For surface markers, five-passage BMSCs were used with flow cytometry to identify the mesenchymal markers, CD44 and CD90, and the hematopoietic cell surface marker, CD45. For multidirectional differentiation potential, osteogenic induction medium, adipogenic induction medium and chondrogenic induction medium (all from Cyagen, China) were used to replace the complete medium to induce differentiation. The process of inducing differentiation was performed as described in the manufacturer's manual.
Adipogenic induction medium
Manufactured by Cyagen
Sourced in United States, China
Adipogenic induction medium is a specialized cell culture medium designed to induce and promote the differentiation of cells into adipocytes, or fat cells. The core function of this medium is to provide the necessary nutrients, hormones, and other factors required for the adipogenic differentiation process.
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Lab products found in correlation
Osteogenic induction medium, by Cyagen (4 mentions)
Chondrogenic induction medium, by Cyagen (4 mentions)
Adipogenic maintenance medium, by Cyagen (2 mentions)
Alizarin red, by Cyagen (2 mentions)
Oil red o, by Cyagen (2 mentions)
Oil red o solution, by Cyagen (2 mentions)
Chondrogenic medium, by Cyagen (1 mentions)
Osteogenic inducing medium, by Cyagen (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Hematoxylin, by Merck Group (1 mentions)
Pancreatic enzyme, by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Oil red o, by Merck Group (1 mentions)
Eclipse ti, by Nikon (1 mentions)
Flow cytometry, by BD (1 mentions)
Oil red o solution, by Merck Group (1 mentions)
Chondrogenic differentiation medium, by Cyagen (1 mentions)
Alizarin red stain, by Cyagen (1 mentions)
Aspirin, by Merck Group (1 mentions)
19 protocols using adipogenic induction medium
1
Mesenchymal Stem Cell Characterization
2
Multilineage Differentiation of hADSCs
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Multilineage differentiation capacity of hADSCs was detected as requested by the International Society for Cellular Therapy. Briefly, cells were cultured in adipogenic induction medium (Cyagen Biosciences, USA) for designated time and were stained by Oil Red‐O staining. For osteogenic differentiation, cells were cultured in osteogenic inducing medium (Cyagen Biosciences, USA) for designated time and were stained by alizarin red S. Chondrogenic differentiation was performed using the micromass culture technique maintained in chondrogenic medium (Cyagen Biosciences, USA) for up to 5 weeks, and were stained by Alcian blue.
3
Chondrogenic, Osteogenic, Adipogenic Differentiation
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Fetal bovine serum (FBS), streptomycin, penicillin, 0.25% pancreatic enzyme and DMEM were obtained from Gibco, USA. Chondrogenic induction medium, osteogenic induction medium and adipogenic induction medium were obtained from Cyagen US Inc. Hematoxylin and BSA were obtained from Sigma-Aldrich, USA. Mitochondrial fusion promoter M1 was obtained from Selleckchem (Shanghai, China).
4
Multilineage Differentiation of Adipose-Derived Stem Cells
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ASCs were identified via multipotent differentiation potential and flow cytometry. For adipogenic induction, ASCs were cultured in adipogenic induction medium (Cyagen Biosciences, Santa Clara, CA, USA). The adipocytes were stained with Oil red O staining on day 14. To induce osteogenic differentiation, a specific SD rat osteogenic induction medium (Cyagen Biosciences) was used. The cells were stained with ARS on day 14. For chondrogenic differentiation, ASCs were cultured for 2 weeks in chondrogenic induction medium (Cyagen Biosciences). On day 14, the cells were stained with toluidine blue to detect the secretion of sulfated glycosaminoglycans. Additionally, all of the media were replaced every 3 days. In addition, the identification of ASCs was measured via flow cytometry. ASCs were detected with antibodies against CD34 (Invitrogen, Waltham, MA, USA), CD45 (eBioscience, San Diego, CA, USA), CD44 (Invitrogen) and CD90 (BioLegend, San Diego, CA, USA). Results were analyzed via Flowjo software.
5
Adipogenic Differentiation Induction
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When the cells were 70 %–80 % confluent, the complete medium was replaced with an adipogenic induction medium (Cyagen Bioscience Inc.). After incubation for 2 weeks, lipid droplets were stained by using Oil Red O.
