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Yeast Concentration and Viability using Image-Based Fluorescence Analysis

Yeasts are an economically important organism
used for ethanol production in the beverage and
alternative fuels industries as well as a leavening
agent in the baking industry. In addition, pathogenic
strains of yeasts are involved in both plant and animal
diseases. Concentration and viability determinations
are routinely performed for quality control purposes
in yeast production, fermentation processes, and
fungicides research to monitor proliferation of
pathogenic yeasts.

The most common method for determining yeast
cell number and viability is manual counting on a
standard microscope using a hemacytometer and
methylene blue. One advantage of this method is that
visual inspection of each sample allows the operator to
check for contamination, presence of interfering debris,
and obvious dilution errors. A major disadvantage is
that the manual method is laborious, error-prone and
the data acquired is not easily traceable. Although
the equipments used to perform manual counting are
relatively inexpensive, the cost of human labor and
counting errors can be high enough to render manual
counting less than practical in a production facility
where accuracy, consistency, and record-keeping are
highly desirable.

Here we describe an automated, image-based
cytometry method that is rapid, highly accurate, and
consistent for determining concentration and viability
of yeasts. This method utilizes the Cellometer® X2
instrument for image-acquisition and associated
software for image analysis and data management.
Viability can concurrently be determined by staining
with the fluorescent viability dye propidium iodide
(PI). The instrument acquires both bright field and
fluorescence images of a counting chamber loaded
with stained yeast samples. Total cell numbers are
counted by the software using the bright field images
while the viability of each counted cell is determined
by analysis of the fluorescence images of the same
fields of view. The performance of the Cellometer
X2 system with respect to accuracy and reliability
for determining cell concentration and viability is
presented here.

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