FLOW-CYTOMETRIC DETERMINATION OF THE RESPIRATORY ACTIVITY IN GROWING SACCHAROMYCES-CEREVISIAE POPULATIONS

Flow cytometry allows one to measure relevant physical or chemical properties on a single cell, yielding the distribution of these properties in the cell population, Typically, flow cytometry has been used to determine DNA or protein distributions, but it could be extended to the determination of other relevant parameters, such as intracellular pH, membrane potential, intracellular Ca2+ concentration, mitochondrial activity, etc. In the present work, we used flow cytometry to determine the respiratory activity in intact Saccharomyces cerevisiae cells after staining with the cationic lipophilic dye rhodamine 123 (Rh123). We found a good correlation between the Rh123 fluorescence distribution in yeast populations and the degree of respiratory activity, which can be varied by changing the carbon source used for yeast growth. In addition, we developed a vital staining procedure which allows one to measure fast changes in the respiratory activity. We used this technique to follow the kinetics of glucose repression and to measure the apparent K, for the substrate. Our results demonstrate that flow cytometry is a fast and very sensitive method to evaluate the respiratory activity in yeast cells and is also suitable for the determination of rapid changes in yeast metabolism. Biotechnological implications of this study are also discussed.