Diatoms of the genus Pseudo-nitzschia are potentially toxic microalgae, whose blooms can trigger amnesic shellfish poisoning. The purpose of this study was to test and adapt different probes and procedures in order to assess the physiological status of Pseudo-nitzschia multiseries at the cell level using flow cytometry. To perform these analyses, probes and procedures were first optimized for concentration and incubation time. The percentage of dead Pseudo-nitzschia cells, the metabolic activity of live cells and their intracellular lipid content were then measured following a complete growth cycle. In addition, chlorophyll autofluorescence and efficiency of photosynthesis (quantum yield) were monitored. The concentration and viability of bacteria present in the medium were also assessed. Domoic acid (DA) was quantified as well. Just before the exponential phase, cells exhibited high metabolic activity, but low DA content. DA content per cell became most abundant at the beginning of the exponential phase when lipid storage was high, which provided a metabolic energy source, and when they were surrounded by a high number of bacteria (high bacteria/P. multiseries ratio). These physiological measurements tended to decrease during exponential phase and until stationary phase, at which time P. multiseries cells did not contain any DA nor store any lipids, and started to die.