Assessment of Candida shehatae viability by flow cytometry and fluorescent probes

Quantification of different physiological states of Candida shehatae cells was performed by flow cytometry associated with two fluorescent probes. Propidium iodide and carboxyfluorescein diacetate acetoxymethyl ester fluorescent dyes were chosen based on data from the literature. A staining procedure, developed from the previous works was applied to the yeast. Then, the protocol was improved to fit with fermentation constraints such as no physiological interference between the staining procedure and the cells, shortest preparation time and small amounts of dyes. From this optimisation, propidium iodide was included in the sample at 8 mg/L whereas carboxyfluorescein was first diluted in Pluronic® agent and used at 3mg/L, samples were incubated for 10 min at 40°C. Repeatability and accuracy were evaluated to validate this flow cytometry procedure for viability determination.