6
Adipogenic and Osteogenic Differentiation of hBMSCs
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The ability of hBMSCs at passage 4 to differentiate into adipocytes and osteoblasts were examined. To induce adipogenic differentiation, hBMSCs are plated in 24-well tissue culture plates with 1 ml medium per well. When the cells reached 100% confluence, they were carefully aspirated off the medium from the well, and 1 ml of adipogenic induction medium (Cyagen Biosciences, Guang Zhou, CHINA) was added. After 3 days, it was replaced with adipogenic maintenance medium (Cyagen Biosciences, Guang Zhou, CHINA), and was repeated. After 3 complete cycles of induction/maintenance, the adipocyte-induced cells were identified by staining with oil red O (Cyagen Biosciences, Guang Zhou, CHINA). To induce osteogenic differentiation, hBMSCs were plated in 24-well tissue culture plates, and when the cells reached 100% confluence, they were cultured with a osteogenic induction medium (Cyagen Biosciences, Guang Zhou, CHINA) for 21 days.The osteoblast induced cells were identified by staining with alizarin red (Cyagen Biosciences, Guang Zhou, CHINA).
7
Adipogenic Differentiation of Mouse BMSCs
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BMSCs were cultured in adipogenic induction medium (Cyagen, USA) for twenty-eight days and stained with Oil Red O (Sigma-Aldrich, USA) according to the provided instructions to detect the adipogenic capacity of mouse BMSCs.
8
Adipogenic Differentiation of BMSCs
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To induce adipogenic differentiation, the BMSCs were seeded at 2×104 cells/cm2 on 35-mm plastic dishes and grown for 3 days in adipogenic induction medium (Cyagen Biosciences, Inc.) containing additional SD rat MSC adipogenic differentiation basal medium A, FBS, insulin, glutamine, rosiglitazone, dexamethasone, 3-isobutyl-1-methylxanthine and penicillin/streptomycin, followed by 1 day in adipogenic maintenance medium containing SD rat MSC adipogenic differentiation basal medium A, FBS, insulin, glutamine and penicillin/streptomycin (1 cycle). Both steps were repeated up to day 21 (indicated as the 5th cycle), when the cell culture was terminated for Oil Red O staining. UTP (125 µM) was added to the culture medium every 3 days. Briefly, the cells were washed with PBS and fixed with 4% paraformaldehyde, for 30 min at room temperature. After washing with PBS, the fixed cells were stained with Oil Red O in deionized water. After 20 min, the cells were washed with PBS twice and observed under a light microscope (Eclipse Ti; Nikon). Image-Pro Plus 5.0 was used to quantify the nodule areas.
9
Characterization and Differentiation of Adult BMSCs
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Adult BMSCs were purchased from Cyagen Biosciences (Guangzhou, China) and cultured in complete growth medium (DMEM/F-12 medium that was supplemented with 10% FBS and 1% P/S) at 37 °C in a 5% CO2 cell culture incubator, and the adherent cells were passaged after reaching 70–80% confluence. P4 cells were used for subsequent experiments.
For phenotypic characterization, BMSCs were stained with mouse anti-human polyclonal antibodies against CD14, CD19, CD45, CD90, CD105, and CD73 (BD Pharmingen, San Diego, CA, USA) and analyzed by flow cytometry (BD Biosciences, Franklin Lakes, NJ, USA). The data were analyzed using FlowJoTM V10 software.
For multidirectional differentiation potential analysis, osteogenic induction medium, adipogenic induction medium, and chondrogenic induction medium (all from Cyagen, Suzhou, China) were used to replace the complete medium and induce differentiation. The process of inducing differentiation was performed as described in the manufacturer’s instructions and then assessed by Alizarin red staining, Oil Red O staining, and Alimin blue staining.
For phenotypic characterization, BMSCs were stained with mouse anti-human polyclonal antibodies against CD14, CD19, CD45, CD90, CD105, and CD73 (BD Pharmingen, San Diego, CA, USA) and analyzed by flow cytometry (BD Biosciences, Franklin Lakes, NJ, USA). The data were analyzed using FlowJoTM V10 software.
For multidirectional differentiation potential analysis, osteogenic induction medium, adipogenic induction medium, and chondrogenic induction medium (all from Cyagen, Suzhou, China) were used to replace the complete medium and induce differentiation. The process of inducing differentiation was performed as described in the manufacturer’s instructions and then assessed by Alizarin red staining, Oil Red O staining, and Alimin blue staining.
10
Adipogenic Differentiation of BMSCs
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BMSCs were cultured in 60 mm culture dishes until 60%–70% confluency and further incubated in an adipogenic induction medium (Cyagen Biosciences Inc, China). Cells were maintained in a CO2 incubator (37°C, 5% CO2), and the adipogenic differentiation medium was changed every 3 days. After 28 days of culture, cells were fixed and stained with Oil Red O (Ye et al., 2021 (link)).
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