated in six-well plates at a density of 10000 cells/cm2 in triplicates. After 48 hr, osteogenic differentiation medium (Sigma-Aldrich) was added to the cells, and the supplemented DMEM culture medium was added to the control cells. Medium refreshment was performed every 3–4 days for 21 days. Finally, the cells were washed with PBS, fixed with 4% paraformaldehyde and stained with alizarin red (Sigma-Aldrich) to detect the presence of calcium deposition in osteocytes.
For induction of adipogenic differentiation, the MSCs were plated in six-well plates at a density of 10000 cells/cm2 in triplicates. After 48 hr, adipocyte differentiation medium (Sigma-Aldrich) was added to the cells. In case of controls, the supplemented DMEM culture medium was added to the cells (used as control group). The culture media were replaced every 3 days for 21 days. Finally, the cells were washed with PBS, fixed with 4% paraformaldehyde and stained with Oil Red (Sigma-Aldrich) to detect the presence of neutral lipid vacuoles in adipocytes.
For induction of adipogenic differentiation, the MSCs were plated in six-well plates at a density of 10000 cells/cm2 in triplicates. After 48 hr, adipocyte differentiation medium (Sigma-Aldrich) was added to the cells. In case of controls, the supplemented DMEM culture medium was added to the cells (used as control group). The culture media were replaced every 3 days for 21 days. Finally, the cells were washed with PBS, fixed with 4% paraformaldehyde and stained with Oil Red (Sigma-Aldrich) to detect the presence of neutral lipid vacuoles in adipocytes.
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