To confirm the adipogenic potential of young and elderly KNT cells, these cells were incubated in αMEM with 5% PLTMax until cells were confluent. Thereafter, KNT cells were cultured with adipogenic induction medium (Lonza, Basel, Switzerland). After 3 days, maintenance medium was added to cells, and three cycles of induction and maintenance media were completed. Cells were fixed with 10% formalin (Sigma-Aldrich) and stained with 0.5% Oil Red O (Sigma-Aldrich) in methanol (Sigma-Aldrich). To confirm the osteogenic potential of KNT cells, they were incubated in αMEM with 5% PLTMax until a confluent layer was achieved. Thereafter, osteogenic differentiation medium (Lonza) was added. Medium was changed every 3–4 days. After 17 days, cells were fixed in 10% formalin and stained with 10% alizarin red (Sigma-Aldrich).
Osteogenic differentiation medium
Manufactured by Lonza
Sourced in Switzerland, United States
Osteogenic differentiation medium is a specialized cell culture medium designed to support the differentiation of stem cells or progenitor cells into osteoblasts, the cells responsible for bone formation. The core function of this medium is to provide the necessary growth factors, vitamins, and other nutrients to facilitate the commitment and maturation of cells towards the osteogenic lineage.
Automatically generated - may contain errors
Lab products found in correlation
Adipogenic differentiation medium, by Lonza (2 mentions)
Methanol, by Merck Group (1 mentions)
Formalin, by Merck Group (1 mentions)
Alizarin red, by Merck Group (1 mentions)
Adipogenic induction medium, by Lonza (1 mentions)
Oil red o, by Merck Group (1 mentions)
Tgf β3, by ProSpec (1 mentions)
F actin probe, by Merck Group (1 mentions)
Sybr green master mix reagent, by Thermo Fisher Scientific (1 mentions)
Ascorbic acid 2 phosphate, by Merck Group (1 mentions)
Cdna synthesis kit, by Thermo Fisher Scientific (1 mentions)
Alizarin red s ar, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Glutamine, by Merck Group (1 mentions)
Live dead double staining kit, by Merck Group (1 mentions)
Chondrogenic differentiation medium, by Lonza (1 mentions)
Mscgm cd, by Lonza (1 mentions)
Human mesenchymal stem cell growth medium, by Lonza (1 mentions)
Lookout mycoplasma detection kit, by Merck Group (1 mentions)
Hmscs, by Lonza (1 mentions)
12 protocols using osteogenic differentiation medium
1
Adipogenic and Osteogenic Potential of KNT Cells
2
Multifaceted Stem Cell Differentiation
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For osteogenic and adipogenic differentiation, 3x103 cells were seeded in wells of a MW-6 plate and cultured in growth medium for 24 hours. Osteogenesis was induced by culturing the cells in Osteogenic Differentiation Medium (Lonza, Biowhittaker). Adipogenesis was induced by culturing cells in hMSC Commercial Adipogenic Differentiation Medium (Lonza, Biowhittaker), following the manufacturer's instructions. Cultures were alternated between induction and maintenance medium every 3 days. Both differentiation processes were maintained for 9 days.
Chondrogenesis was assessed using the micropellet formation (2.5x104) technique [34 (link)], with some modifications. 3a6 and 3a6 Rho-0 cells were detached using trypsin and centrifuged at 300xg for 10 minutes. The resulting pellet was cultured for 21 days in hMSC Commercial Chondrogenic Differentiation Medium (Lonza, Biowhittaker) supplemented with 10 ng/ml transforming growth factor-β3 (TGFβ-3) (Prospec, Ness-Ziona, Israel). During this process the culture medium was changed every 3 days.
Chondrogenesis was assessed using the micropellet formation (2.5x104) technique [34 (link)], with some modifications. 3a6 and 3a6 Rho-0 cells were detached using trypsin and centrifuged at 300xg for 10 minutes. The resulting pellet was cultured for 21 days in hMSC Commercial Chondrogenic Differentiation Medium (Lonza, Biowhittaker) supplemented with 10 ng/ml transforming growth factor-β3 (TGFβ-3) (Prospec, Ness-Ziona, Israel). During this process the culture medium was changed every 3 days.
