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Applied and Environmental Microbiology, September 2009, p. 5615-5620, Vol. 75, No. 17
0099-2240/09/$08.00+0     doi:10.1128/AEM.00650-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Application of a Short Intracellular pH Method to Flow Cytometry for Determining Saccharomyces cerevisiae Vitality {triangledown}

Claudia Weigert,1 Fabian Steffler,1 Tomas Kurz,2 Thomas H. Shellhammer,3 and Frank-Jürgen Methner4*

Research Institute for Brewing and Malting Technology at Research and Teaching Institute for Brewing in Berlin (VLB), Seestrasse 13, 13353 Berlin, Germany,1 Baldung-Grien-Strasse 26, 75173 Pforzheim, Germany,2 Department of Food Science and Technology, Oregon State University, Corvallis, Oregon 97331-6602,3 Institute of Biotechnology, Technical University of Berlin, Seestrasse 13, 13353 Berlin, Germany4

Received 19 March 2009/ Accepted 15 June 2009

The measurement of yeast's intracellular pH (ICP) is a proven method for determining yeast vitality. Vitality describes the condition or health of viable cells as opposed to viability, which defines living versus dead cells. In contrast to fluorescence photometric measurements, which show only average ICP values of a population, flow cytometry allows the presentation of an ICP distribution. By examining six repeated propagations with three separate growth phases (lag, exponential, and stationary), the ICP method previously established for photometry was transferred successfully to flow cytometry by using the pH-dependent fluorescent probe 5,6-carboxyfluorescein. The correlation between the two methods was good (r2 = 0.898, n = 18). With both methods it is possible to track the course of growth phases. Although photometry did not yield significant differences between exponentially and stationary phases (P = 0.433), ICP via flow cytometry did (P = 0.012). Yeast in an exponential phase has a unimodal ICP distribution, reflective of a homogeneous population; however, yeast in a stationary phase displays a broader ICP distribution, and subpopulations could be defined by using the flow cytometry method. In conclusion, flow cytometry yielded specific evidence of the heterogeneity in vitality of a yeast population as measured via ICP. In contrast to photometry, flow cytometry increases information about the yeast population's vitality via a short measurement, which is suitable for routine analysis.

* Corresponding author. Mailing address: Technical University of Berlin, Institute of Biotechnology, Seestr. 13, 13353 Berlin, Germany. Phone: 49-30-31427504. Fax: 49-30-31427503. E-mail: brauwesen{at}tu-berlin.de

{triangledown} Published ahead of print on 6 July 2009.

Applied and Environmental Microbiology, September 2009, p. 5615-5620, Vol. 75, No. 17
0099-2240/09/$08.00+0     doi:10.1128/AEM.00650-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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