Isolation of lipid-rich marine microalgae by flow cytometric screening with Nile Red staining

Microalgae provide an optimal lipid source for commercial applications. The development a rapid, easy, and efficient screening procedure to isolate lipid-rich microalgae is becoming an extraordinarily active field of study. An automated flow cytometric cell-sorting technique in combination with Nile Red (NR) staining was used to isolate lipid-rich microalgae strains from a field sample. Cell sorting was based on the two-dimensional distribution of chlorophyll autofluorescence against forward scatter and NR fluorescence at 576 nm against that at 620 nm, corresponding to the optimal emission wavelength of NR in neutral and in polar lipid fractions, respectively. Of the 24 strains isolated using this method, 13 marine microalgae strains were successfully grown in culture medium and characterized for biomass production and cellular content of neutral and polar lipids based on NR fluorescence after 12 days of cultivation. The production of neutral lipids was higher in two strains (strains 9 and 12) compared to the marine diatom Phaeodactylum tricornutum, which is well known for its high lipid productivity. Three strains (strains 7, 8, and 9) exhibited a higher productivity of polar lipids than observed for P. tricornutum. These results indicate that our proposed method, an automated isolation technique coupled with the use of a specific cell stain such as NR, is efficient for selecting and isolating lipid-rich microalgae strains from the natural environment.