Image-Based Cytometric Analysis of Fluorescent Viability and Vitality Staining Methods for Saccharomyces Cerevisiae

Leo L. Chan, Dan Driscoll, Stephanie Saldi, and Dmitry Kuksin

Saccharomyces cerevisiae has been an essential component for the production of beer for centuries. The viability and vitality of yeast during standard brewing process is especially important for proper cell growth,
consistent production of flavor, and optimal yield for fermentation. Viability refers to the ability of the yeast to live and continue dividing, while the vitality refers to the metabolic activity of the yeast. Yeast may be viable and dividing, while not vital and allowing for fermentation. Traditional method for yeast viability measurement depended on mainly manual counting of methylene blue stained yeast cells in a hemacytometer. However, this method can be time-consuming and has user-dependent variations. In the recent years, fluorescent viability and vitality stains have become widely used for flow and image-based cytometry methods. Specifically for image cytometry, it has been previously demonstrated for rapid yeast concentration and viability measurements. In this work, we demonstrate the capability of Cellometer Vision image cytometry for yeast viability and vitality measurement, validating the methods against methylene blue. Various fluorescent stains were employed for viability and vitality measurement, such as nucleic acid
stains (PI, EB, 7-AAD and DAPI), membrane potential, intracellular, and enzymatic stains (oxonol, MgANS and CFDA-AM), and dual-fluorescent stains (AO/PI and CFDA-AM/PI). In addition, we performed a timecourse study to compare viability and vitality of lager and ale yeast, in order to understand yeast physical and metabolic characteristics during a standard fermentation process.