3
Adipogenic and Osteogenic Potential of MSCs
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The potential of AD- and BM-MSCs to differentiate into adipogenic and osteogenic lineages were examined. To induce adipogenic differentiation, cells were treated with an adipogenic differentiation medium (Lonza, Basel, Switzerland) for 3 weeks. Adipogenesis was assessed by Oil Red O staining (Supplementary Figure S1B ). For osteogenic differentiation, cells were treated with osteogenic differentiation medium (Lonza, Basel, Switzerland) for 3 weeks. Osteogenesis was assessed by Alizarin Red S staining (Supplementary Figure S1C ). Medium changes were performed twice weekly for the two assays.
4
Thermoresponsive Poloxamer 407 for Mesenchymal Stem Cell Culture
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Thermoresponsive poloxamer 407 (P407; non-ionic, ≤1% water content, pH 7.5) was procured from Sigma-Aldrich, St. Louis, USA. A PASCO® 850 Universal Interface (standard waveforms, frequency range 0.001–100 Hz, amplitude range ± 15 V, and output current range 61 μA at 10 V) with two Helmholtz coils (200 turns; inner radius 10.06 cm, outer radius 10.58 cm. Coil width 1.6 cm, and maximum current change 2 A) was obtained from PASCO Scientific, California, USA. Dulbecco's modified eagle media (DMEM), αMEM, fetal bovine serum (FBS), phosphate-buffered saline (PBS), and 1% P/S were procured from Welgene, Republic of Korea. Mesenchymal stem cell identification kit, live/dead double staining kit, ascorbic acid-2-phosphate, glutamine, alizarin Red-S (ARS), bovine serum albumin (BSA), plasminogen activator inhibitor-1 (PAI-1), and F-actin probe were procured from Sigma-Aldrich, St. Louis, USA. The osteogenic differentiation medium was obtained from Lonza, Maryland, USA. Primary and secondary antibodies against CD34, CD13, CD90, and CD146 were procured from BD Biosciences, San Jose, California, USA. cDNA synthesis kit and SYBR green master mix reagent were procured from Invitrogen, Maryland, USA, and Bio-Rad Laboratories, USA. A fully automated CELLINK® BIO-X 3D printer was procured from CELLINK® Corporation, Sweden.
5
Multilineage Differentiation of hAFSCs
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To investigate the differentiation ability, hAFSCs were differentiated in vitro into osteogenic, adipogenic, and chondrogenic lineages. hAFSCs were independently cultured either in “adipogenic differentiation medium,” “osteogenic differentiation medium,” or “chondrogenic differentiation medium” (Lonza, Basel, Switzerland) at 37°C in 5% CO2 for the appropriate time according to the manufacturer's recommended protocol. Osteogenesis was assessed by Alizarin staining (Cosmo Bio Co., Ltd., Tokyo, Japan) of the calcified extracellular matrix deposition. Oil Red O staining was used to detect intracellular lipid droplet formation to evaluate adipogenesis. Chondrogenic differentiation was determined by Alcian blue staining.
6
Evaluating Osteogenic Potential of Scaffold Designs
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hGMSCs were seeded at 8 × 103 cells/cm2 in chemically defined MSC growth medium (MSCGM-CD) (control medium) (Lonza, Basel, Switzerland) and in osteogenic differentiation medium (Lonza) in the presence of all scaffold designs (Fig. 1a–e ). To evaluate the performance of different scaffold designs in terms of osteogenic differentiation, the expression of RUNX2 after 1 week of culture was performed by Western blotting analysis (Fig. 1g ).
7
Osteogenic Differentiation of h-MSCs
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105 h-MSCs were seeded in 12-well cell-culture plasticware and cultured in either (i) α-MEM supplemented with 10% FBS, (ii) medium containing α-MEM:h-iPSC CM (1:2), or (iii) osteogenic differentiation medium (Lonza, USA). At different days of culture, cell mRNA was extracted and analyzed using real-time quantitative RT-PCR targeting select human, osteogenic-differentiation markers. The presence of calcium-containing deposits in the extracellular matrix was assessed using Alizarin Red stain.
8
Osteogenic Differentiation of Human Mesenchymal Stem Cells
9
Osteogenic Differentiation of Adult MSCs
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Osteogenic differentiation of sorted adult MSC was induced as previously described9 (link),10 (link). Briefly, cell cultures at 80% confluence were incubated for three weeks in osteogenic differentiation medium (Lonza, Basel, Switzerland). Differentiation was confirmed by staining with Alizarin Red S (cat. no. 130-22-3; Sigma-Aldrich, Saint Louis, MI, USA).
10
Osteogenic Differentiation of hMSCs on Fibronectin-Coated Scaffolds
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