A Simple Method for Labeling Human Embryonic Stem Cells Destined to Lose Undifferentiated Potency

Mitochondrial oxidative phosphorylation is a major source of cellular ATP. Its usage as an energy source varies, not only according to the extracellular environment, but also during development and differentiation, as indicated by the reported changes in the flux ratio of glycolysis to oxidative phosphorylation during embryonic stem (ES) cell differentiation. The fluorescent probe JC‐1 allows visualization of changes in the mitochondrial membrane potential produced by oxidative phosphorylation. Strong JC‐1 signals were localized in the differentiated cells located at the edge of H9 ES colonies that expressed vimentin, an early differentiation maker. The JC‐1 signals were further intensified when individual adjacent colonies were in contact with each other. Time‐lapse analyses revealed that JC‐1‐labeled H9 cells under an overconfluent condition were highly differentiated after subculture, suggesting that monitoring oxidative phosphorylation in live cells might facilitate the prediction of induced pluripotent stem cells, as well as ES cells, that are destined to lose their undifferentiated potency. Significance Skillful cell manipulation is a major factor in both maintaining and disrupting the undifferentiation potency of human embryonic stem (hES) cells. Staining with JC‐1, a mitochondrial membrane potential probe, is a simple monitoring method that can be used to predict embryonic stem cell quality under live conditions, which might help ensure the future use of hES and human induced pluripotent stem cells after subculture. Mitochondrial oxidative phosphorylation is a major source of cellular ATP. The fluorescent probe JC‐1 allows visualization of changes in the mitochondrial membrane potential produced by oxidative phosphorylation. JC‐1‐labeled H9 cells under an overconfluent condition were highly differentiated after subculture, suggesting that monitoring oxidative phosphorylation in live cells might facilitate the prediction of induced pluripotent stem cells and ES cells destined to lose their undifferentiated potency.