Nile Red: A Fluorophore Useful in Assessing the Relative Lipid Content of Single Cells

The energy crisis in 1973 brought home to many people the fact that the world’s supply of crude mineral oil was not unlimited. Since that time there have been various attempts to investigate alternate sources of energy and many of these have centered on the use of plants to convert solar radiation to a more conventional energy source. A particular example is the production of fuel-grade oil from microalgal biomass grown in brackish water ponds in the southwestern deserts of the USA. The algae used in this process must have a high lipid content as well as being able to grow at elevated temperatures and light levels. One of the objectives of our research was to design a method to select algal strains producing large quantities of neutral lipid. For the purposes of this study, neutral lipid was defined as the triacylglyceride and hydrocarbon fraction of the cell. Since neither of these groups of compounds have functional groups that are easy to assay chemically, bulk lipid is often weighed after extraction from tissues with non-polar solvents or solvent mixtures (e.g., Bligh and Dyer, 1959). The method is time consuming and requires a sample large enough to produce a weighing (for the extracted lipid) in the milligram range. Greenspan and Fowler (1985) drew attention to the fact that Nile Red was fluorescent in non-polar but not in polar environments and pointed out that the stain could serve as a fluorescent lipid probe (Greenspan et al., 1985). We have extended this idea into a semiquantitative method for the determination of neutral lipid in small populations of single cells.