Nile Red Staining of C. elegans: A Method to Visualize Lipid Droplets in Fixed Animals

- When visualizing lipid droplets, the intracellular storage organelles for neutral lipids, begin by adding a detergent solution, such as one containing Triton X-100, to the sample. This will permeabilize the worm's cells. Next, fix the sample in the appropriate concentration of isopropanol, for example 40%.

Then, add Nile Red to the sample and protect it from light, since the dye is light sensitive. Fixation allows the dye to access lipid droplets throughout the animal. Nile Red is a lipophilic dye whose fluorescence emission spectrum depends on the polarity of the local lipid environment.

When exposed to polar lipids, such as phospholipids, Nile Red's emission spectrum is in the higher, red wavelengths. When exposed to neutral lipids, such as triacylglycerols and sterol esters in the neutral core of lipid droplets, Nile Red's emission spectrum shifts to the lower, yellow-gold wavelengths.

Therefore, use the green channel of a fluorescent microscope, which will collect the yellow-gold wavelength emissions, to visualize Nile Red staining of lipid droplets throughout the animal. In the example protocol, we will see a Nile Red staining procedure and imaging of lipid droplets in fixed