Metabolic Assays for Detection of Neutral Fat Stores.

Abstract

Lipid droplets (LDs) are ubiquitous intracellular structures whose formation, growth, and maintenance are highly regulated (Wang et al., 2013; Ranall et al., 2011; Goodman, 2009). Lipid metabolism and droplet dynamics are of considerable interest to agriculture, biofuel production, viral pathology, nutrition, and cancer biology (Walther and Farese, 2009; Liu et al., 2010). Accumulation of fatty acids and neutral lipids in nonadipose tissues is cytotoxic (Kourtidis et al., 2009). BODIPY 493/503 (4,4-Difluoro-1,3,5,7,8-Pentamethyl-4-Bora-3a,4a-Diaza-s-Indacene) is the standard dye to study LDs within adipocytes. BODIPY 493/503 contains a nonpolar structure that, upon binding to neutral lipid, emits a green fluorescence signal with a narrow wavelength range, making it an ideal fluorophore for multi-labeling experiments. The hydrophobic nature of the dye molecules promotes rapid entry into the nonpolar environment of LDs (Listenberge and Brown, 2007). Gocze and Freeman showed that the lipid fluorescent variability is significantly lower when using BODIPY493/503 compared to Nile Red, suggesting that it may be more specific for the LD (Gocze and Freeman, 1994). Here, we describe a BODIPY 493/503 assay for the detection of neural fat stores in cultured cells (Figure 1) (Wang et al., 2013).