Ltured in medium supplemented with aHS, we observed that aHS provides a high proliferation rate compared with the FBS supplementation, which was attributed for the first time to higher c-MYC protein expression in aHS cultures. Additionally it was shown that hASCs maintain their phenotypic, functional and genetic stability without any evidence of cell transformation.Materials and methodsPooled allogeneic human serumaHS was obtained from the whole blood of distinct blood-group-typed donors, as previously described by our group [40]. Donors gave written informed consent according to the approval of this study by the Ethics Committee in Research of the Universidade Federal de Minas Gerais (n?ETIC 11668613.7.0000.5149), advisory board of National Health Council (CNS). No clinical or laboratory evidence of metabolic diseases, hepatitis, HIV, or other systemic complications was observed for the donors. Briefly, the blood was collected from each donor using vacutainer tubes (Biosciences, USA) and allowed to clot spontaneously at 4 . The serum was separated by centrifugation at 252 g at 4 for 15 minutes. Different blood types (A, B, AB and O) from 20 (five of each blood type) different donors were mixed n-Phenylpiperazine-1-carboxamide to produce thePaula et al. Stem Cell Research Therapy (2015) 6:Page 3 ofbatches of aHS. The aHS was inactivated at 56 for 30 minutes prior to use and kept frozen at -20 . The batches of aHS were used in the culture of all hASCs studied in this work.Culture mediaBasal medium consisted of Dulbecco's modified Eagle's medium-high glucose (Sigma-Aldrich, USA) supplemented with 5 mM sodium bicarbonate (Cin ica Qu ica Ltda, Brazil), penicillin (100 units/mL; Sigma-Aldrich), streptomycin (0.1 mg/mL; Sigma-Aldrich), amphotericyn B (0.25 g/mL; Sigma-Aldrich), gentamicin (60 mg/L; Schering-Plough, USA), and 10 of either PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/9638577 aHS or FBS (Cripion Biotecnologia Ltda, Brazil). Adipogenic medium consisted of basal medium with 0.5 mM isobutylmethylxanthine (Sigma-Aldrich), 200 M indomethacin (SigmaAldrich), 1 M dexamethasone (Ach? Brazil), and 10 M insulin (Eli Lilly and Company, USA). Osteogenic medium consisted of basal medium with 50 g/mL ascorbate-2phosphate (Ecibra, Brazil), 10 mM -glycerophosphate (Sigma-Aldrich), and 0.1 M dexamethasone (Ach?. Chondrogenic medium consisted of basal medium with 1 mM dexamethasone (Ach?, 125 g/mL bovine serum albumin (PAA, Austria), 1 mM pyruvate (Sigma-Aldrich), 200 U/mL insulin (Eli Lilly and Company), 3.25 g/mL transferrin (Wako, Brazil), PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/9544797 0.01 g/mL transforming 3-Fluoro-2-(trifluoromethyl)aniline growth factor-1 (Sigma-Aldrich), 5 mg/mL ascorbate-2-phosphate (Ecibra) and a reduced concentration of serum supplements - 1 aHS or 1 FBS [5].Isolation and culture of human adipose tissue-derived stem cellsin a humidified 5 CO2 atmosphere (Water Jacketed CO2 Incubator; Thermo Scientific, USA). The hASCs were maintained under two distinct culture conditions: basal medium with 10 aHS or 10 gahmj.2015.132 FBS. After 12 to 28 hours of incubation, the medium was changed and nonadherent cells were removed. The hASCs were maintained at subconfluent levels, with three weekly medium changes. When the cells reached 80 to 90 confluence, they were washed with PBS and treated with 0.05 trypsin-EDTA (Invitrogen, Canada) for 3 to 5 minutes to detach from the surface of the culture flask. The resulting suspension was replated in new cell culture flask. The hASCs were expanded this way until they were used in the assays. To perform the assays the cells were harvested with basal medium afte.