title: Dissolved oxygen concentration in the medium during cell culture: Defects and improvements.
abstract: In vitro cell culture has provided a useful model to study the effects of oxygen on cellular behavior. However, it remains unknown whether the in vitro operations themselves affect the medium oxygen levels and the living states of cells. In addition, a prevailing controversy is whether reactive oxygen species (ROS) production is induced by continuous hypoxia or reoxygenation. In this study, we have measured the effects of different types of cell culture containers and the oxygen environment where medium replacement takes place on the actual oxygen tension in the medium. We found that the deviations of oxygen concentrations in the medium are much greater in 25-cm(2) flasks than in 24-well plates and 35-mm dishes. The dissolved oxygen concentrations in the medium were increased after medium replacement in normoxia, but remained unchanged in glove boxes in which the oxygen tension remained at a low level (11.4, 5.7, and 0.5% O2 ). We also found that medium replacement in normoxia increased the number of ROS-positive cells and reduced the cell viability; meanwhile, medium replacement in a glove box did not produce the above effects. Therefore, we conclude that the use of 25-cm(2) flasks should be avoided and demonstrate that continuous hypoxia does not produce ROS, whereas the reoxygenation that occurs during the harvesting of cells leads to ROS and induces cell death.
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.|Brain Research Center, College of Basic Medical Sciences, Third Military Medical University, Chongqing, 400038, China.
authors.forename: Kuan
authors.initials: K
authors.lastname: Zhang
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.
authors.forename: Tong
authors.initials: T
authors.lastname: Zhao
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Huang
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.
authors.forename: Yunlin
authors.initials: Y
authors.lastname: He
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.
authors.forename: Yanzhao
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.
authors.forename: Liying
authors.initials: L
authors.lastname: Wu
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.
authors.forename: Kuiwu
authors.initials: K
authors.lastname: Wu
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.|Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, Jiangsu, 226001, China.
authors.forename: Ming
authors.initials: M
authors.lastname: Fan
authors.affiliation: Department of Brain Protection and Plasticity, Institute of Basic Medical Sciences, Beijing, 100850, China.|Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, Jiangsu, 226001, China.
authors.forename: Lingling
authors.initials: L
authors.lastname: Zhu
mesh_terms: D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D015687:Cell Hypoxia; D002470:Cell Survival; D003470:Culture Media / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D003470:Culture Media; D017382:Reactive Oxygen Species; D010100:Oxygen
keywords: cell viability; in vitro culture; medium replacement; oxygen concentration; reactive oxygen species